SKENARIO 1

.... lebih baik sakit gigi, dari pada sakit hati ...
Nn. Cantique, 18 tahun, datang ke poliklinik RSI Unisma dengan keluhan sulit membuka
mulut. Dari hasil anamnesa dan pemeriksaan fisik oleh dokter, diketahui kalau 2 hari
yang lalu pasien mulai merasa sakit pada gusi dan gigi belakang kanan bawah. Sewaktu
bercermin, pasien melihat gusi pada daerah gigi bungsu membengkak, kemerahan dan
terasa sakit. Pasien lalu berkumur-kumur dengan air rebusan daun sirih untuk
mengurangi keluhan tersebut. Keesokan harinya pasien merasa demam dan mulai agak
sulit membuka mulut.
1. Bagaimanakah mekanisme biomolekuler yang terjadi pada kasus diatas ?
2. mengapa pasien minum rebusan daun sirih ?

1. identifikasi kata sulit

a. Poliklinik
b. sulit membuka mulut
c. anamnesa
d. gusi
e. gigi bungsu
f. air rebusan daun sirih
g. demam

2. identifikasi permasalahan
1. mengapa pasien mengalami kesulitan membuka mulut ?
2. mengapa gusi pada daerah gigi yang paling belakang membengkak,
kemerahan dan terasa sakit ?
3. mengapa pasien mengalami demam ?
4. mengapa pasien minum air rebusan daun sirih ?

3. curah pendapat
Learn about :
a. pericoronitis caused by wisdom teeth eruption  penjelasan untuk no.1,
2,3
Wisdom Teeth Eruption Symptoms
Wisdom teeth, technically called as third molar, are the last teeth to erupt in the mouth.
They usually erupt when a person is between the age 17 and 22, which in ancient times
was considered as the age in which a person was suppose to gain wisdom. A normal
person has four wisdom teeth, two in the upper arch, and two in the lower. Generally,
most adults have four wisdom teeth, but in some cases they may be more or
less. Wisdom teeth cause a lot of problems when they erupt in mouth because they
usually get trapped in the gum and jaw bones. They may break some of way
through your gums causing the food to trap there, which in turn can cause
the gum tissues to grow over them, thus triggering a gum infection. This
creates pain and irritation in mouth.

Signs & Symptoms Of Wisdom Tooth

Chewing Problem : Impacted wisdom teeth may pierce into the cheek and causes mouth
ulcers. The constant pain in that area caused by these ulcers makes it difficult for one to
chew.
Infection: In dental terminology, a wisdom tooth that has failed to emerge completely
into its expected position is called 'impacted wisdom tooth’. Impacted wisdom teeth can
form a gum flap near the top of the partially exposed tooth. Bacteria grows from food
particles can infect it. This infection can cause pain and irritation.
Irritation: Sometimes, a wisdom tooth gets trapped due to the space insufficiency in the
mouth and that lead to irritation. You may not able to concentrate on anything because
of this pain. This can be a real problem when you are handling some important
assignment.
Pain: Wisdom teeth pain is the most noticed symptom amongst the infected wisdom
teeth symptoms. Impacted wisdom teeth, or a tooth that experiences trouble breaking
through the gum, can lead to red, swollen, and painful gums.
Fever: Pericoronitis can cause fever and fatigue too. It can also cause muscle spasms in
the jaw.
Facial Swelling: The 'impacted wisdom teeth' may have erupted at wrong angles or at
wrong positions. Sometimes, infection like pericoronitis causes this condition. It may
cause pain around throat also.
Pus : Formation of pus near the gum can be found in some cases. It happens because of
severe infection.
Difficulty In Swallowing : You may feel difficulty in opening the jaw wide and
swallowing. These two are very common wisdom tooth symptoms.
Bad Breath: An infected wisdom teeth can lead to bad breath.
Reddened Gum: It’s a basic symptom of any tooth disease. The gum around the wisdom
is generally seen reddened and inflamed due to the erupting wisdom teeth.
Headache : An impacted wisdom tooth can cause headache and constant jaw pain
Pericoronitis

Pericoronitis is the inflammation of the soft tissues of varying severity around an

erupting or partially erupted tooth with breach of the follicle. It results in
establishing a communication with the oral environment. Even though this can occur in
relation to any tooth mandibular third molar is more often involved. Statistically, this
condition seems to be the third odontogenic infection and one of the common causes for
the removal of the impacted 3rd molars erupt in the lateral direction, which is the non-
functional region of the dental arc with unfavorable drainage. It is termed as
pericementitis.

Pathogenesis of Pericoronitis

It is widely believed that factors that predispose to pericoronitis are emotional stress,
fatigue, upper respiratory tract infections, second trimester of pregnancy
and menstruation. Impinging maxillary molars aggravate this condition by
traumatizing the pericoronal soft tissues of the mandibular 3rd molars. Repeated
occlusal trauma leads to oedematous swelling there by disturbing the drainage of the
inflammatory exudates. Any of these conditions may upset the delicate balance by
lowering the general resistance of the patient. During the eruptive phase, significant
collagenase activity in the operculum is present. The pericoronal pocket with soft tissue
debris provides a favorable environment for the microorganisms. With the associated
osteitis distal bony pocket increases the infection. Depending on the local condition
virulence or the organisms and varying degree of resistance to infection, edema spreads
by the toxins. Pathology of pericoronitis is similar to the abscess formation or
cellulites. The resistance of the surrounding tissues like fascia, muscle, etc guides the
spread of infection.

Wisdom tooth infections / Pericoronitis

Some wisdom teeth have to be extracted because the tissues around them repeatedly
become infected. The term for this condition is "pericoronitis." Pericoronitis refers to a
bacterial infection located in the tissues that surround the crown portion of a partially
erupted tooth.

A "partially erupted tooth" refers to a tooth that has only part way penetrated through
the gums. This may be a transitory positioning because the tooth is in the process of
erupting or, in the case of a partially erupted impacted tooth, a permanent one.

The "crown" of a tooth is its non-root portion. That part that is typically positioned above
the gum line

Pericoronitis is caused by bacteria.

Pericoronitis is caused by bacteria (dental plaque) that cannot be adequately cleansed

from around the tooth. And as long as the tooth remains partially erupted and
impossible to properly clean (which might be forever in the case of an impacted wisdom
tooth), a person may experience repeated episodes of pericoronitis.
Why does pericoronitis form?

The crown portion of a tooth forms within a pouch termed the follicular sac (drawn in
yellow in our pictures).

At that point in time during its eruption when a tooth first penetrates through the gum
tissue, the integrity of this developmental sac is lost and bacterial from the mouth
easily enter into this space and find harbor.

Unfortunately, there is no way for a person to effectively clean away these bacteria. And
due to this inability, from time to time it is possible for them to cause an active infection,
which then spreads to adjacent tissues surrounding the wisdom tooth (as illustrated in
our picture below).

Where does the term "pericoronitis" come from?

The term "pericoronitis" breaks down as follows. "Peri-" means "around." The "-coron-"
portion of the term refers to the "crown" of the tooth. The suffix "-itis" refers to the
presence of infection. So, the word pericoronitis literally means "an infection around
the crown portion of a tooth." There can also be severe pain, an unpleasant mouth odor,
and even a bad taste coming from the infected area.

Pericoronitis Symptoms and Signs

The signs of pericoronitis are tenderness and swelling in the gum tissue surrounding the
wisdom tooth. This swelling can extend into the face and may be associated with a
limited ability to open one's mouth. In extreme cases, the swelling may become so
pronounce as to threaten the patency of the person's airway and their ability to breathe.

Based on patient’s history, it is convenient to group the sings and symptoms of

Pericoronitis as acute, sub acute and chronic phases.

Acute Phase Sub Acute Phase Chronic Phase

Severe throbbing pain Jaw stiffness Dull Pain

Restricted Mouth Opening Pus discharge Unpleasant taste
Acute phase: In the early stage, it is very similar to teething. Patient is aware of the
eruption of the tooth and discomfort which aggravates by traumatic occlusion at the
mandibular retro molar region. Patient experiences severe throbbing pain radiating to
the adjacent regions. Development of some degree of restricted mouth opening may be
due to the stimulation of pain receptors, extra oral swelling and dysphagia. On
examination, patient has pyrexia, increased pulse and respiration rate. Regional sub-
mandibular lymph nodes are tender. There may be Halitosis, leucocytosis and malaise.
Intra-orally, swelling and redness with purulent discharge from pericoronal space may
be noticed. Increase in edema may lead to rapid separation of muscles of mastication
and the subsequent trismus. Dysphagia indicates that infection has spread into
sublingual and Para-pharyngeal spaces. Sloughing or ulceration may be noticed around
the operculum.

Sub-acute phase: During this phase, the systemic features become less acute. The
patient experiences swelling, jaw stiffness regional lymphadenopathy and pus discharge.

Chronic phase: This phase is characterized by the complete absence of all the systemic
features except during acute exacerbation. Usually the patient complains of dull pain and
discomfort with unpleasant taste in the oral cavity. Intraoral periapical radiograph may
reveal a craterlike bony defect around the third molar.

Diagnosis of Pericoronitis Infection

Pericoronitis diagnosis does not present any problem with proper history and careful
clinical examination. It is advantageous to record patient’s body temperature, pulse and
respiration rate. Special investigations include radiographic examination, total and
differential counts of leucocytes. If facilities exist, bacteriological examinations done to
determine the nature of causative organisms so that it is helpful to confirm the
sensitivity to various antibiotics. Whenever a fungal infection is considered, a special
request must be made to the microbiologist, since it requires special anaerobic methods
of culture.

b. air rebusan daun sirih  baca jurnal “ Piper betle Linn. a maligned Pan-Asiatic plant
with an array of pharmacological activities and prospects for drug discovery ”
4. Mapping
Mapping Konsep

SEL

STRUKTUR FUNGSI
-Membran
-Organela
-Sitoskeleton

Reproduksi Respirasi & Komunikasi Ekskresi & Gerakan

Mitosis Metabolisme - Endokrin Sekresi Amoeboid
Meiosis OxFos - Parakrin -Endositosis Diapedesi
(Nukleus) Siklus - Autokrin -Eksositosis (Sitoskeleton)
Krebs - Sinaps (Membran)
Sintesa - Junction (Aparatus G)
(Tight, Gap Jun) (Ribosom)
Glikolisis
(Mitokondria) (Membran Sel)
(Sitoplasma)
(Ribosom) Regulasi
Transport

Co Transport
(Transport aktif 2 nd )

Active
Facilitated Transport
Passive Diffusion - Perlu Protein spesifik
Diffusion - Melawan gradien
- Digerakan perpindahan
- Perlu Protein spesifik ion ko transport
- Melawan gradien menuruni gradien
- Perlu Protein spesifik - Bahan yg ditransport
Bahan yg ditransport : - Perlu ATP
- Bahan yg ditransport Glukosa & Asam
O2, CO2, Hormon - Bahan yg ditransport
Glukosa, Asam Amino (Symporters)
Steroid , Obat Ion, molekul hidrofil,
Amino (uniporter), ion, sukrosa (anti
Lipid
ion, air (channel). porters)
Mapping Konsep Keradangan

Fisik
Radiasi, Iritan
Benda Tajam, Suhu

Kimiawi
Mikroorganisme INJURY SEL Toksin
Alergen i Sel Free Radikal

KERUSAKAN SEL / JARINGAN

AKTIVASI Sekresi IL-10

MAKROFAGE

Aktivasi sistim imun tubuh dan sel yg

terkena jejas, PD, jar interstitiel

Pembuluh Sel Matriks

Darah & Extrasel
Komponennya

INFLAMASI
AKUT
PELEPASAN MEDIATOR INFLAMASI Tumor
antara lain : PG, Bradikinin, Histamin, Leukotrien Kalor
Rubor
Dolor
Gagal
WOUND Berhasil Gagal
INFLAMASI KRONIS NEKROSIS
HEALING/
APOPTOSIS
SEMBUH
Mapping Kasus
Nona Cantique Usia 17-25 tahun

Geraham bungsu erupsi (M3 RB)

Erupsi optimal posisi Erupsi optimal tapi posisi Erupsi tidak optimal
baik buruk
Aktivitas kolagenase
meningkat Terhalang corpus mandibulae Space sempit

Gusi “robek”  flap Tidak mencapai posisi anatomis

Terbetuk celah antara gusi dan gigi

Iritan bagi mukosa
Gusi yang menutupinya tergigit Makanan mudah terjebak, sulit
yang berhadapan
oleh gigi diatasnya dijangkau oleh air dan sikat gigi shg
dengan bagian oklusal
sulit dibersihkan
gigi

jejas bagi gusi atau mukosa pipi Lingkungan kondusif bagi opportunistic infection

merusak membran sel epitel / fibrosit Toksin bakteri

Aktivasi metabolisme
asam arachidonat

Aktivasi mediator
inflamasi yang lain

PROSTAGLANDINE A group of lipids TROMBOXANE : agregasi tromosit LEUKOTRIENE

which can cause vasodilation, fever, and Able to mediate leukocyte adhesion and
pain activation, allowing them to bind to the
Melambatkan aliran darah  endothelium and migrate across it. In
hemostasis neutrophils, it is also a potent
Volume darah Cairan jaringan chemoattractant, and is able to induce
meningkat meningkat the formation of reactive oxygen species
and the release of lysosome enzymes by
these cells.
Gusi Edema menyebar
bengkak dan musculus
kemerahan massetericus Aktivitas anti infeksi

Otot
Menekan pengunyahan
nerve ending kaku
 Sulit membuka Sulit menyikat gigi Berkumur rebusan daun sirih
nyeri mulut
hipersalivasi Berkumur rebusan daun sirih Aktivasi fitokimia sirih

Saliva mengandung Allylpyrocatechol (APC) Hydroxychavicol (HC)/

Hydroxychavicol acetate
komponen imun  (HCA)
lisosom Targets the inflammatory
Peroksisome response of macrophages via
inhibition of iNOS, COX-2 and Probably works through the
sIgA IL-12 p40 through down- disruption of the permeability
regulation of the NF-kappaB barrier of microbial membrane
pathway, indicating that APC structures
may have therapeutic potential
in inflammation associated
disorders Oral hygiene/antibacterial/
antifungal for :
Staphylococcus aureus
Staphylococcus mutans
Staphylococcus faecalis
Candida albicans
Vibrio cholerae ogawa
Diplococcus pneumoniae and
Klebsiella aerogenes

Upaya penangan terhadap keradangan lokal, sementara sitokin yang berperan sbg pirogen endogen mulai teraktivasi

demam

5. Self Directed Learning

6. Learning Objective
Mengetahui dan memahami :
1. sel : definisi, macam, perbedaan sel prokariot-eukariot dan macam2 sel dalam
tubuh manusia dan fungsinya
2. anatomi, fisiologi dan sistim biokimia sel tubuh
3. agen-agen pertahanan sel : mediator anti inflamasi dan anti oksidan
4. agen2 yang melemahkan sel : jejas fisik, kimia dan bakteri-toksin, radiasi
5. mekanisme pertahanan sel melalui keradangan (inflamasi) : definisi, jenis, proses,
faktor2 pemicu, peran/kegunaan, mekanisme trjadinya tanda2 radang , upaya
pencegahan, penghambatan dan penanganan
 6. patomekanisme kasus
7. terapi herbal untuk keradangan (penanganan pada kasus)

7. Reporting
1. sel : definisi, macam, perbedaan sel prokariot-eukariot dan macam2 sel
dalam tubuh manusia dan fungsinya
Sel adalah segumpal protoplasma yang berinti, sebagai individu yang berfungsi
menyelenggarakan seluruh aktivitas untuk kebutuhan hidupnya. Sel itu setelah tumbuh
dan berdeferensiasi, akan berubah bentuknya sesuai dengan fungsinya, ada yang
menjadi epidermis berfungsi untuk melindungi sel-sel sebelah dalamnya ada yang
menjadi tempat penyediaan makanan, ada yang berfungsi menjadi tempat persediaan
makanan dan lain-lain . Atau dengan kata lain juga sel merupakan unit struktural
kehidupan dan merupakan unit fungsional dari kehidupan dikarenakan didalam organ
tumbuhan dan hewan tersusun dari sel-sel rumusan yang penting bukannya dinding sel
tetapi isi sel yang disebut protoplasma

Ada tiga keistimewaan yang khas pada sel tumbuhan : dinding sel dengan selulosa,
vakuola (yang memberi tekanan dan memperbesar volume serta luas permukaan
meskipun dengan protoplasma sedikit), dan plastida, khususnya kloroplas. Vakuola
dapat ditemui pada anggota kelima dunia, namun vakuola besar di pusat sel ada pada
hampir semua sel tumbuhan, cendawan, dan beberapa protista. Kloroplas hanya
terdapat pada tumbuhan dan beberapa protista (bergantung pada golongannya). Sel
sendiri sebagai dasar menyusun suatu organisme yang terdiri dari inti (nukleus) yang
terbungkus oleh membran atau struktur serupa tanpa membran. Tidak ada kehidupan
dalam satuan yang lebih kecil dari pada sel. Sel terbentuk hanya dengan pembelahan sel-
sel sebelumnya. Sel dicirikan oleh adanya molekul makro khusus, seperti pati dan
selulosa, yang terjadi dari ratusan sampai ribuan gula atau molekul lain selain itu sel
juga dapat dicirikan oleh adanya molekul makro seperti protein dan asam nukleat baik
DNA atau RNA yang tersusun sebagai rantai yang terdiri dari ratusan sampai ribuan
molekul. Pada tumbuhan istilah sel meliputi protoplasma dan dinding sel yang ada
sedangkan pada organisme multi sel yang ada membentuk struktur kompleks yaitu
jaringan dan organ. Sel pada organisme multi sel tidak sama satu dengan lainnya tetapi
masing-masing mempunyai struktur dan fungsi yang berbeda. Pada awalnya struktur
dinding sel yang ada pada tumbuhan merupakan sebagai sel mati hasil ekskresi zat
hidup dalam sel akan tetapi baru-baru ini makin banyak ditemui bukti bahwa ada satuan
organik yang ada diantara protoplasma dan dinding, khususnya pada sel muda.

Meskipun antara sel hewan dan sel tumbuhan berbeda namun terdapat persamaan-
persamaan dasar tertentu mengenai sifat, bentuk, dan fungsi dari bagian sel tersebut.
Secara umum bagian-bagian sel tersebut adalah membran sel, sitoplasma, mitokondria,
retikulum endoplasma, aparatus golgi, lisosom, plastida, kloroplas, sentrosom, ribosom,
vakuola, inti sel, membran inti, mikrofilamen, dan dinding sel . Sel tumbuhan
mempunyai bentuk yang bermacam-macam. Ada yang berbentuk peluru, prisma, dan
memanjang seperti rambut atau seperti ular.

Sel tumbuhan mempunyai dua bagian pokok yang berbeda dari hewan yaitu vakuola,
plastida dan dinding sel. Vakuola dan plastida merupakan bagian hidup dari sel
tumbuhan dan disebut protoplas. Sedangkan dinding sel yang berfungsi untuk
melindungi isi sel atau lumen yang ada di protoplasma disebut bagian sel yang mati. Hal
ini terlihat pada sel gabus tumbuhan yang tergolong sel mati karena hanya memiliki inti
sel dan sitoplasma, sehingga ruang antar selnya kosong. Bentuk sel gabus heksagonal,
tersusun rapat antara satu dan lainnya. Meskipun antara sel hewan dan sel tumbuhan
berbeda namun terdapat persamaan-persamaan dasar tertentu mengenai sifat, bentuk,
dan fungsi dari bagian sel tersebut. Secara umum bagian-bagian sel tersebut adalah
membran sel, sitoplasma, mitokondria, retikulum endoplasma, aparatus golgi, lisosom,
plastida, kloroplas, sentrosom, ribosom, vakuola, inti sel, membran inti, mikrofilamen,
dan dinding sel. Sel tumbuhan memiliki sifat yang khas yaitu adanya dinding sel yang
bukan komponen protoplasma.Penelitian terhadap dinding sel yang dipadu dengan
berbagai penemuanyang serba canggih pada saat sekarang telah dapat mengembangkan
cara-carapengungkapannya. Dinding sel mempunyai struktur yang komplek tetapi tiga
bagian fundamentalnya dapat dibedakan yaitu lamela tengah,dinding sel primer dan
dinding sel sekunder.Dinding sel terbuat dari zat selulosa,dinding sel relatif tebal. Ruang
sel adalah tempat organel-organel yang lain yang berada didalam sel. Ruang sel ini
meliputi bagian-bagian dalam sel yang mencakupnya protoplasma atau cairan sel.
Sedangkan ruang antar sel adalah penghubung antar sel yang satu dengan yang lainnya.

Secara umum setiap sel memiliki

a. Membran sel

b. Sitoplasma,dan

c. Inti sel atau nukleus.

Sel tumbuhan dan sel hewan mempunyai beberapa perbedaan seperti berikut:

 Sel tumbuhan lebih besar daripada sel hewan

 Sel tumbuhan mempunyai bentuk yang tetap sebaliknya pada sel hewan tidak
mempunyai bentuk yang tetap.
 Pada sel tumbuhan terdapat dinding sel sedangkan pada sel hewan tidak
 Sel tumbuhan terdapat klorofil namun pada sel hewan tidak
 Pada sel tumbuhan terdapat vakuola atau rongga sel yang besar sedangkan pada
sel hewan tidak memiliki vakuola, namun pada sel beberapa hewan uniseluler
memiliki vakuola tapi tidak sebesar yang terdapat pada sel tumbuhan
 Sel tumbuhan menyimpan tenaga dalam bentuk biji kanji sedangkan pada hewan
dalam bentuk glikogen.

Adapun perbedaan organel-organel pada sel tumbuhan dan hewan adalah sebagai
berikut
1. DINDING SEL

a. Hanya ada pada tumbuhan

b. Tersusun atas :

 Yang utama zat kayu yaitu selulosa yang terbuat dari glukosa
 Pectin terbuat dari polisakarida
 Hemiselulosaterbuat dari polisakarida
 Glikoprotein

c. Dibentuk oleh diktiosom

d. Berperan dalam turgiditas sel (kekakuan sel)
e. Pada dinding sel terdapat noktah (pori) yaitu : bagian dinding sel yang tidak
mengalami penebalan untuk berhubungan dengan sel tetangga
f. Pada noktah tersebut terdapat plasmodesmata
g. Terdapat 2 jenis dinding sel :

1. Dinding sel primer

Pada sel-sel muda yang sedang tumbuh, sel parenkim dan sel kolenkim Tersusun atas:
 9-25 % selulosa
 hemiselulosa
 pectin
 beberapa senyawa

2. Dinding sel sekunder

Pada sel dewasa yang dibentuk disebelah dalam dinding primer Tersusun atas :
1. Kandungan selulosa lebih banyak (41-45)%

2. Hemiselulosa

3. Lignin

Beberapa sel mengalami penambahan lignin yang keras dan kaku

2. PLASTIDA

1. Hanya pada tumbuhan

2. Bermembran rangkap

 Membrane luar „³ untuk melewatkan molekul berukuran kecil

 Membrane dalam bersifat selektif permeable, memilih molekul yang keluar
masuk dengan transport aktif

3. Organel yang mengandung pigmen

Jenis-jenisnya adalah :

1. Kloroplas

 Plastida yang mengandung klorofil

 Hanya dijumpai pada sel autotrof eukariotik
 Membrane dalam membentuk tilakoid (tempat terjadinya fotosintesis) dan
Membungkus cairan kloroplas
 Stroma untuk penyimpanan hasil fotosintesis
 Tilakoid bertumpuk disebut grana

2. Kromoplas

Bertugas menyintesis dan menyimpan pigmen merah, jingga, atau kuning Misal :
 pada tomat dan wortel

3. Leukoplas

 Tidak mengandung pigmen warna

 Terdapat pada jaringan yang tidak terkena cahaya
 Yaitu terdapat pada sel-sel embrional, empulur batang, bagian tumbuhan di
dalam tanah yang berwarna putih

4. Amiloplas

 Tidak mengandung pigmen

 Berfungsi dalam penyimpanan amilum (pati) Misal pada umbi

3. VAKUOLA
1. Bermembran
2. Berisi cairan vakuola
3. Pada hewan vesikula
4. Pada tumbuhan :
 Tumbuhan muda vakuola kecil
 Tumbuhan dewasa vakuola besar-besar

5. Berhubungan dengan tekanan turgor

6. Tekanan turgor berguna untuk mengatur gerakan osmosis cairan dari luar ke dalam
sel

Vakuola berisi:

 Senyawa cadangan makanan (as.amino, gula, beberapa asam.organik dan

protein)
 Metabolit sekunder (senyawa yang tidak diperlukan oleh sel itu sendiri)

4. SENTRIOL
1. Hanya terdapat pada sel hewan
2. Sepasang structur seperti silinder yang memiliki lubang tengah
3. Tersusun dari protein mikrotubulus
4. Pasangan sentriol terletak menyudut
5. Tampak seperti jala berlekatan dengan kromosom saat sel membelah
6. Jala tersebut disebut benang spindle fungsi :
 Mengatur polaritas pembelahan sel hewan
 Mengatur pemisahan kromosom selama pembelahan sel

Dapat kami simpulkan bahwa bentuk dan susunan sel yang terdapat pada preparat
berbeda-beda, namun secara mendasar perbedaan tersebut dapat kita lihat antara sel
hewan dan sel tumbuhan yang dimana antara keduanya memiliki perbedaan yang sangat
mencolok
JENIS SEL TUBUH MANUSIA
 1 Cells that are derived primarily from endoderm
o 1.1 Gland cells
 1.1.1 Exocrine secretory epithelial cells
o 1.2 Hormone secreting cells
 2 Epithelial cells lining closed internal body cavities
o 2.1 Ciliated cells with propulsive function
 3 Derived primarily from ectoderm
o 3.1 Integumentary system
 3.1.1 Keratinizing epithelial cells
 3.1.2 Wet stratified barrier epithelial cells
o 3.2 Nervous system
 3.2.1 Sensory transducer cells
 3.2.2 Autonomic neuron cells
 3.2.3 Sense organ and peripheral neuron supporting cells
 3.2.4 Central nervous system neurons and glial cells
 3.2.5 Lens cells
 4 Derived primarily from mesoderm
o 4.1 Metabolism and storage cells
o 4.2 Barrier function cells (Lung, Gut, Exocrine Glands and Urogenital
Tract)
 4.2.1 Kidney
o 4.3 Extracellular Matrix Cells
o 4.4 Contractile cells
o 4.5 Blood and immune system cells
o 4.6 Pigment cells
o 4.7 Germ cells
o 4.8 Nurse cells

o 4.9 Interstitial cells

Cells that are derived primarily from endoderm

Gland cells

Exocrine secretory epithelial cells

 Salivary gland mucous cell (polysaccharide-rich secretion)
 Salivary gland serous cell (glycoprotein enzyme-rich secretion)
 Von Ebner's gland cell in tongue (washes taste buds)
 Mammary gland cell (milk secretion)
 Lacrimal gland cell (tear secretion)
 Ceruminous gland cell in ear (ear wax secretion)
 Eccrine sweat gland dark cell (glycoprotein secretion)
 Eccrine sweat gland clear cell (small molecule secretion)
 Apocrine sweat gland cell (odoriferous secretion, sex-hormone sensitive)
 Gland of Moll cell in eyelid (specialized sweat gland)
 Sebaceous gland cell (lipid-rich sebum secretion)
 Bowman's gland cell in nose (washes olfactory epithelium)
 Brunner's gland cell in duodenum (enzymes and alkaline mucus)
  Seminal vesicle cell (secretes seminal fluid components, including fructose for
swimming sperm)
 Prostate gland cell (secretes seminal fluid components)
 Bulbourethral gland cell (mucus secretion)
 Bartholin's gland cell (vaginal lubricant section)
 Gland of Littre cell (mucus secretion)
 Uterus endometrium cell (carbohydrate secretion)
 Isolated goblet cell of respiratory and digestive tracts (mucus secretion)
 Stomach lining mucous cell (mucus secretion)
 Gastric gland zymogenic cell (pepsinogen secretion)
 Gastric gland oxyntic cell (hydrochloric acid secretion)
 Pancreatic acinar cell (bicarbonate and digestive enzyme secretion
 Paneth cell of small intestine (lysozyme secretion)
 Type II pneumocyte of lung (surfactant secretion)
 Clara cell of lung

Hormone secreting cells

 Anterior pituitary cells
o Somatotropes
o Lactotropes
o Thyrotropes
o Gonadotropes
o Corticotropes
 Intermediate pituitary cell, secreting melanocyte-stimulating hormone
 Magnocellular neurosecretory cells
o secreting oxytocin
o secreting vasopressin
 Gut and respiratory tract cells
o secreting serotonin
o secreting endorphin
o secreting somatostatin
o secreting gastrin
o secreting secretin
o secreting cholecystokinin
o secreting insulin
o secreting glucagon
o secreting bombesin
 Thyroid gland cells
o thyroid epithelial cell
o parafollicular cell
 Parathyroid gland cells
o Parathyroid chief cell
o Oxyphil cell
 Adrenal gland cells
o chromaffin cells
o secreting steroid hormones (mineralcorticoids and gluco corticoids)
 Leydig cell of testes secreting testosterone
 Theca interna cell of ovarian follicle secreting estrogen
 Corpus luteum cell of ruptured ovarian follicle secreting progesterone
o Granulosa lutein cells
 o Theca lutein cells
 Juxtaglomerular cell (renin secretion)
 Macula densa cell of kidney
 Peripolar cell of kidney
 Mesangial cell of kidney

Epithelial cells lining closed internal body cavities

 Blood vessel and lymphatic vascular endothelial fenestrated cell
 Blood vessel and lymphatic vascular endothelial continuous cell
 Blood vessel and lymphatic vascular endothelial splenic cell
 Synovial cell (lining joint cavities, hyaluronic acid secretion)
 Serosal cell (lining peritoneal, pleural, and pericardial cavities)
 Squamous cell (lining perilymphatic space of ear)
 Squamous cell (lining endolymphatic space of ear)
 Columnar cell of endolymphatic sac with microvilli (lining endolymphatic space
of ear)
 Columnar cell of endolymphatic sac without microvilli (lining endolymphatic
space of ear)
 Dark cell (lining endolymphatic space of ear)
 Vestibular membrane cell (lining endolymphatic space of ear)
 Stria vascularis basal cell (lining endolymphatic space of ear)
 Stria vascularis marginal cell (lining endolymphatic space of ear)
 Cell of Claudius (lining endolymphatic space of ear)
 Cell of Boettcher (lining endolymphatic space of ear)
 Choroid plexus cell (cerebrospinal fluid secretion)
 Pia-arachnoid squamous cell
 Pigmented ciliary epithelium cell of eye
 Nonpigmented ciliary epithelium cell of eye
 Corneal endothelial cell
 Peg cell (of Fallopian tube)

Ciliated cells with propulsive function

 Respiratory tract ciliated cell
 Oviduct ciliated cell (in female)
 Uterine endometrial ciliated cell (in female)
 Rete testis ciliated cell (in male)
 Ductulus efferens ciliated cell (in male)
 Ciliated ependymal cell of central nervous system (lining brain cavities)

Derived primarily from ectoderm

Integumentary system

Keratinizing epithelial cells

 Epidermal keratinocyte (differentiating epidermal cell)
 Epidermal basal cell (stem cell)
 Keratinocyte of fingernails and toenails
 Nail bed basal cell (stem cell)
 Medullary hair shaft cell
 Cortical hair shaft cell
  Cuticular hair shaft cell
 Cuticular hair root sheath cell
 Hair root sheath cell of Huxley's layer
 Hair root sheath cell of Henle's layer
 External hair root sheath cell
 Hair matrix cell (stem cell)

Wet stratified barrier epithelial cells

 Surface epithelial cell of stratified squamous epithelium of cornea, tongue, oral
cavity, esophagus, anal canal, distal urethra and vagina
 basal cell (stem cell) of epithelia of cornea, tongue, oral cavity, esophagus, anal
canal, distal urethra and vagina
 Urinary epithelium cell (lining urinary bladder and urinary ducts)

Nervous system

Sensory transducer cells

 Auditory inner hair cell of organ of Corti
 Auditory outer hair cell of organ of Corti
 Basal cell of olfactory epithelium (stem cell for olfactory neurons)
 Cold-sensitive primary sensory neurons
 Heat-sensitive primary sensory neurons
 Merkel cell of epidermis (touch sensor)
 Olfactory receptor neuron
 Pain-sensitive primary sensory neurons (various types)
 Photoreceptor cells of retina in eye:
o Photoreceptor rod cells
o Photoreceptor blue-sensitive cone cell of eye
o Photoreceptor green-sensitive cone cell of eye
o Photoreceptor red-sensitive cone cell of eye
 Proprioceptive primary sensory neurons (various types)
 Touch-sensitive primary sensory neurons (various types)
 Type I carotid body cell (blood pH sensor)
 Type II carotid body cell (blood pH sensor)
 Type I hair cell of vestibular apparatus of ear (acceleration and gravity)
 Type II hair cell of vestibular apparatus of ear (acceleration and gravity)
 Type I taste bud cell

Autonomic neuron cells

 Cholinergic neural cell (various types)
 Adrenergic neural cell (various types)
 Peptidergic neural cell (various types)

Sense organ and peripheral neuron supporting cells

 Inner pillar cell of organ of Corti
 Outer pillar cell of organ of Corti
 Inner phalangeal cell of organ of Corti
 Outer phalangeal cell of organ of Corti
 Border cell of organ of Corti
 Hensen cell of organ of Corti
  Vestibular apparatus supporting cell
 Taste bud supporting cell
 Olfactory epithelium supporting cell
 Schwann cell
 Satellite cell (encapsulating peripheral nerve cell bodies)
 Enteric glial cell

Central nervous system neurons and glial cells

 Astrocyte (various types)
 Neuron cells (large variety of types, still poorly classified)
 Oligodendrocyte
 Spindle neuron

Lens cells
 Anterior lens epithelial cell
 Crystallin-containing lens fiber cell

Derived primarily from mesoderm

Metabolism and storage cells

 Hepatocyte (liver cell)
 Adipocytes:
o White fat cell
o Brown fat cell
 Liver lipocyte

Barrier function cells (Lung, Gut, Exocrine Glands and Urogenital Tract)

Kidney
 Kidney glomerulus parietal cell
 Kidney glomerulus podocyte
 Kidney proximal tubule brush border cell
 Loop of Henle thin segment cell
 Kidney distal tubule cell
 Kidney collecting duct cell
 Type I pneumocyte (lining air space of lung cell)
 Pancreatic duct cell (centroacinar cell)
 Nonstriated duct cell (of sweat gland, salivary gland, mammary gland, etc.)
o principal cell
o Intercalated cell
 Duct cell (of seminal vesicle, prostate gland, etc.)
 Intestinal brush border cell (with microvilli)
 Exocrine gland striated duct cell
 Gall bladder epithelial cell
 Ductulus efferens nonciliated cell
 Epididymal principal cell
 Epididymal basal cell
Extracellular Matrix Cells
 Ameloblast epithelial cell (tooth enamel secretion)
 Planum semilunatum epithelial cell of vestibular apparatus of ear (proteoglycan
secretion)
 Organ of Corti interdental epithelial cell (secreting tectorial membrane covering
hair cells)
 Loose connective tissue fibroblasts
 Corneal fibroblasts (corneal keratocytes)
 Tendon fibroblasts
 Bone marrow reticular tissue fibroblasts
 Other nonepithelial fibroblasts
 Pericyte
 Nucleus pulposus cell of intervertebral disc
 Cementoblast/cementocyte (tooth root bonelike cementum secretion)
 Odontoblast/odontocyte (tooth dentin secretion)
 Hyaline cartilage chondrocyte
 Fibrocartilage chondrocyte
 Elastic cartilage chondrocyte
 Osteoblast/osteocyte
 Osteoprogenitor cell (stem cell of osteoblasts)
 Hyalocyte of vitreous body of eye
 Stellate cell of perilymphatic space of ear
 Hepatic stellate cell (Ito cell)
 Pancreatic stelle cell

Contractile cells
 Skeletal muscle cells
o Red skeletal muscle cell (slow)
o White skeletal muscle cell (fast)
o Intermediate skeletal muscle cell
o nuclear bag cell of muscle spindle
o nuclear chain cell of muscle spindle
 Satellite cell (stem cell)
 Heart muscle cells
o Ordinary heart muscle cell
o Nodal heart muscle cell
o Purkinje fiber cell
 Smooth muscle cell (various types)
 Myoepithelial cell of iris
 Myoepithelial cell of exocrine glands

Blood and immune system cells

 Erythrocyte (red blood cell)
 Megakaryocyte (platelet precursor)
 Monocyte
 Connective tissue macrophage (various types)
 Epidermal Langerhans cell
 Osteoclast (in bone)
 Dendritic cell (in lymphoid tissues)
 Microglial cell (in central nervous system)
  Neutrophil granulocyte
 Eosinophil granulocyte
 Basophil granulocyte
 Mast cell
 Helper T cell
 Suppressor T cell
 Cytotoxic T cell
 Natural Killer T cell
 B cell
 Natural killer cell
 Reticulocyte
 Stem cells and committed progenitors for the blood and immune system (various
types)

Pigment cells
 Melanocyte
 Retinal pigmented epithelial cell

Germ cells
 Oogonium/Oocyte
 Spermatid
 Spermatocyte
 Spermatogonium cell (stem cell for spermatocyte)
 Spermatozoon

Nurse cells
 Ovarian follicle cell
 Sertoli cell (in testis)
 Thymus epithelial cell

Interstitial cells
 Interstitial kidney cells
 ,

Human cell types / list derived primarily from endoderm

Pneumocyte (Type I pneumocyte, Type II pneumocyte) ·

Respiratory
Clara cell · Goblet cell

enteroendocrine: G cell · D cell · ECL cell

Foregut
Stomach exocrine: Gastric chief cell · Parietal cell
Digestive Foveolar cell

Intestine enteroendocrine: K cell · S cell · D cell · I cell

 Goblet cell · Paneth cell

Enterocyte (Microfold cell)

Hepatocyte · Hepatic stellate cell · (Kupffer cell

Liver
from mesoderm)

GallbladderCholecystocyte

Exocrine
Centroacinar cell · Pancreatic stellate cell
pancreas

endocrine pancreas (alpha cell, beta cell, delta cell, PP cell,

Endocrine
epsilon cell)

Pharyngeal thyroid (Follicular cell) · parathyroid (Parathyroid chief cell,

Endocrine
pouch Oxyphil cell)

Hindgut/cloacaUrogenitalUrothelial cells

Human cell types / list derived primarily from mesoderm

bone: Osteoblast → Osteocyte

OCP
cartilage: Chondroblast → Chondrocyte
Cartilage/bone/
muscle Fibroblast → Fibrocyte
(MSC)
Paraxial muscle: Myoblast → Myocyte · Myosatellite
Myofibroblast
cell · Tendon cell · Cardiac muscle cell

adipose: Lipoblast → Adipocyte

Digestive
Interstitial cell of Cajal
system

Angioblast → Endothelial cell · Mesangial cell

(Intraglomerular, Extraglomerular) · Juxtaglomerular cell ·
Macula densa cell
Intermedia Urinary
te system
Stromal cell → Interstitial cell → Telocytes
(RSC)
Simple epithelial cell → Podocyte · Kidney proximal tubule
brush border cell
 ReproductiveSertoli cell · Leydig cell · Granulosa cell · Peg cell ·
system (spermatozoon and ovum are germ cells)

Lymphoi
d (CFU- see lymphocytes
L)
Blood/immune
Lateral (HSC)
plate/ Myeloid
hemangiob (CFU- see myeloid cells
last GEMM)

Circulatory Endothelial progenitor cell · Endothelial stem cell ·

system Angioblast/Mesoangioblast · Pericyte · Mural cell

M: BON/CAR anat(c/f/k/f, u, t/p, noco/cong/tumr, proc, drug(M5)

l)/phys/devp/cell sysi/epon, injr

M: URI anat/phys/devp/cell noco/acba/cong/tumr, proc/itvp, drug

sysi/epon, urte (G4B), blte, urte

M: anat(a:h/u/t/a/l,v:h/u/t/a/l)/phys/dev noco/syva/cong/lyvd/ proc,

VA p/cell/prot tumr, sysi/epon, injr drug(C2s+n/3/4/5/7
S /8/9)

Human cell types / list derived primarily from ectoderm

Integumentary
Trichocyte · Keratinocyte
system
Surface
ectoderm
Nervous anterior pituitary (Gonadotrope, Corticotrope, Thyrotrope,
system Somatotrope, Lactotroph )

Endocrine
Chromaffin cell · Parafollicular cell
system

IntegumentaryMelanoblast → Melanocyte (Nevus cell)

system Merkel cell
Neural
crest Teeth Odontoblast · Cementoblast

Nervous
glia: Schwann cell · Satellite glial cell
system

Eyes Corneal keratocyte

 Digestive
Enterochromaffin cell
system

NervousNeuroblastNeuron
system
Neural
(Neural
tube Oligodendrocyte progenitor → Oligodendrocyte ·
stem Glioblast
cell) Astrocyte · Ependymal cell · Pinealocyte

2. anatomi (struktur sel, organel dan fungsi masing2), fisiologi (sistim

homeostasis sel) dan biokimia sel tubuh (proses sel dalam menjalankan
fungsi masing2 organel)  100%

FUNCTIONAL MORPHOLOGY OF THE CELL

Revolutionary advances in the understanding of cell structure and function have been
made through use of the techniques of modern cellular and molecular biology. There
have also been major advances in the study of embryology and development at the
cellular level. Developmental biology and the details of cell biology are beyond the scope
of this book. However, a basic knowledge of cell biology is essential to an understanding
of the organ systems in the body and the way they function.
The specialization of the cells in the various organs is very great, and no cell can
be called "typical" of all cells in the body. However, a number of structures (organelles)
are common to most cells. These structures are shown in Figure 1-4. Many of them can
be isolated by ultracentrifugation combined with other techniques. When cells are
homogenized and the resulting suspension is centrifuged, the nuclei sediment first,
followed by the mitochondria. High-speed centrifugation that generates forces of
100,000 times gravity or more causes a fraction made up of granules called the
microsomes to sediment. This fraction includes organelles such as the ribosomes and
peroxisomes.

Cell Membrane
The membrane that surrounds the cell is a remarkable structure. It is made up of
lipids and proteins and is semipermeable, allowing some substances to pass through it
and excluding others. However, its permeability can also be varied because it contains
numerous regulated ion channels and other transport proteins that can change the
amounts of substances moving across it. It is generally referred to as the plasma
membrane. The nucleus is also surrounded by a membrane of this type, and the
organelles are surrounded by or made up of a membrane.
Although the chemical structure of membranes and their properties vary
considerably from one location to another, they have certain common features. They are
generally about 7.5 nm (75 Angstrom units) thick. They are made up primarily of protein
and lipids. The chemistry of proteins and lipids is discussed in Chapter 17. The major
lipids are phospholipids such as phosphatidylcholine and phosphatidylethanolamine.
The shape of the phospholipid molecule is roughly that of a clothespin (Figure 1-5). The
head end of the molecule contains the phosphate portion and is relatively soluble in
water (polar, hydrophilic). The tails are relatively insoluble (nonpolar, hydrophobic).
In the membrane, the hydrophilic ends of the molecules are exposed to the aqueous
environment that bathes the exterior of the cells and the aqueous cytoplasm; the
hydrophobic ends meet in the water-poor interior of the membrane. In prokaryotes
(cells such as bacteria in which there is no nucleus), the membranes are relatively
simple, but in eukaryotes (cells containing nuclei), cell membranes contain various
glycosphingolipids, sphingomyelin, and cholesterol.
There are many different proteins embedded in the membrane. They exist as
separate globular units and many pass through the membrane (integral proteins),
whereas others (peripheral proteins) stud the inside and outside of the membrane
(Figure 1-5). The amount of protein varies with the function of the membrane but makes
up on average 50% of the mass of the membrane; ie, there is about one protein molecule
per 50 of the much smaller phospholipid molecules. The proteins in the membranes
carry out many functions. Some are cell adhesion molecules that anchor cells to their
neighbors or to basal laminas. There are proteins that function as pumps, actively
transporting ions across the membrane. Other proteins function as carriers,
transporting substances down electrochemical gradients by facilitated diffusion. Still
others are ion channels, which, when activated, permit the passage of ions into or out
of the cell. The role of the pumps, carriers, and ion channels in transport across the cell
membrane is discussed below. Proteins in another group function as receptors that
bind neurotransmitters and hormones, initiating physiologic changes inside the cell.
Proteins also function as enzymes, catalyzing reactions at the surfaces of the
membrane. In addition, some glycoproteins function in antibody processing and
distinguishing self from nonself (see Chapter 27).
The uncharged, hydrophobic portions of the proteins are usually located in the
interior of the membrane, whereas the charged, hydrophilic portions are located on the
surfaces. Peripheral proteins are attached to the surfaces of the membrane in various
ways. One common way is attachment to glycosylated forms of phosphatidylinositol.
Proteins held by these glycosylphosphatidylinositol (GPI) anchors (Figure 1-5)
include enzymes such as alkaline phosphatase, various antigens, a number of cell
adhesion molecules, and three proteins that combat cell lysis by complement (see
Chapter 27). Over 40 GPI-linked cell surface proteins have now been described. Other
proteins are lipidated, ie, they have specific lipids attached to them (Figure 1-6). They
may be myristolated, palmitoylated, or prenylated, ie, attached to geranylgeranyl
or farnesyl groups.
The protein structure—and particularly the enzyme content—of biologic
membranes varies not only from cell to cell but also within the same cell. For example,
there are different enzymes embedded in cell membranes than in mitochondrial
membranes. In epithelial cells, the enzymes in the cell membrane on the mucosal surface
differ from those in the cell membrane on the basal and lateral margins of the cells; ie,
the cells are polarized. This is what makes transport across epithelia possible (see
below). The membranes are dynamic structures, and their constituents are being
constantly renewed at different rates. Some proteins are anchored to the cytoskeleton,
but others move laterally in the membrane. For example, receptors move in the
membrane and aggregate at sites of endocytosis (see below).
Underlying most cells is a thin, fuzzy layer plus some fibrils that collectively make
up the basement membrane or, more properly, the basal lamina. The basal lamina
and, more generally, the extracellular matrix are made up of many proteins that hold
cells together, regulate their development, and determine their growth. These include
collagens, laminins (see below), fibronectin, tenascin, and proteoglycans.

Mitochondria

Although the morphology of mitochondria varies somewhat from cell to cell, each
mitochondrion (Figure 1-4) is in essence a sausage-shaped structure. It is made up of an
outer membrane and an inner membrane. The latter is folded to form shelves (cristae).
The space between the two membranes is called the intracristal space, and the space
inside the inner membrane is called the matrix space. The mitochondria are the power-
generating units of the cell and are most plentiful and best developed in parts of cells
where energy-requiring processes take place. The chemical reactions occurring in them
are discussed in detail in Chapter 17. The outer membrane of each mitochondrion is
studded with the enzymes concerned with biologic oxidations, providing raw materials
for the reactions occurring inside the mitochondrion. In the interior, the enzymes that
convert the products of carbohydrate, protein, and fat metabolism to CO 2 and water are
located in the internal mitochondrial membrane. Four complex enzymes are involved
(Figure 1-7): NADH dehydrogenase, succinic dehydrogenase, cytochrome c,
and cytochrome oxidase. During these reactions, protons (H+) are pumped from the
matrix to the intracristal space, establishing a proton gradient. Protons diffusing back
along this gradient drive the synthesis of adenosine triphosphate (ATP) by the
enzyme ATP synthase. ATP is the energy-rich triphosphate which provides the energy
for many key metabolic processes, not only in animals (see Chapter 17) but in bacteria
and plants as well, which synthesize it in different ways. The coupling of oxidation with
formation of ATP in the mitochondria of animals is called oxidative
phosphorylation.

ATP synthase is a unique enzyme made up of many subunits with a base embedded in
the inner mitochondria membrane, a neck, and a globular head in the mitochondrial
matrix. Most of the neck and the base actually rotate as ATP is formed.
It is almost certain that mitochondria were once autonomous microorganisms
that developed a symbiotic relation with and became incorporated into ancestral
eukaryotic cells. Consistent with this origin is the fact that mitochondria have their own
genome. There is much less DNA in the mitochondrial genome than in the nuclear
genome (see below), and 99% of the proteins in the mitochondria are the products of
nuclear genes, but mitochondrial DNA is responsible for certain key components of the
pathway for oxidative phosphorylation. Specifically, human mitochondrial DNA is a
double-stranded circular molecule containing 16,569 base pairs (compared with over a
billion in nuclear DNA). It codes for 13 protein subunits that are associated with proteins
encoded by nuclear genes to form four enzyme complexes plus two ribosomal RNAs and
22 transfer RNAs (see below) that are needed for protein production by the
intramitochondrial ribosomes.
Sperm contributes no mitochondria to the zygote; therefore, mitochondria come
from the ovum and inheritance is strictly maternal. There is no effective DNA repair
system in the mitochondria, and the mutation rate for mitochondrial DNA is over ten
times the rate for nuclear DNA. A large number of relatively rare diseases have now been
traced to mutations in mitochondrial DNA. These include for the most part disorders of
tissues with high metabolic rates in which energy production is defective as a result of
abnormalities in the production of ATP.
Evidence is accumulating that mammalian cell death (apoptosis) is initiated in
the mitochondria—or at least involves them at an early stage (see below).

Lysosomes

In the cytoplasm of the cell, there are large, somewhat irregular structures surrounded
by membrane. The interior of these structures, which are called lysosomes, is more
acidic than the rest of the cytoplasm, and external material such as endocytosed bacteria
as well as worn-out cell components are digested in them. Some of the enzymes involved
are listed in Table 1-3.
When a lysosomal enzyme is congenitally absent, the lysosomes become
engorged with the material the enzyme normally degrades. This eventually leads to one
of the lysosomal storage diseases. For example, α-galactosidase A deficiency causes
Fabry's disease, and β-galactocerebrosidase deficiency causes Gaucher's disease. These
diseases are rare, but they are serious and can be fatal. Another example is the lysosomal
storage disease called Tay-Sachs disease, which causes mental retardation and blindness.

Peroxisomes

Peroxisomes are found in the microsomal fraction of cells. They are 0.5 mm in diameter
and are surrounded by a membrane. This membrane contains a number of peroxisome-
specific proteins that are concerned with transport of substances into and out of the
matrix of the peroxisome. The matrix contains more than 40 enzymes, which operate in
concert with enzymes outside the peroxisome to catalyze a variety of anabolic and
catabolic reactions. Several years ago, a number of synthetic compounds were found to
cause proliferation of peroxisomes by acting on receptors in the nuclei of cells. These
receptors (PPARs) are members of the nuclear receptor superfamily, which includes
receptors for steroid hormones, thyroid hormones, certain vitamins, and a number of
other substances (see below). When activated, they bind to DNA, producing changes in
the production of mRNAs. Three PPAR receptors—α, β, and γ—have been characterized.
PPAR-α and PPAR-γ have received the most attention because PPAR-γ's are activated by
feeding and initiate increases in enzymes involved in energy storage, whereas PPAR-α's
are activated by fasting and increase energy-producing enzyme activity.
Thiazolidinediones are synthetic ligands for PPAR-γ's and they increase sensitivity to
insulin, though their use in diabetes has been limited by their toxic side effects. Fibrates,
which lower circulating triglycerides, are ligands for PPAR-α's.

Cytoskeleton

All cells have a cytoskeleton, a system of fibers that not only maintains the structure of
the cell but also permits it to change shape and move. The cytoskeleton is made up
primarily of microtubules, intermediate filaments, and microfilaments, along with
proteins that anchor them and tie them together. In addition, proteins and organelles
move along microtubules and microfilaments from one part of the cell to another
propelled by molecular motors.

Microtubules (Figures 1-8 and 1-9) are long, hollow structures with 5-nm walls
surrounding a cavity 15 nm in diameter. They are made up of two globular protein
subunits, α- and β-tubulin. A third subunit, γ-tubulin, is associated with the production
of microtubules by the centrosomes (see below). The α and β subunits form
heterodimers (Figure 1-9), which aggregate to form long tubes made up of stacked rings,
with each ring usually containing 13 subunits. The tubules also contain other proteins
that facilitate their formation. The assembly of microtubules is facilitated by warmth and
various other factors, and disassembly is facilitated by cold and other factors. The end
where assembly predominates is called the + end, and the end where disassembly
predominates is the - end. Both processes occur simultaneously in vitro.
Because of their constant assembly and disassembly, microtubules are a dynamic
portion of the cell skeleton. They provide the tracks for transport of vesicles, organelles
such as secretory granules, and mitochondria from one part of the cell to another. They
also form the spindle, which moves the chromosomes in mitosis. Microtubules can
transport in both directions.
Microtubule assembly is prevented by colchicine and vinblastine. The anticancer
drug paclitaxel (Taxol) binds to microtubules and makes them so stable that
organelles cannot move. Mitotic spindles cannot form, and the cells die.
Intermediate filaments are 8-14 nm in diameter and are made up of various
subunits. Some of these filaments connect the nuclear membrane to the cell membrane.
They form a flexible scaffolding for the cell and help it resist external pressure. In their
absence, cells rupture more easily; and when they are abnormal in humans, blistering of
the skin is common.

Microfilaments (Figure 1-8) are long solid fibers 4-6 nm in diameter. They are made
up of actin, the protein that by its interaction with myosin brings about contraction of
muscle (see Chapter 3). Actin and its mRNA (see below) are present in all types of cells.
It is the most abundant protein in mammalian cells, sometimes accounting for as much
as 15% of the total protein in the cell. Its structure is highly conserved; for example, 88%
of the amino acid sequences in yeast and rabbit actin are identical. The actin molecules
(G-actin) polymerize in vivo to form F-actin, the long filamentous chains that are the
microfilaments. They also depolymerize in vivo, with polymerization often occurring at
one end of a microfilament (the + end, as in microtubules) and depolymerization at the
other (the - end). They attach to various parts of the cytoskeleton (Figure 1-10). They
reach to the tips of the microvilli on the epithelial cells of the intestinal mucosa. They are
also abundant in the lamellipodia that cells put out when they crawl along surfaces. The
actin filaments interact with integrin receptors and form focal adhesion complexes
(focal adhesions; see below) which serve as points of traction with the surface over which
the cell pulls itself.

Molecular Motors
The molecular motors that move proteins, organelles, and other cell parts (their
cargo) to all parts of the cell are 100-500 kDa ATPases that attach to the cargo. Their
heads form cross-bridges to the microtubules or other actin polymers, and they convert
ATP into energy that produces bending of the cross-bridges, causing the ATPase
molecules to move. There are two types of molecular motors: those producing motion
along microtubules and those producing motion along actin (Table 1-4). Examples are
shown in Figure 1-11, but each type is a superfamily, with many forms throughout the
animal kingdom.
The conventional form of kinesin is a double-headed molecule that moves its
cargo toward the + ends of microtubules. One head binds to the microtubule and then
bends its neck while the other head swings forward and binds, producing almost
continuous movement. Some kinesins are associated with mitosis and meiosis. Other
kinesins perform other functions, including, in some instances, moving cargo to the - end
of microtubules.
Dyneins have two heads, with their neck pieces embedded in a complex of
proteins (Figure 1-11). Cytoplasmic dynein has a function like that of conventional
kinesin, except that it moves particles and membranes to the - end of the microtubules.
Axonemal dynein oscillates and is responsible for the beating of flagella and cilia (see
below).
Myosins form cross-bridges to actin filaments and the myosin heads move,
generating force. This produces movement that literally ranges from contraction of
intestinal villi to cell migration and contraction of all the skeletal and other muscles in
the human body. The myosin superfamily is divided into 15 classes. Myosin-I and
myosin-II are shown in Figure 1-11. Myosin-I is associated with the actin in many cells,
whereas myosin-II (discussed in detail in Chapter 3) is the form present in skeletal
muscle.

Myosin molecules have globular heads, which contain the ATPase activity, and tails of
various lengths (Figure 1-11). Myosin-I molecules have a single head. Along with actin,
myosin-I is often associated with cell membranes. Myosin-II molecules have two heads,
but only one head is active in a given molecule

Centrosomes

Near the nucleus in the cytoplasm of eukaryotic animal cells is a centrosome. The
centrosome is made up of two centrioles and surrounding amorphous pericentriolar
material. The centrioles are short cylinders arranged so that they are at right angles to
each other. Microtubules in groups of three run longitudinally in the walls of each
centriole (Figure 1-4). There are nine of these triplets spaced at regular intervals around
the circumference.

The centrosomes are microtubule-organizing centers (MTOCs) that contain γ-

tubulin. The microtubules grow out of this γ-tubulin in the pericentriolar material. When
a cell divides, the centrosomes duplicate themselves, and the pairs move apart to the
poles of the mitotic spindle, where they monitor the steps in cell division. In
multinucleate cells, there is a centrosome near each nucleus.

Cilia

There are various types of projections from cells. True cilia are dynein-driven motile
processes that are used by unicellular organisms to propel themselves through the water
and by multicellular organisms to propel mucus and other substances over the surface of
various epithelia. They resemble centrioles in having an array of nine tubular structures
in their walls, but they have in addition a pair of microtubules in the center, and there
are two rather than three microtubules in each of the nine circumferential structures.
The basal granule, on the other hand, is the structure to which each cilium is
anchored. It has nine circumferential triplets, like a centriole, and there is evidence that
basal granules and centrioles are interconvertible.

Cell Adhesion Molecules

Cells are attached to the basal lamina and to each other by cell adhesion molecules
(CAMs) that are prominent parts of the intercellular connections described below.
These adhesion proteins have attracted great attention in recent years because they are
important in embryonic development and formation of the nervous system and other
tissues; in holding tissues together in adults; in inflammation and wound healing; and in
the metastasis of tumors. Many pass through the cell membrane and are anchored to the
cytoskeleton inside the cell. Some bind to like molecules on other cells (homophilic
binding), whereas others bind to other molecules (heterophilic binding). Many bind to
laminins, a family of large cross-shaped molecules with multiple receptor domains in
the extracellular matrix.
Nomenclature in the CAM field is somewhat chaotic, partly because the field is
growing so rapidly and partly because of the extensive use of acronyms, as in other areas
of modern biology. However, the CAMs can be divided into four broad families: (1)
integrins, heterodimers that bind to various receptors; (2) adhesion molecules of the
IgG superfamily of immunoglobulins; (3) cadherins, Ca2+-dependent molecules that
mediate cell-to-cell adhesion by homophilic reactions; and (4) selectins, which have
lectin-like domains that bind carbohydrates. The functions of CAMs in granulocytes and
platelets are described in Chapter 27, and their roles in inflammation and wound healing
are discussed in Chapter 33.

The CAMs not only fasten cells to their neighbors—they also transmit signals into and
out of the cell. Cells that lose their contact with the extracellular matrix via integrins have
a higher rate of apoptosis (see below) than anchored cells, and interactions between
integrins and the cytoskeleton are involved in cell movement.

Intercellular Connections
Two types of junctions form between the cells that make up tissues: junctions
that fasten the cells to one another and to surrounding tissues, and junctions that permit
transfer of ions and other molecules from one cell to another. The types of junctions that
tie cells together and endow tissues with strength and stability include the tight
junction, which is also known as the zonula occludens. The desmosome and
zonula adherens (Figure 1-12) hold cells together, and the hemidesmosome and
focal adhesion attach cells to their basal laminas. Tight junctions between epithelial
cells are also essential for transport of ions across epithelia. The junction by which
molecules are transferred is the gap junction.
Tight junctions characteristically surround the apical margins of the cells in
epithelia such as the intestinal mucosa, the walls of the renal tubules, and the choroid
plexus. They are made up of ridges—half from one cell and half from the other—which
adhere so strongly at cell junctions that they almost obliterate the space between the
cells. They permit the passage of some ions and solute, and the degree of this "leakiness"
varies. Extracellular fluxes of ions and solute across epithelia at these junctions are a
significant part of overall ion and solute flux. In addition, tight junctions prevent the
movement of proteins in the plane of the membrane, helping to maintain the different
distribution of transporters and channels in the apical and basolateral cell membranes
that make transport across epithelia possible (see above and Chapters 25 and 38).
In epithelial cells, each zonula adherens is usually a continuous structure on the basal
side of the zonula occludens, and it is a major site of attachment for intracellular
microfilaments. It contains cadherins.
Desmosomes are patches characterized by apposed thickenings of the
membranes of two adjacent cells. Attached to the thickened area in each cell are
intermediate filaments, some running parallel to the membrane and others radiating
away from it. Between the two membrane thickenings, there is filamentous material in
the intercellular space that includes cadherins and the extracellular portions of several
other transmembrane proteins.
Hemidesmosomes look like half-desmosomes that attach cells to the underlying
basal lamina and are connected intracellularly to intermediate filaments. However, they
contain integrins rather than cadherins. Focal adhesions also attach cells to their basal
laminas. As noted above, they are labile structures associated with actin filaments inside
the cell, and they play an important role in cell movement.

Gap Junctions
At gap junctions, the intercellular space narrows from 25 nm to 3 nm, and
hexagonal arrays of protein units called connexons in the membrane of each cell are
lined up with one another (Figure 1-13). Each connexon is made up of six subunits
surrounding a channel that, when lined up with the channel in the corresponding
connexon in the adjacent cell, permits substances to pass between the cells without
entering the ECF. The diameter of the channel is normally about 2 nm, which permits
the passage of ions, sugars, amino acids, and other solutes with molecular weights up to
about 1000. Gap junctions thus permit the rapid propagation of electrical activity from
cell to cell (see Chapter 4) and the exchange of various chemical messengers. The
diameter of each channel is regulated by intracellular Ca 2+, and an increase in Ca2+
concentration causes the subunits to slide together, reducing the diameter of the
channel. The diameter may also be regulated by pH and voltage.
Connexon isoforms are encoded by at least 13 different genes in rodents, and it
seems likely that there are at least this many different genes that are involved in humans.
There is considerable variation in distribution of the isoforms from one tissue to another.
In mice, the ovum is connected to granulosa cells by tight junctions, and mice deficient
in one type of connexon fail to ovulate and complete meiosis. A number of diseases in
humans have been linked to mutant connexons. One is the X-linked form of Charcot-
Marie-Tooth disease, a peripheral neuropathy, and another is heterotaxia, in which
there are multiple abnormalities with failure to establish normal left-right asymmetry. It
is not known how connexon abnormalities lead to these diseases, and at least for
heterotaxia, other genes are probably involved.

Nucleus & Related Structures

A nucleus is present in all eukaryotic cells that divide. If a cell is cut in half, the
anucleate portion eventually dies without dividing. The nucleus is made up in large part
of the chromosomes, the structures in the nucleus that carry a complete blueprint for
all the heritable species and individual characteristics of the animal. Except in germ cells,
the chromosomes occur in pairs, one originally from each parent (see Figure 23-2). Each
chromosome is made up of a giant molecule of deoxyribonucleic acid (DNA). The
DNA strand is about 2 m long, but it can fit in the nucleus because at intervals it is
wrapped around a core of histone proteins to form a nucleosome. There are about 25
million nucleosomes in each nucleus. Thus, the structure of the chromosomes has been
likened to a string of beads. The beads are the nucleosomes, and the linker DNA between
them is the string. The whole complex of DNA and proteins is called chromatin. During
cell division, the coiling around histones is loosened, probably by acetylation of the
histones, and pairs of chromosomes become visible, but between cell divisions only
clumps of chromatin can be discerned in the nucleus. The ultimate units of heredity are
the genes on the chromosomes (see below), and each gene is a portion of the DNA
molecule.
During normal cell division by mitosis, the chromosomes duplicate themselves
and then divide in such a way that each daughter cell receives a full complement
(diploid number) of chromosomes. During their final maturation, germ cells undergo
a division in which half the chromosomes go to each daughter cell (see Chapter 23). This
reduction division (meiosis) is actually a two-stage process, but the important
consideration is that as a result of it, mature sperms and ova contain half the normal
number (the haploid number) of chromosomes. When a sperm and ovum unite, the
resultant cell (zygote) has a full diploid complement of chromosomes, one-half from the
female parent and one-half from the male. The chromosomes undergo recombination,
which mixes maternal and paternal genes.
The nucleus of most cells contains a nucleolus (Figure 1-4), a patchwork of
granules rich in ribonucleic acid (RNA). In some cells, the nucleus contains several of
these structures. Nucleoli are most prominent and numerous in growing cells. They are
the site of synthesis of ribosomes, the structures in the cytoplasm in which proteins are
synthesized (see below).
The interior of the nucleus has a skeleton of fine filaments that are attached to
the nuclear membrane, or envelopee (Figure 1-4), which surrounds the nucleus.
This membrane is a double membrane, and spaces between the two folds are called
perinuclear cisterns. The membrane is permeable only to ions and small molecules.
However, it contains nuclear pore complexes. Each complex has eightfold symmetry
and is made up of about 100 proteins organized to form a tunnel through which
transport of proteins and mRNA occurs. There are many transport pathways, and
proteins called importins and exportins have been isolated and characterized. A
protein named Ran appears to play an organizing role. Much current research is focused
on transport into and out of the nucleus, and a more detailed understanding of these
processes should emerge in the near future.

Endoplasmic Reticulum
The endoplasmic reticulum is a complex series of tubules in the cytoplasm of
the cell (Figure 1-4). The outer limb of its membrane is continuous with a segment of the
nuclear membrane, so in effect this part of the nuclear membrane is a cistern of the
endoplasmic reticulum. The tubule walls are made up of membrane. In rough or
granular endoplasmic reticulum, granules called ribosomes are attached to the
cytoplasmic side of the membrane, whereas in smooth or agranular endoplasmic
reticulum, the granules are absent. Free ribosomes are also found in the cytoplasm.
The granular endoplasmic reticulum is concerned with protein synthesis and the initial
folding of polypeptide chains with the formation of disulfide bonds. The agranular
endoplasmic reticulum is the site of steroid synthesis in steroid-secreting cells and the
site of detoxification processes in other cells. As the sarcoplasmic reticulum (see Chapter
3), it plays an important role in skeletal and cardiac muscle.

Ribosomes

The ribosomes in eukaryotes measure approximately 22 by 32 nm. Each is made up of a

large and a small subunit called, on the basis of their rates of sedimentation in the
ultracentrifuge, the 60S and 40S subunits. The ribosomes are complex structures,
containing many different proteins and at least three ribosomal RNAs (see below). They
are the sites of protein synthesis. The ribosomes that become attached to the
endoplasmic reticulum synthesize all transmembrane proteins, most secreted proteins,
and most proteins that are stored in the Golgi apparatus, lysosomes, and endosomes. All
these proteins have a hydrophobic signal peptide at one end (see below). The polypeptide
chains that form these proteins are extruded into the endoplasmic reticulum. The free
ribosomes synthesize cytoplasmic proteins such as hemoglobin (see Chapter 27) and the
proteins found in peroxisomes and mitochondria.

The Golgi apparatus, which is involved in processing proteins formed in the

ribosomes, and secretory granules, vesicles, and endosomes are discussed below in the
context of protein synthesis and secretion.

STRUCTURE & FUNCTION OF DNA & RNA

The Genome
DNA is found in bacteria, in the nuclei of eukaryotic cells, and in mitochondria. It
is made up of two extremely long nucleotide chains containing the bases adenine (A),
guanine (G), thymine (T), and cytosine (C) (Figure 1-14). The chemistry of these purine
and pyrimidine bases and of nucleotides is discussed in Chapter 17. The chains are
bound together by hydrogen bonding between the bases, with adenine bonding to
thymine and guanine to cytosine. The resultant double-helical structure of the molecule
is shown in Figure 1-15. An indication of the complexity of the molecule is the fact that
the DNA in the human haploid genome (the total genetic message) is made up of 3 × 10 9
base pairs.

DNA is the component of the chromosomes that carry the "genetic message," the
blueprint for all the heritable characteristics of the cell and its descendants. Each
chromosome contains a segment of the DNA double helix. The genetic message is
encoded by the sequence of purine and pyrimidine bases in the nucleotide chains. The
text of the message is the order in which the amino acids are lined up in the proteins
manufactured by the cell. The message is transferred to ribosomes, the sites of protein
synthesis in the cytoplasm, by RNA. RNA differs from DNA in that it is single-stranded,
has uracil in place of thymine, and its sugar moiety is ribose rather than 2´-deoxyribose
(see Chapter 17). The proteins formed from the DNA blueprint include all the enzymes,
and these in turn control the metabolism of the cell. A gene used to be defined as the
amount of information necessary to specify a single protein molecule. However, the
protein encoded by a single gene may be subsequently divided into several different
physiologically active proteins. In addition, different mRNAs can be formed from a gene,
with each mRNA dictating formation of a different protein. Genes also contain
promoters, DNA sequences that facilitate the formation of RNA. Mutations occur when
the base sequence in the DNA is altered by x-rays, cosmic rays, or other mutagenic
agents.

The Human Genome

When the human genome was finally mapped several years ago, there was
considerable surprise that it contained only about 30,000 genes and not the 50,000 or
more that had been expected. Yet humans differ quite markedly from their nearest
simian relatives. The explanation appears to be that rather than a greater number of
genes in humans, there is a greater number of mRNAs—perhaps as many as 85,000. The
implications of this increase are discussed below.

DNA Polymorphism

The protein-coding genes (exons) make up only 3% of the human genome; the remaining
97% is made up of introns (see below) and other DNA of unsettled or unknown function.
This 97% is sometimes called junk DNA. A characteristic of human DNA is its
structural variability from one individual to another. Most of the variations occur in
noncoding regions, but they can also occur in coding regions, where they can be silent or
expressed as a detectable alteration in a protein. A common form of these variations is
variable repetition of base pairs (tandem repeats) from one to hundreds of times. This
variation alters the length of the DNA chain between points where it is cut by various
restriction enzymes, so that there is restriction fragment length polymorphism
(RFLP) in the DNA fragments from different individuals. Consequently, analysis of
RFLP in a population gives a pattern that is in effect a DNA fingerprint. The value of
DNA fingerprinting has been improved by additional specialized techniques. The chance
of obtaining identical DNA patterns by using these techniques in individuals who are not
identical twins varies with the number of enzymes used, the relatedness of the
individuals, and other factors, and there has been debate about the appropriate statistics
to use for analysis. However, the possibility that an RFLP match is due to chance has
been estimated at 1 in 1,000,000 to 1 in 100,000. Furthermore, RFLP analysis can be
carried out on small specimens of semen, blood, or other tissue, and multiple copies of
pieces of DNA can be made by using the polymerase chain reaction (PCR), an
ingenious technique for making DNA copy itself. Therefore, DNA fingerprinting is of
obvious value in investigating crimes and determining paternity, although reliable
techniques must be used and the results interpreted with care. RFLP analysis is also of
value in studying animal and human evolution and in identifying the chromosomal
location of genes causing inherited diseases.

Mitosis
At the time of each somatic cell division (mitosis), the two DNA chains separate, each
serving as a template for the synthesis of a new complementary chain. DNA
polymerase catalyzes this reaction. One of the double helices thus formed goes to one
daughter cell and one goes to the other, so the amount of DNA in each daughter cell is
the same as that in the parent cell.

Telomeres

Cell replication involves not only DNA polymerase but a special reverse transcriptase
that synthesizes the short repeats of DNA that characterize the ends (telomeres) of
chromosomes. Without this transcriptase and related enzymes known collectively as
telomerase, somatic cells lose DNA as they divide 40-60 times and then become
senescent and undergo apoptosis (see below). On the other hand, cells with high
telomerase activity, which includes most cancer cells, can in theory keep multiplying
indefinitely. Not surprisingly, there has been considerable interest in the telomerase
mechanism, both in terms of aging and in terms of cancer. However, it now seems clear
that the mechanism for replicating chromosome ends is complex, and much additional
research will be needed before a complete understanding is achieved and therapeutic
applications emerge.

Meiosis

In germ cells, reduction division (meiosis) takes place during maturation. The net
result is that one of each pair of chromosomes ends up in each mature germ cell;
consequently, each mature germ cell contains half the amount of chromosomal material
found in somatic cells. Therefore, when a sperm unites with an ovum, the resulting
zygote has the full complement of DNA, half of which came from the father and half from
the mother. The chromosomal events that occur at the time of fertilization are discussed
in detail in Chapter 23. The term "ploidy" is sometimes used to refer to the number of
chromosomes in cells. Normal resting diploid cells are euploid and become tetraploid
just before division. Aneuploidy is the condition in which a cell contains other than the
haploid number of chromosomes or an exact multiple of it, and this condition is common
in cancerous cells.

Cell Cycle
Obviously, the initiation of mitosis and normal cell division depends on the orderly
occurrence of events during what has come to be called the cell cycle. A diagram of
these events is shown in Figure 1-16. There is intense interest in the biochemical
machinery that produces mitosis, in part because of the obvious possibility of its relation
to cancer. When DNA is damaged, entry into mitosis is inhibited, giving the cell time to
repair the DNA; failure to repair damaged DNA leads to cancer. The cell cycle is
regulated by proteins called cyclins and cyclin-dependent protein kinases, which
phosphorylate other proteins. However, the regulation is complex, and a detailed
analysis of it is beyond the scope of this book.

Transcription & Translation

The strands of the DNA double helix not only replicate themselves, but also serve
as templates by lining up complementary bases for the formation in the nucleus of
messenger RNA (mRNA), transfer RNA (tRNA), the RNA in the ribosomes
(rRNA), and various other RNAs. The formation of mRNA is called transcription
(Figure 1-17) and is catalyzed by various forms of RNA polymerase. Usually after some
posttranscriptional processing (see below), mRNA moves to the cytoplasm and
dictates the formation of the polypeptide chain of a protein (translation). This process
occurs in the ribosomes. tRNA attaches the amino acids to mRNA. The mRNA molecules
are smaller than the DNA molecules, and each represents a transcript of a small segment
of the DNA chain. The molecules of tRNA contain only 70-80 nitrogenous bases,
compared with hundreds in mRNA and 3 billion in DNA.
It is worth noting that DNA is responsible for the maintenance of the species; it
passes from one generation to the next in germ cells. RNA, on the other hand, is
responsible for the production of the individual; it transcribes the information coded in
the DNA and forms a mortal individual, a process that has been called "budding off from
the germ line."

Genes

Information is accumulating at an accelerating rate about the structure of genes and

their regulation. The structure of a typical eukaryotic gene is shown in diagrammatic
form in Figure 1-18. It is made up of a strand of DNA that includes coding and noncoding
regions. In eukaryotes, unlike prokaryotes, the portions of the genes that dictate the
formation of proteins are usually broken into several segments (exons) separated by
segments that are not translated (introns). A pre-mRNA is formed from the DNA, and
then the introns and sometimes some of the exons are eliminated in the nucleus by
posttranscriptional processing, so that the final mRNA which enters the cytoplasm is
made up of exons (Figure 1-19). Introns are eliminated and exons are joined by several
different processes. The introns of some genes are eliminated by spliceosomes,
complex units that are made up of small RNAs and proteins. Other introns are
eliminated by self-splicing by the RNA they contain. Two different mechanisms
produce self-splicing. RNA can catalyze other reactions as well and there is great interest
today in the catalytic activity of RNA.

Because of introns and splicing, more than one mRNA is formed from the same gene. As
noted above, the formation of multiple proteins from one gene is perhaps one of the
explanations of the surprisingly small number of genes in the human genome. Other
physiologic functions of the introns are still unsettled, though they may foster changes in
the genetic message and thus aid evolution.

Near the transcription start site of the gene is a promoter, which is the site at
which RNA polymerase and its cofactors bind. It often includes a TATA sequence (TATA
box), which ensures that transcription starts at the proper point. Farther out in the 5´
region are regulatory elements, which include enhancer and silencer sequences. It
has been estimated that there are an average of five regulatory sites per gene. Regulatory
sequences are sometimes found in the 3´-flanking region as well, and there is evidence
that sequences in this region can also affect the function of other genes.

Regulation of Gene Expression

Each nucleated somatic cell in the body contains the full genetic message, yet
there is great differentiation and specialization in the functions of the various types of
adult cells. Only small parts of the message are normally transcribed. Thus, the genetic
message is normally maintained in a repressed state. However, genes are controlled both
spatially and temporally. What turns on genes in one cell and not in other cells? What
turns on genes in a cell at one stage of development and not at other, inappropriate
stages? What maintains orderly growth in cells and prevents the uncontrolled growth
that we call cancer? Obviously, DNA sequences such as the TATA box promote orderly
transcription of the gene of which they are a part (cis regulation). However, the major
key to selective gene expression is the proteins that bind to the regulatory regions of the
gene and increase or shut off its activity. These transcription factors are products of
other genes and hence mediate trans regulation. They are extremely numerous and
include activated steroid hormone receptors and many other factors.
It is common for stimuli such as neurotransmitters that bind to the cell
membrane to initiate chemical events which activate immediate-early genes. These
in turn produce transcription factors that act on other genes. The best-characterized
immediate-early genes are c-fos, and c-jun. The proteins produced by these genes, c-
Fos, c-Jun, and several related proteins, form homodimer or heterodimer transcription
factors which bind to a specific DNA regulatory sequence called an AP-1 site (Figure 1-
20). Some of the dimers enhance transcription, and others inhibit it. The appearance of
c-Fos, c-Jun, and related proteins is such a common sign of cell activation that
immunocytochemistry for them or measurement of their mRNAs is used to determine
which cells in the nervous system and elsewhere are activated by particular stimuli.

Over 80% of the known transcription factors have one of four DNA-binding motifs. The
most common is the zinc finger motif, in which characteristically shaped complexes are
formed by coordinate binding of Zn 2+ between two cysteine and two histidine residues or
between four cysteine residues. Various transcription factors contain 2-37 of these zinc
fingers, which mediate the binding to DNA. Another motif is the leucine zipper, in
which α-helical regions of dimers have regularly spaced leucine residues that interact
with one another to form a coiled coil. Extensions of the dimer beyond the zippered
region are rich in arginine and lysine, and these bind to DNA. The other common DNA-
binding motifs are helix-turn-helix and helix-loop-helix structures.
It is now possible by using molecular biologic techniques to augment the function
of particular genes, to transfer human genes into animals, and to disrupt the function of
single genes (gene knockout). The gene knockout technique is currently being used in
numerous experiments.

Protein Synthesis
The process of protein synthesis is a complex but fascinating one that, as noted
above, involves four steps: transcription, posttranscriptional modification, translation,
and posttranslational modification.

When suitably activated, transcription of the gene starts at the cap site (Figure 1-19)
and ends about 20 bases beyond the signal sequence AATAAA. The RNA transcript is
capped in the nucleus by addition of 7-methylguanosine triphosphate to the 5´ end; this
cap is necessary for proper binding to the ribosome (see below). A poly(A) tail of about
100 bases is added to the untranslated segment at the 3´ end. The function of the poly(A)
tail is unsettled, but it may help maintain the stability of the mRNA. The pre-mRNA
formed by capping and addition of the poly(A) tail is then processed by elimination of
the introns (Figure 1-19), and once this posttranscriptional modification is complete, the
mature mRNA moves to the cytoplasm. Posttranscriptional modification of the pre-
mRNA is a regulated process, and, as noted above, differential splicing can occur, with
the formation of more than one mRNA from a single pre-mRNA.
When a definitive mRNA reaches a ribosome in the cytoplasm, it dictates the
formation of a polypeptide chain. Amino acids in the cytoplasm are activated by
combination with an enzyme and adenosine monophosphate (adenylate), and each
activated amino acid then combines with a specific molecule of tRNA. There is at
least one tRNA for each of the 20 unmodified amino acids found in large quantities in
the body proteins of animals (see Chapter 17), but there is more than one tRNA for some
amino acids. The tRNA-amino acid-adenylate complex is next attached to the mRNA
template, a process that occurs in the ribosomes. This process is shown diagrammatically
in Figure 1-17. The tRNA "recognizes" the proper spot to attach on the mRNA template
because it has on its active end a set of three bases that are complementary to a set of
three bases in a particular spot on the mRNA chain. The genetic code is made up of such
triplets, sequences of three purine or pyrimidine bases (or both); each triplet stands for
a particular amino acid

Translation starts in the ribosomes with an AUG (transcribed from ATG in the gene)
which codes for methionine. The amino terminal amino acid is then added, and the chain
is lengthened one amino acid at a time. The mRNA attaches to the 40S subunit of the
ribosome during protein synthesis; the polypeptide chain being formed attaches to the
60S subunit; and the tRNA attaches to both. As the amino acids are added in the order
dictated by the triplet code, the ribosome moves along the mRNA molecule like a bead
on a string. Translation stops at one of three stop, or nonsense, codons—UGA, UAA, or
UAG—and the polypeptide chain is released. The tRNA molecules are used again. The
mRNA molecules are also reused approximately 10 times before being replaced.

Typically, there is more than one ribosome on a given mRNA chain at one time. The
mRNA chain plus its collection of ribosomes is visible under the electron microscope as
an aggregation of ribosomes called a polyribosome (polysome).

At least in theory, synthesis of particular proteins can be stopped by

administering antisense oligonucleotides, short synthetic stretches of bases
complementary to segments of the bases on the mRNA for the protein. These bind to the
mRNA, blocking translation. Early results with this technology were disappointing
because of nonspecific binding and immune responses, but research continues and there
is hope for products that will be useful in the treatment of a variety of diseases, including
cancer.

Posttranslational Modification.After the polypeptide chain is formed, it is modified

to the final protein by one or more of a combination of reactions that include
hydroxylation, carboxylation, glycosylation, or phosphorylation of amino acid residues;
cleavage of peptide bonds that converts a larger polypeptide to a smaller form; and the
folding and packaging of the protein into its ultimate, often complex configuration.

It has been claimed that a typical eukaryotic cell synthesizes about 10,000 different
proteins during its lifetime. How do these proteins get to the right locations in the cell?
Synthesis starts in the free ribosomes. As noted above, most proteins that are going to be
secreted or stored in organelles and most transmembrane proteins have at their amino
terminal a signal peptide (leader sequence) that guides them into the endoplasmic
reticulum. It is made up of 15-30 predominantly hydrophobic amino acid residues. The
signal peptide, once synthesized, binds to a signal recognition particle (SRP), a
complex molecule made up of six polypeptides and 7S RNA, one of the small RNAs. The
SRP stops translation until it binds to a translocon, a pore in the endoplasmic
reticulum that is a heterotrimeric structure made up of Sec 61 proteins. The ribosome
also binds, and the signal peptide leads the growing peptide chain into the cavity of the
endoplasmic reticulum (Figure 1-21). The signal peptide is next cleaved from the rest of
the peptide by a signal peptidase while the rest of the peptide chain is still being
synthesized.

The signals that direct nascent proteins to some of the other parts of the cell are
fashioned in the Golgi apparatus (see below) and involve specific modifications of the
carbohydrate residues on glycoproteins.

Secreted Proteins

Many and perhaps all proteins that are secreted by cells are synthesized as larger
proteins, and polypeptide sequences are cleaved off from them during maturation. In the
case of the hormones, these larger forms are called preprohormones and
prohormones (Figures 1-19 and 1-22). Parathyroid hormone (see Chapter 21) is a good
example. It is synthesized as a molecule containing 115 amino acid residues
(preproparathyroid hormone). The leader sequence, 25 amino acid residues at the amino
terminal, is rapidly removed to form proparathyroid hormone. Before secretion, an
additional 6 amino acids are removed from the amino terminal to form the secreted
molecule. The function of the 6-amino-acid fragment is unknown.

Although most secreted polypeptides and proteins have a leader sequence that targets
them to the endoplasmic reticulum and are secreted by exocytosis (see below), a growing
list of proteins that are secreted lack a signal sequence. These include in humans the
cytokines interleukin-1α (IL-1α) and IL-1β, three growth factors, and various factors
involved in hemostasis. Secretion probably occurs via ATP-dependent membrane
transporters. There is a large family of these ATP-binding-cassette (ABC) transport
proteins, and they transport ions and other substances as well as proteins between
organelles and across cell membranes. In general, they are made up of two cytoplasmic
ATP-binding domains and two membrane domains, each of which probably spans the
membrane and in general contains six long α-helical sequences (Figure 1-23). The cystic
fibrosis transmembrane conductance regulator (CFTR) is one of those ABC
transport proteins that also has a region for regulation by cAMP. It transports Cl - and is
abnormal in individuals with cystic fibrosis (see Chapter 37).

Protein Folding

Protein folding is an additional posttranslational modification. It is a complex process

that is dictated primarily by the sequence of the amino acids in the polypeptide chain. In
some instances, however, nascent proteins associate with other proteins called
chaperones, which prevent inappropriate contacts with other proteins and ensure that
the final "proper" conformation of the nascent protein is reached. Misfolded proteins
and other proteins targeted for degradation are conjugated to ubiquitin and broken
down in the organelles called 26S proteasomes (see Chapter 17).

Apoptosis

In addition to dividing and growing under genetic control, cells can die and be absorbed
under genetic control. This process is called programmed cell death, or apoptosis
(Gr apo "away" + ptosis "fall"). It can be called "cell suicide" in the sense that the cell's
own genes play an active role in its demise. It should be distinguished from necrosis
("cell murder"), in which healthy cells are destroyed by external processes such as
inflammation.

Apoptosis is a very common process during development and in adulthood. In the

central nervous system, large numbers of neurons are produced and then die during the
remodeling that occurs during development and synapse formation (see Chapter 4). In
the immune system, apoptosis gets rid of inappropriate clones of immunocytes (see
Chapter 27) and is responsible for the lytic effects of glucocorticoids on lymphocytes (see
Chapter 20). Apoptosis is also an important factor in processes such as removal of the
webs between the fingers in fetal life and regression of duct systems in the course of
sexual development in the fetus (see Chapter 23). In adults, it participates in the cyclic
breakdown of the endometrium that leads to menstruation (see Chapter 23). In epithelia,
cells that lose their connections to the basal lamina and neighboring cells undergo
apoptosis. This is responsible for the death of the enterocytes sloughed off the tips of
intestinal villi (see Chapter 26). Abnormal apoptosis probably occurs in autoimmune
disease, neurodegenerative diseases, and cancer. It is interesting that apoptosis occurs in
invertebrates, including nematodes and insects. However, its molecular mechanism is
much more complex than that in vertebrates.

The final common pathway bringing about apoptosis is activation of caspases, a group
of cysteine proteases. Thirteen of these have been characterized to date in mammals.
They exist in cells as inactive proenzymes until activated by the cellular machinery. The
net result is DNA fragmentation, cytoplasmic and chromatin condensation, and
eventually membrane bleb formation, with cell breakup and removal of the debris by
phagocytes.

Apoptosis can be triggered by external and internal stimuli. One ligand that activates
receptors triggering apoptosis is Fas, a transmembrane protein that projects from
natural killer cells and T lymphocytes (see Chapter 27) but also exists in a circulating
from. Another is tumor necrosis factor.

Between initiating stimuli and caspase activation is a complex network of excitatory and
inhibitory intracellular proteins. One of the important pathways goes through the
mitochondria, which release cytochrome c and a protein called smac/DIABLO.
Cytochrome c acts with several cytoplasmic proteins to facilitate caspase activation.
Smac/DIABLO binds to several inhibiting proteins, lifting the inhibition of caspase-9
and thus increasing apoptotic activity.

Molecular Medicine

Fundamental research in molecular aspects of genetics, regulation of gene expression,

and protein synthesis has been paying off in clinical medicine at a rapidly accelerating
rate.
One early dividend was an understanding of the mechanisms by which antibiotics exert
their effects. Almost all act by inhibiting protein synthesis at one or another of the steps
described above. Antiviral drugs act in a similar way; for example, acyclovir and
ganciclovir act by inhibiting DNA polymerase. Some of these drugs have this effect
primarily in bacteria, but others inhibit protein synthesis in the cells of other animals,
including mammals. This fact makes antibiotics of great value for research as well as for
treatment of infections.

Single genetic abnormalities that cause over 600 human diseases have now been
identified. Many of the diseases are rare, but others are more common and some cause
conditions that are severe and eventually fatal. Examples include the defectively
regulated Cl- channel in cystic fibrosis (see above and Chapter 34) and the unstable
trinucleotide repeats in various parts of the genome that cause Huntington's disease,
the fragile X syndrome, and several other neurologic diseases (see Chapter 12).
Abnormalities in mitochondrial DNA can also cause human diseases such as Leber's
hereditary optic neuropathy and some forms of cardiomyopathy. Not surprisingly,
genetic aspects of cancer are probably receiving the greatest current attention. Some
cancers are caused by oncogenes, genes which are carried in the genomes of cancer
cells and are responsible for producing their malignant properties. These genes are
derived by somatic mutation from closely related proto-oncogenes, which are normal
genes that control growth. Over 100 oncogenes have been described. Another group of
genes produce proteins that suppress tumors, and more than 10 of these tumor
suppressor genes have been described. The most studied of these is the P53 gene (p53
in some publications) on human chromosome 17. The P53 protein produced by this gene
triggers apoptosis. It is also a nuclear transcription factor that appears to increase
production of a 21-kDa protein which blocks two cell cycle enzymes, slowing the cycle
and permitting repair of mutations and other defects in DNA. The P53 gene is mutated
in up to 50% of human cancer patients, with the production of P53 proteins that fail to
slow the cell cycle and permit other mutations in DNA to persist. The accumulated
mutations eventually cause cancer.

Gene therapy is still in its infancy, but various ingenious approaches to the problem of
getting genes into cells are now being developed. One that is already in clinical trials for
some diseases involves removal of cells from the patient with the disease, transfection of
the cells with normal genes in vitro, and reinjection of the cells into the patient as an
autotransplant. Another is insertion of appropriate genes into relatively benign viruses
that are then administered to patients to carry the genes to the cells they invade.

Golgi Apparatus & Vesicular Transport in Cells

The Golgi apparatus is a collection of membrane-enclosed sacs (cisterns) that are stacked
like dinner plates (Figure 1-4). There are usually about six sacs in each apparatus, but
there may be more. One or more Golgi apparatuses are present in all eukaryotic cells,
usually near the nucleus. The Golgi apparatus is a polarized structure, with cis and trans
sides (Figure 1-24). Membranous vesicles containing newly synthesized proteins bud off
from the granular endoplasmic reticulum and fuse with the cistern on the cis side of the
apparatus. The proteins are then passed via other vesicles to the middle cisterns and
finally to the cistern on the trans side, from which vesicles branch off into the cytoplasm.
From the trans Golgi, vesicles shuttle to the lysosomes and to the cell exterior via a
constitutive and a nonconstitutive pathway, both involving exocytosis (see below).
Mannose-6-phosphate receptors (MPRs) on the lysosomes capture hydrolytic enzymes
destined for these organelles. Conversely, vesicles are pinched off from the cell
membrane by endocytosis (see below) and pass to endosomes and eventually to the
lysosomes. From the lysosomes, some of the proteins are shuttled back to the trans
Golgi. The initial glycosylation of proteins occurs with the attachment of preformed
oligosaccharides in the endoplasmic reticulum, but these oligosaccharides are altered to
a variety of different carbohydrate moieties in the Golgi apparatus.

Quality Control

The processes involved in protein synthesis, folding, and migration to the various parts
of the cell are so complex that it is remarkable that more errors and abnormalities do not
occur. This is due to mechanisms at each level that are responsible for "quality control."
Damaged DNA is detected and repaired or bypassed. The various RNAs are also checked
during the translation process. Finally, when the protein chains are in the endoplasmic
reticulum and Golgi apparatus, defective structure is detected and the abnormal proteins
are degraded in lysosomes and proteasomes. The net result is a remarkable accuracy in
the production of the proteins needed for normal body function.

TRANSPORT ACROSS CELL MEMBRANES

Transport across cell membranes is accomplished primarily by exocytosis, endocytosis,

movement through ion channels, and primary and secondary active transport.

Exocytosis

Proteins that are secreted by cells move from the endoplasmic reticulum to the Golgi
apparatus, and from the trans Golgi, they are extruded into secretory granules or vesicles
(Figure 1-24). The granules and vesicles move to the cell membrane. Their membrane
then fuses to the cell membrane (Figure 1-25), and the area of fusion breaks down. This
leaves the contents of the granules or vesicles outside the cell and the cell membrane
intact. The extrusion process is called exocytosis. It requires Ca2+ and energy, along
with docking proteins (see below and Chapter 4).

Note that there are two pathways by which secretion from the cell occurs (Figure 1-24).
In the nonconstitutive pathway, proteins from the Golgi apparatus initially enter
secretory granules, where processing of prohormones to the mature hormones occurs
before exocytosis. The other pathway, the constitutive pathway, involves the prompt
transport of proteins to the cell membrane in vesicles, with little or no processing or
storage. The nonconstitutive pathway is sometimes called the regulated pathway, but
this term is misleading because the output of proteins by the constitutive pathway is also
regulated.

Endocytosis

Endocytosis is the reverse of exocytosis. There are various types. Phagocytosis ("cell
eating") is the process by which bacteria, dead tissue, or other bits of material visible
under the microscope are engulfed by cells such as the polymorphonuclear leukocytes of
the blood. The material makes contact with the cell membrane, which then invaginates.
The invagination is pinched off, leaving the engulfed material in the membrane-enclosed
vacuole and the cell membrane intact. Pinocytosis ("cell drinking") is essentially the
same process, the difference being that the substances ingested are in solution and not
visible under the microscope.

Endocytosis can be constitutive or clathrin-mediated. Constitutive endocytosis is not

a specialized process, whereas clathrin-mediated endocytosis occurs at membrane
indentations where the protein clathrin accumulates. Clathrin molecules have the
shape of a triskelion, with three legs radiating from a central hub (Figure 1-26). As
endocytosis progresses, the clathrin molecules form a geometric array that surrounds the
endocytotic vesicle. At the neck of the vesicle, a guanosine triphosphatase protein called
dynamin is involved, either directly or indirectly, in pinching off the vesicle, and this
protein has been called a "pinchase." Once the complete vesicle is formed, the clathrin
falls off and the three-legged proteins recycle to form another vesicle. The vesicle fuses
with and dumps its contents into an early endosome (Figure 1-24). From the early
endosome, a new vesicle can bud off and return to the cell membrane (see Figure 4-4).
Alternatively, the early endosome can become a late endosome and fuse with a
lysosome (Figure 1-24) in which the contents are digested by the lysosomal proteases.

Clathrin-mediated endocytosis is responsible for the internalization of many receptors

and the ligands bound to them—including, for example, nerve growth factor and low-
density lipoproteins (LDL; see Chapter 17). It also plays a major role in synaptic function
(see Chapter 4).

It is apparent that exocytosis adds to the total amount of membrane surrounding the
cell, and if membrane were not removed elsewhere at an equivalent rate, the cell would
enlarge. However, removal of cell membrane occurs by endocytosis, and such exocytosis-
endocytosis coupling maintains the surface area of the cell at its normal size.

Rafts & Caveolae

Some areas of the cell membrane are especially rich in cholesterol and sphingolipids and
have been called rafts. These rafts are probably the precursors of flask-shaped
membrane depressions called caveolae (little caves) when their walls become infiltrated
with a protein called caveolin that resembles clathrin. Three isoforms of caveolin—
caveolins-1, -2, and -3—have been identified. There is considerable debate about the
functions of rafts and caveolae, with evidence that they are involved in cholesterol
regulation and transcytosis (see below). However, mice in which the gene for caveolin-1
is knocked out are relatively healthy, with only some poorly understood blood vessel and
pulmonary abnormalities.

Mechanisms Involved in Vesicle Transport

Considerable progress has been made in analyzing the biochemical basis of vesicle
formation, transport, and docking in cells. It is convenient to consider transport within
the cell and exocytosis and endocytosis together in this regard, since very similar
processes are involved.

It now appears that all vesicles involved in transport have protein coats. In addition to
the caveolin, there are at least four types of coat proteins: AP-1 clathrin, AP-2
clathrin, COPI, and COPII. In humans, 53 coat complex subunits have been
identified. Vesicles that transport proteins from the trans Golgi to lysosomes have AP-1
clathrin coats, and endocytotic vesicles that transport to endosomes have AP-2 clathrin
coats. Vesicles that transport between the endoplasmic reticulum and the Golgi have
COPI and COPII coats. Certain amino acid sequences or attached groups on the
transported proteins ticket the proteins for particular locations. For example, the amino
acid sequence Asn-Pro-any amino acid-Tyr tickets transport from the cell surface to the
endosomes, and, as noted above, mannose-6-phosphate groups ticket transfer from the
Golgi to MPR receptors on the lysosomes. Dynamin appears to be involved in the
formation of vesicles in the Golgi as well as in endocytosis from the cell surface. Much of
the work on proteins involved in vesicle docking and fusion has been done on synaptic
vesicles, and these are discussed in Chapter 4. In general, vesicles move along
microtubules, and when a vesicle reaches its target, it docks when V-snare proteins on
the vesicle latch with T-snare proteins on the target. Each target has a unique set of T-
snare proteins, thus ensuring that only the vesicle with the corresponding set of V-snare
proteins will dock. Various small GTP-binding proteins of the Rab family (see below) are
associated with the various types of vesicles. They appear to guide and facilitate orderly
attachments of these vesicles. In humans, there are 60 Rab proteins and 35 snare
proteins.

Distribution of Ions & Other Substances Across Cell Membranes

The unique properties of the cell membranes are responsible for the differences in the
composition of intracellular and interstitial fluid. Specific values for one mammalian
tissue are shown in Table 1-2. Average values for humans are shown in Figure 1-27.

Membrane Permeability & Membrane Transport Proteins

Small, nonpolar molecules—including O2 and N2—and small uncharged polar molecules

such as CO2 diffuse across the lipid membranes of cells. However, the membranes have
very limited permeability to other substances. Instead, they cross the membranes by
endocytosis and exocytosis and by passage through highly specific transport proteins,
transmembrane proteins that form channels for ions or transport substances such as
glucose, urea, and amino acids. The limited permeability applies even to water, with
simple diffusion being supplemented throughout the body with various water channels
(aquaporins; see Chapters 14 and 38). For reference, the sizes of ions and other
biologically important substances are summarized in Table 1-5.

An important technique that has permitted major advances in our knowledge about
transport proteins is patch clamping. A micropipette is placed on the membrane of a
cell and forms a tight seal to the membrane. The patch of membrane under the pipette
tip usually contains only a few transport proteins, and they can be studied in detail
(Figure 1-28). The cell can be left intact (cell-attached patch clamp). Alternatively,
the patch can be pulled loose from the cell, forming an inside-out patch. A third
alternative is to suck out the patch with the micropipette still attached to the rest of the
cell membrane, providing direct access to the interior of the cell (whole cell
recording).Some transport proteins are simple aqueous ion channels, though many
of these have special features that make them effective for a given substance such as Ca 2+
or, in the case of aquaporins, for water. Some of these transport proteins are
continuously open, whereas others are gated; ie, they have gates that open or close.
Some are gated by alterations in membrane potential (voltage-gated), whereas others
are opened or closed when they bind a ligand (ligand-gated). The ligand is often
external, eg, a neurotransmitter or a hormone. However, it can also be internal;
intracellular Ca2+, cAMP, lipids, or one of the G proteins produced in cells (see below)
can bind directly to channels and activate them. Some channels are also opened by
mechanical stretch, and these mechanosensitive channels play an important role in cell
movement. A typical voltage-gated channel is the Na + channel (see below), and a typical
ligand-gated channel is the acetylcholine receptor (see Chapter 4).

Other transport proteins are carriers that bind ions and other molecules and then
change their configuration, moving the bound molecule from one side of the cell
membrane to the other. Molecules move from areas of high concentration to areas of low
concentration (down their chemical gradient), and cations move to negatively charged
areas whereas anions move to positively charged areas (down their electrical
gradient). When carrier proteins move substances in the direction of their chemical or
electrical gradients, no energy input is required and the process is called facilitated
diffusion. A typical example is glucose transport by the glucose transporter, which
moves glucose down its concentration gradient from the ECF to the cytoplasm of the cell
(see Chapter 19). Other carriers transport substances against their electrical and
chemical gradients. This form of transport requires energy and is called active
transport. In animal cells, the energy is provided almost exclusively by hydrolysis of
ATP (see above and Chapter 17). Not surprisingly, therefore, the carrier molecules are
ATPases, enzymes that catalyze the hydrolysis of ATP. One of these ATPases is sodium-
potassium-activated adenosine triphosphatase (Na+-K+ ATPase), which is also
known as the Na+-K+ pump. There are also H+-K+ ATPases in the gastric mucosa (see
Chapter 26) and the renal tubules (see Chapter 38). Ca2+ ATPase pumps Ca2+ out of cells.
Proton ATPases acidify many intracellular organelles, including parts of the Golgi
complex and lysosomes. F-ATPases are present in mitochondria and synthesize ATP.
Some cell membranes contain ATPases that transport Ca2+.

Some of the transport proteins are called uniports, because they transport only one
substance. Others are called symports, because transport requires the binding of more
than one substance to the transport protein and the substances are transported across
the membrane together. An example is the symport in the intestinal mucosa that is
responsible for the cotransport by facilitated diffusion of Na + and glucose from the
intestinal lumen into mucosal cells (see Chapter 25). Other transporters are called
antiports because they exchange one substance for another. The Na +-K+ ATPase
mentioned above is a typical antiport; it moves three Na + out of the cell in exchange for
each two K+ that it moves into the cell.

Ion Channels

There are ion channels for K+, Na+, Ca2+, and Cl-, and each exists in multiple forms with
diverse properties. Most are made up of identical or very similar subunits. A Ca 2+
channel and a Na+ channel are shown in extended diagrammatic form in Figure 1-29,
and an acetylcholine-gated ion channel is shown in Figure 4-18. Figure 1-30 shows the
multiunit structure of various channels in diagrammatic cross-section. Most K + channels
are tetramers, with each of the four subunits forming part of the pore through which K +
ions pass. Fast-inactivating voltage-gated K+ channels have a unique feature. Each of the
tetramers has a polypeptide ball structure on the end of a polypeptide chain, and the ball
moves into the channel, producing inactivation (Figure 1-31). There are ball-and-chain
structures on all four tetramers even though there is only one K + pore. In the
acetylcholine ion channel and other ligand-gated cation or anion channels, five subunits
make up the pore. Members of the CLC family of Cl - channels are dimers, but they have
two pores, one in each subunit. Finally, aquaporins are tetramers with a water pore in
each of the subunits. Recently, a number of ion channels with intrinsic enzyme activity
have been cloned.

More than 30 different voltage-gated or cyclic nucleotide-gated Na + and Ca2+ channels of

this type have been described. The toxins tetrodotoxin (TTX) and saxitoxin (STX) bind to
the Na+ channels and block them. The number and distribution of the Na + channels can
be determined by tagging them with labeled TTX or STX carrying a suitable label and
analyzing the distribution of the label.

Another family of Na+ channels with a different structure has been found in the apical
membranes of epithelial cells in the kidneys, colon, lungs, and brain. Those epithelial
sodium channels (ENaCs) are made up of three subunits encoded by three different
genes. Each of the subunits probably spans the membrane twice, and the amino terminal
and carboxyl terminal are located inside the cell. The α subunit transports Na +, whereas
the β and γ subunits do not. However, the addition of the β and γ subunits increases Na +
transport through the α subunit. ENaCs are inhibited by the diuretic amiloride, which
binds to the α subunit, and they used to be called amiloride-inhibitable Na+
channels. The ENaCs in the kidney play an important role in the regulation of ECF
volume by aldosterone (see Chapter 38). ENaC knockout mice are born alive but
promptly die because they cannot pump Na+ and hence water out of their lungs.

There are several types of Cl- channels in humans. The CLC dimeric channels described
above (Figure 1-29) are found in plants, bacteria, and animals, and there are nine
different CLC genes in humans. Other Cl - channels have the same pentameric form as the
acetylcholine receptor; examples include the GABA A and glycine receptors in the CNS
(see Chapter 4). The CFTR receptor that is mutated in cystic fibrosis is also a Cl - channel.
Ion channel mutations cause a variety of channelopathies—diseases that mostly affect
muscle and brain tissue and produce episodic paralyses or convulsions.

Na+-K+ ATPase

As noted above, Na+-K+ ATPase catalyzes the hydrolysis of ATP to ADP and uses the
energy to extrude three Na+ from the cell and take two K + into the cell for each mole of
ATP hydrolyzed. It is an electrogenic pump in that it moves three positive charges out
of the cell for each two that it moves in, and it is therefore said to have a coupling ratio
of 3:2. It is found in all parts of the body. Its activity is inhibited by ouabain and related
digitalis glycosides used in the treatment of heart failure. It is a heterodimer made up of
an α subunit with a molecular weight of approximately 100,000 and a β subunit with a
molecular weight of approximately 55,000. Both extend through the cell membrane
(Figure 1-32). Separation of the subunits eliminates activity. However, the β subunit is a
glycoprotein, whereas Na+ and K+ transport occur through the α subunit. The β subunit
has a single membrane-spanning domain and three extracellular glycosylation sites, all
of which appear to have attached carbohydrate residues. These residues account for one-
third of its molecular weight. The α subunit probably spans the cell membrane ten times,
with the amino and carboxyl terminals both located intracellularly. This subunit has
intracellular Na+- and ATP-binding sites and a phosphorylation site; it also has
extracellular binding sites for K + and ouabain. When Na+ binds to the α subunit, ATP
also binds and is converted to ADP, with a phosphate being transferred to Asp 376, the
phosphorylation site. This causes a change in the configuration of the protein, extruding
Na+ into the ECF. K+ then binds extracellularly, dephosphorylating the α subunit, which
returns to its previous conformation, releasing K+ into the cytoplasm.
The α and β subunits are heterogeneous, with α1, α2, and α3 subunits and β1, β2, and β3
subunits described so far. The α1 isoform is found in the membranes of most cells,
whereas α2 is present in muscle, heart, adipose tissue, and brain, and α 3 is present in
heart and brain. The β1 subunit is widely distributed but is absent in certain astrocytes,
vestibular cells of the inner ear, and glycolytic fast-twitch muscles. The fast-twitch
muscles contain only β2 subunits. The different α and β subunit structures of Na +-K+
ATPase in various tissues probably represents specialization for specific tissue functions.

Regulation of Na+-K+ ATPase Activity

The amount of Na+ normally found in cells is not saturating, so if it increases, the pump
activity and hence the amount of Na+ extruded from the cell increase. Pump activity is
affected by second messengers produced in cells, including cAMP and diacylglycerol
(DAG; see below); the magnitude and direction of the observed effects vary with the
experimental conditions. Thyroid hormones increase pump activity by a genomic action
to increase the formation of Na +-K+ ATPase molecules. Aldosterone also increases the
number of pumps, although this effect is probably secondary (see Chapters 20 and 38).
Dopamine in the kidney inhibits the pump by phosphorylating it, causing a natriuresis.
Insulin increases pump activity, probably by a variety of different mechanisms. Finally,
Na+-K+ ATPase is anchored in the membrane by the cytoskeleton, and it is interesting
that G-actin (see above) increases pump activity.

Secondary Active Transport

In many situations, the active transport of Na + is coupled to the transport of other
substances (secondary active transport). For example, the luminal membranes of
mucosal cells in the small intestine contain a symport that transports glucose into the
cell only if Na+ binds to the protein and is transported into the cell at the same time.
From the cells, the glucose enters the blood. The electrochemical gradient for Na + is
maintained by the active transport of Na + out of the mucosal cell into ECF (see Chapter
25). Other examples are shown in Figure 1-33. In the heart, Na+-K+ ATPase indirectly
affects Ca2+ transport. An antiport in the membranes of cardiac muscle cells normally
exchanges intracellular Ca2+ for extracellular Na+. The rate of this exchange is
proportionate to the concentration gradient for Na + across the cell membrane. If the
operation of the Na+-K+ ATPase is inhibited (eg, by ouabain), the intracellular Na +
concentration increases, the Na+ gradient across the cell membrane decreases, and Ca2+
extrusion decreases. The resulting increase in intracellular Ca 2+ concentration facilitates
the contraction of cardiac muscle (positively inotropic effect; see Chapter 3).
Active transport of Na+ and K+ is one of the major energy-using processes in the
body. On the average, it accounts for about 24% of the energy utilized by cells, and in
neurons it accounts for 70%. Thus, it accounts for a large part of the basal metabolism.

Transport Across Epithelia

In the gastrointestinal tract, the pulmonary airways, the renal tubules, and other
structures, substances enter one side of a cell and exit another, producing movement of
the substance from one side of the epithelium to the other. For transepithelial transport
to occur, the cells need to be bound by tight junctions and, obviously, have different ion
channels and transport proteins in different parts of their membranes. Most of the
instances of secondary active transport cited in the preceding paragraph involve
transepithelial movement of ions and other molecules.
THE CAPILLARY WALL

Filtration

The capillary wall separating plasma from interstitial fluid is different from the cell
membranes separating interstitial fluid from intracellular fluid because the pressure
difference across it makes filtration a significant factor in producing movement of
water and solute. By definition, filtration is the process by which fluid is forced through a
membrane or other barrier because of a difference in pressure on the two sides.

Oncotic Pressure
The structure of the capillary wall varies from one vascular bed to another (see
Chapter 30). However, in skeletal muscle and many other organs, water and relatively
small solutes are the only substances that cross the wall with ease. The apertures in the
junctions between the endothelial cells are too small to permit plasma proteins and other
colloids to pass through in significant quantities. The colloids have a high molecular
weight but are present in large amounts. Small amounts cross the capillary wall by
vesicular transport (see below), but their effect is slight. Therefore, the capillary wall
behaves like a membrane impermeable to colloids, and these exert an osmotic pressure
of about 25 mm Hg. The colloid osmotic pressure due to the plasma colloids is called the
oncotic pressure. Filtration across the capillary membrane as a result of the
hydrostatic pressure head in the vascular system is opposed by the oncotic pressure. The
way the balance between the hydrostatic and oncotic pressures controls exchanges across
the capillary wall is considered in detail in Chapter 30.

Transcytosis

There are vesicles in the cytoplasm of endothelial cells, and tagged protein molecules
injected into the bloodstream have been found in the vesicles and in the interstitium.
This indicates that small amounts of protein are transported out of capillaries across
endothelial cells by endocytosis on the capillary side followed by exocytosis on the
interstitial side of the cells. The transport mechanism makes use of coated vesicles that
appear to be coated with caveolin and is called transcytosis, vesicular transport, or
cytopempsis.

INTERCELLULAR COMMUNICATION

Cells communicate with each other via chemical messengers. Within a given tissue, some
messengers move from cell to cell via gap junctions (see above) without entering the
ECF. In addition, cells are affected by chemical messengers secreted into the ECF. These
chemical messengers bind to protein receptors on the surface of the cell or, in some
instances, in the cytoplasm or the nucleus, triggering sequences of intracellular changes
that produce their physiologic effects. There are three general types of intercellular
communication mediated by messengers in the ECF: (1) neural communication, in
which neurotransmitters are released at synaptic junctions from nerve cells and act
across a narrow synaptic cleft on a postsynaptic cell (see Chapter 4); (2) endocrine
communication, in which hormones and growth factors reach cells via the circulating
blood (see Chapters 18-24); and (3) paracrine communication, in which the
products of cells diffuse in the ECF to affect neighboring cells that may be some distance
away (Figure 1-34). In addition, cells secrete chemical messengers that in some
situations bind to receptors on the same cell, ie, the cell that secreted the messenger
(autocrine communication). The chemical messengers include amines, amino acids,
steroids, polypeptides, and in some instances lipids, purine nucleotides, and pyrimidine
nucleotides. It is worth noting that in various parts of the body, the same chemical
messenger can function as a neurotransmitter, a paracrine mediator, a hormone secreted
by neurons into the blood (neural hormone), and a hormone secreted by gland cells into
the blood.

An additional form of intercellular communication is called juxtacrine

communication. Some cells express multiple repeats of growth factors such as
transforming growth factor alpha (TGFα) extracellularly on transmembrane
proteins that provide an anchor to the cell. Other cells have TGFα receptors.
Consequently, TGFα anchored to a cell can bind to a TGFα receptor on another cell,
linking the two. This could be important in producing local foci of growth in tissues.

Radioimmunoassay

Antibodies to the polypeptides and proteins are readily produced, and, by using special
techniques, it is possible to make antibodies to the other chemical messengers as well.
The antibodies can be used to measure the messengers in body fluids and in tissue
extracts by radioimmunoassay. This technique depends on the fact that the naturally
occurring, unlabeled ligand and added radioactive ligand compete to bind to an antibody
to the ligand. The greater the amount of unlabeled ligand in the specimen being
analyzed, the more it competes and the smaller the amount of radioactive ligand that
binds to the antibody. Radioimmunoassays are extensively used in research and in
clinical medicine.

Receptors for Hormones, Neurotransmitters, & Other Ligands

Many of the receptors for chemical messengers have now been isolated and
characterized. These proteins are not static components of the cell, but their numbers
increase and decrease in response to various stimuli, and their properties change with
changes in physiologic conditions. When a hormone or neurotransmitter is present in
excess, the number of active receptors generally decreases (down-regulation),
whereas in the presence of a deficiency of the chemical messenger, there is an increase in
the number of active receptors (up-regulation). Angiotensin II in its actions on the
adrenal cortex is an exception; it increases rather than decreases the number of its
receptors in the adrenal. In the case of receptors in the membrane, receptor-mediated
endocytosis is responsible for down-regulation in some instances; ligands bind to their
receptors, and the ligand-receptor complexes move laterally in the membrane to coated
pits, where they are taken into the cell by endocytosis (internalization). This
decreases the number of receptors in the membrane. Some receptors are recycled after
internalization, whereas others are replaced by de novo synthesis in the cell. Another
type of down-regulation is desensitization, in which receptors are chemically modified in
ways that make them less responsive (see Chapter 4).

Mechanisms by Which Chemical Messengers Act

The principal mechanisms by which chemical messengers exert their intracellular effects
are summarized in Table 1-6. Ligands such as acetylcholine bind directly to ion channels
in the cell membrane, changing their conductance. Thyroid and steroid hormones, 1,25-
dihydroxycholecalciferol, and retinoids enter cells and act on one or another member of
a family of structurally related cytoplasmic or nuclear receptors. The activated receptor
binds to DNA and increases transcription of selected mRNAs. Many other ligands in the
ECF bind to receptors on the surface of cells, and many of them trigger the release of
intracellular mediators such as cAMP, IP 3, and DAG (see below) that initiate changes in
cell function. Consequently, the extracellular ligands are called "first messengers" and
the intracellular mediators are called "second messengers."

Second messengers bring about many short-term changes in cell function by altering
enzyme function, triggering exocytosis, etc, but they also alter transcription of various
genes. They do this in part by activating transcription factors already present in the cell,
and these activated factors induce the transcription of immediate-early genes (Figure 1-
20). The transcription factors that are the products of the immediate-early genes then
activate other genes which produce more long-term effects.

When activated, many of the membrane receptors initiate release of second messengers
or other intracellular events via GTP-binding proteins (G proteins; see below). The
second messengers generally activate protein kinases, enzymes that catalyze the
phosphorylation of tyrosine or serine and threonine residues in proteins. More than 300
protein kinases have been described. Some of the principal ones that are important in
mammals are summarized in Table 1-7. Addition of phosphate groups changes the
configuration of the proteins, altering their functions and consequently the functions of
the cell. In some instances, eg, the insulin receptor, the intracellular portions of the
receptors themselves are protein kinases, and in some instances, they phosphorylate
themselves (autophosphorylation). Other receptors, eg, cytokine receptors, are not
protein kinases themselves but readily initiate phosphorylation of many intracellular
proteins. Obviously, phosphatases are also important, since removal of a phosphate
group inactivates some transport proteins or enzymes whereas it activates others.

Stimulation of Transcription

When thyroid and steroid hormones, 1,25-dihydroxycholecalciferol, and retinoids bind

to their receptors inside cells, the conformation of the receptor protein is changed and a
DNA-binding domain is exposed (Figure 1-35). The receptor-hormone complex moves to
DNA, where it binds to enhancer elements in the untranslated 5´-flanking portions of
certain genes. The estrogen and the triiodothyronine (T 3) receptors bind hormones in the
nucleus. The T3 receptors also bind thyroxine (T4), but with less affinity. The
glucocorticoid receptor is located mainly in the cytoplasm but migrates promptly to the
nucleus as soon as it binds its ligand. The initial location of the other receptors that act in
this fashion is unsettled. In any case, binding of the receptor-hormone complex to DNA
increases the transcription of mRNAs encoded by the gene to which it binds. The mRNAs
are translated in the ribosomes, with the production of increased quantities of proteins
that alter cell function.

At least for the glucocorticoid, estrogen, and progesterone receptors, the receptor is
bound to the heat shock protein Hsp90 and other proteins in the absence of the
steroid, and it appears that the heat shock protein covers the DNA-binding domain.
When the steroid binds to the receptor, the conformation change releases the heat shock
protein, exposing the DNA-binding domain.

Heat shock proteins are a group of intracellular proteins whose amounts increase when
cells are exposed to heat and other stresses, and they help the cells survive a variety of
stresses. Consequently, it is probably more appropriate to call them stress proteins.

Structure of Receptors

The structures of the human glucocorticoid and mineralocorticoid receptors are shown
in Figure 1-36. Two estrogen receptors (α and β) and two T 3 receptors (α and β) have
been identified; the α estrogen receptor and the β T 3 receptor are shown in the figure. All
these receptors are part of a superfamily of receptors that have in common a highly
conserved cysteine-rich DNA-binding domain; a ligand-binding domain at or near the
carboxyl terminal of the receptor; and a relatively variable, poorly conserved amino
terminal region. When a ligand binds to one of them, it becomes a transcription factor
and binds to DNA via zinc fingers (see above). Other receptors in the family include the
receptors for progesterone, androgen, and 1,25-dihydroxycholecalciferol. Many other
factors that regulate genes act via receptors of this type in species ranging from fruit flies
to humans, and over 70 members of this receptor superfamily have been described.
Ligands are now known for about half of these, but the remaining half are orphan
receptors, for which the ligands are still unidentified. Retinoic acid, which is a
derivative of retinol (vitamin A), has an extensive role in fetal development, and there
are three retinoic acid receptors, α, β, and γ, encoded by two families of retinoic acid
receptors, RAR and RXR, each with α, β, and γ forms. T 3 receptors form homodimers
before binding to DNA, but heterodimers with retinoic receptors also form and bind, and
their actions are complex (see Chapter 18).

Rapid Actions of Steroids

Some of the actions of steroids are much more rapid than those known to be mediated
via binding to DNA. Examples include the rapid increase in the Ca 2+ concentration in
sperm heads that is produced by progesterone and prompt steroid-induced alteration in
the functions of various neurons. This has led to the hypothesis that there are
nongenomic actions of steroids which are mediated by putative membrane receptors
and second messengers inside the cells (see below). There is molecular biologic evidence
for the existence of these receptors, though detailed information about them is still
lacking. Steroids also bind to GABAA receptors, facilitating their action (see Chapter 4).

Intracellular Ca2+

Ca2+ regulates a very large number of physiologic processes that are as diverse as
proliferation, neural signaling, learning, contraction, secretion, and fertilization, so
regulation of intracellular Ca2+ is of great importance. The free Ca 2+ concentration in the
cytoplasm at rest is maintained at about 100 nmol/L. The Ca 2+ concentration in the
interstitial fluid is about 12,000 times the cytoplasmic concentration, ie, 1,200,000
nmol/L, so there is a marked inwardly directed concentration gradient as well as an
inwardly directed electrical gradient. Much of the intracellular Ca2+ is bound by the
endoplasmic reticulum and other organelles (Figure 1-37), and these organelles provide a
store from which Ca2+ can be mobilized via ligand-gated channels to increase the
concentration of free Ca2+ in the cytoplasm. Increased cytoplasmic Ca 2+ binds to and
activates calcium-binding proteins, and these in turn activate a number of protein
kinases.

Ca2+ enters cells through many different Ca2+ channels. Some of these are ligand-gated
and others are voltage-gated. There appear to be stretch-activated channels as well. The
voltage-gated Ca2+ channels are often divided into T (transient) or L (long-lasting) types
depending on whether they do or do not inactivate during maintained depolarization.

Ca2+ is pumped out of cells in exchange for two H+ by a Ca2+-H+ ATPase, and it is
transported out of cells by an antiport driven by the Na + gradient that exchanges three
Na+ for each Ca2+.

Many second messengers act by increasing the cytoplasmic Ca 2+ concentration. The

increase is produced by releasing Ca2+ from intracellular stores—primarily the
endoplasmic reticulum—or by increasing the entry of Ca 2+ into cells, or by both
mechanisms. IP3 is the major second messenger that causes Ca 2+ release from the
endoplasmic reticulum. In many tissues, transient release of Ca 2+ from internal stores
into the cytoplasm triggers opening of a population of Ca 2+ channels in the cell
membrane (store-operated Ca2+ channels; SOCCs). The resulting Ca2+ influx
replenishes the intracellular Ca2+ supply and refills the endoplasmic reticulum. The exact
identity of the SOCCs is still unknown, and there is debate about the signal from the
endoplasmic reticulum that opens them. However, evidence is accumulating that IP 3 is
responsible for both the internal release from the endoplasmic reticulum and the
activation of the SOCCs.

Calcium-Binding Proteins

Many different Ca2+-binding proteins have been described, including troponin,

calmodulin, and calbindin. Troponin is the Ca2+-binding protein involved in
contraction of skeletal muscle (see Chapter 3). Calmodulin contains 148 amino acid
residues (Figure 1-38) and has four Ca2+-binding domains. It is unique in that residue 115
is trimethylated, and it is extensively conserved, being found in plants as well as animals.
When calmodulin binds Ca2+, it is capable of activating five different calmodulin-
dependent kinases (Table 1-7). One of these is myosin light-chain kinase, which
phosphorylates myosin. This brings about contraction in smooth muscle. Another is
phosphorylase kinase, which activates phosphorylase (see Chapter 17).
Ca2+/calmodulin kinases I and II are concerned with synaptic function, and
Ca2+/calmodulin kinase III is concerned with protein synthesis. Another calmodulin-
activated protein is calcineurin, a phosphatase that inactivates Ca2+ channels by
dephosphorylating them. It also plays a role in activating T cells and is inhibited by some
immunosuppressants (see Chapter 27).

Mechanisms of Diversity of Ca2+ Actions

It may seem difficult to understand how intracellular Ca 2+ can have so many varied
effects as a second messenger. Part of the explanation is that Ca 2+ may have different
effects at low and at high concentrations. The ion may be in high concentration at the site
of its release from an organelle or a channel (Ca2+ sparks) and at a subsequent lower
concentration after it diffuses throughout the cell. Some of the changes it produces can
outlast the rise in intracellular Ca2+ concentration because of the way it binds to some of
the Ca2+-binding proteins. In addition, once released, intracellular Ca 2+ concentrations
frequently oscillate at regular intervals, and there is evidence that the frequency and, to a
lesser extent, the amplitude of those oscillations codes information for effector
mechanisms. Finally, increases in intracellular Ca2+ concentration can spread from cell to
cell in waves, producing coordinated events such as the rhythmic beating of cilia in
epithelial tissue.
G Proteins

A common way to translate a signal to a biologic effect inside cells is by way of nucleotide
regulatory proteins (G proteins) that bind GTP. GTP is the guanosine analog of ATP
(see Chapter 17). When the signal reaches a G protein, the protein exchanges GDP for
GTP. The GTP-protein complex brings about the effect. The inherent GTPase activity of
the protein then converts GTP to GDP, restoring the resting state. The GTPase activity is
accelerated by a family of RGS (regulators of G protein signaling) proteins that accelerate
the formation of GDP.

Small G proteins are involved in many cellular functions. Members of the Rab family
of these proteins regulate the rate of vesicle traffic between the endoplasmic reticulum,
the Golgi apparatus, lysosomes, endosomes, and the cell membrane (see above). Another
family of small GTP-binding proteins, the Rho/Rac family, mediates interactions
between the cytoskeleton and cell membrane, and a third family, the Ras family,
regulates growth by transmitting signals from the cell membrane to the nucleus. The
members of these three families are related to the product of the ras proto-oncogene.

Another family of G proteins, the larger heterotrimeric G proteins, couple cell

surface receptors to catalytic units that catalyze the intracellular formation of second
messengers or couple the receptors directly to ion channels. These G proteins are made
up of three subunits designated α, β, and γ (Figure 1-39). The α subunit is bound to GDP.
When a ligand binds to a G-coupled receptor, this GDP is exchanged for GTP and the α
subunit separates from the combined β and γ subunits. The separated α subunit brings
about many biologic effects. The β and γ subunits do not separate from each other, and
βγ also activates a variety of effectors. The intrinsic GTPase activity of the α subunit then
converts GTP to GDP, and this leads to reassociation of the α with the βγ subunit and
termination of effector activation.
Heterotrimeric G proteins relay signals from over 1000 receptors, and their
effectors in the cells include ion channels and enzymes. Examples are listed in Table 1-8.
There are 16 α, 6 β, and 12 γ genes, so a large number of subunits are produced, and they
can combine in various ways. They can be divided into five families, each with a
relatively characteristic set of effectors. The families are G s, Gi, Gt, Gq, and G13.

Many G proteins are modified by having specific lipids attached to them, ie, they are
lipidated (Figure 1-6). Trimeric G proteins may be myristolated, palmitoylated, or
prenylated. Small G proteins may be prenylated.

Serpentine Receptors
All the heterotrimeric G protein-coupled receptors that have been characterized
to date are proteins that span the cell membrane seven times (serpentine receptors).
These receptors may be palmitoylated. A very large number have been cloned, and their
functions are multiple and diverse. The structures of two of them are shown in Figure 1-
40. In general, small ligands bind to the amino acid residues in the membrane, whereas
large polypeptide and protein ligands bind to the extracellular domains, which are bigger
and better developed in the receptors for polypeptides and proteins. It is generally amino
acid residues in the third cytoplasmic loop, the loop nearest the carboxyl terminal, that
interact with the G proteins.

Inositol Triphosphate & Diacylglycerol as Second Messengers

The link between membrane binding of a ligand that acts via Ca 2+ and the prompt
increase in the cytoplasmic Ca2+ concentration is often inositol triphosphate
(inositol 1,4,5-triphosphate; IP3). When one of these ligands binds to its receptor,
activation of the receptor produces activation of phospholipase C on the inner surface of
the membrane via Gq. There are at least eight isoforms of phospholipase C (PLC), and the
PLCβ1 and PLCβ2 forms are activated by G proteins. They catalyze the hydrolysis of
phosphatidylinositol 4,5-diphosphate (PIP2) to form IP3 and diacylglycerol (DAG)
(Figure 1-41). Tyrosine kinase-linked receptors (see below) can also produce IP 3 and
DAG by activating PLCγ1. The IP3 diffuses to the endoplasmic reticulum, where it triggers
the release of Ca2+ into the cytoplasm (Figure 1-42). The IP3 receptor resembles the
ryanodine receptor which is the Ca 2+ channel in the sarcoplasmic reticulum of skeletal
muscle (see Chapter 3), except that the IP3 receptor is half as large. DAG is also a second
messenger; it stays in the cell membrane, where it activates one of the seven subspecies
of protein kinase C (Table 1-7). Examples of ligands that act via these second
messengers are listed in Table 1-8.
The precursor of PIP2 is phosphatidylinositol (Figure 1-41). This phospholipid is
found in relatively small amounts in the inner lamella of the cell membrane. It is first
converted to phosphatidyl 4-phosphate (PIP) and then to PIP 2, the derivative that is
hydrolyzed to form IP3 and the DAG. Other inositol phosphates are also formed in cells,
but their function is uncertain. The IP3 is metabolized by stepwise dephosphorylation to
inositol. DAG is converted to phosphatidic acid and then to cytosine diphosphate (CDP)
diacylglycerol, which combines with inositol to form phosphatidylinositol, completing
the cycle.

Cyclic AMP
Another important second messenger is cyclic AMP (cAMP) (Figure 1-43).
Some of the many ligands that act via this compound are listed in Table 1-6. Cyclic AMP
is cyclic adenosine 3´,5´-monophosphate. It is formed from ATP by the action of the
enzyme adenylyl cyclase and converted to physiologically inactive 5´-AMP by the
action of the enzyme phosphodiesterase. Cyclic AMP activates one of the cyclic
nucleotide-dependent protein kinases (protein kinase A) that, like protein kinase C,
catalyzes the phosphorylation of proteins, changing their conformation and altering their
activity. A typical example is the activation of phosphorylase kinase in the liver by
epinephrine via cAMP and protein kinase A (see Figure 17-13). In addition, the active
catalytic subunit of PKA moves to the nucleus and phosphorylates the cAMP-
responsive element-binding protein (CREB) This transcription factor then binds
to DNA and alters transcription of a number of genes.
Cyclic AMP is metabolized by a phosphodiesterase. This phosphodiesterase is
inhibited by methylxanthines such as caffeine and theophylline; consequently, these
compounds augment hormonal and transmitter effects mediated via cAMP.

Activation of Adenylyl Cyclase

Five components are involved in the mechanism by which ligands bring about
changes in the intracellular concentration of cAMP: a catalytic unit, adenylyl cyclase,
which catalyzes the conversion of ATP to cAMP; stimulatory and inhibitory receptors;
and stimulatory and inhibitory G proteins that link the receptor to the catalytic unit
(Figure 1-44). Like the receptors, adenylyl cyclase is a transmembrane protein, and it
crosses the membrane 12 times. Eight isoforms of this enzyme have been described, and,
combined with the many different forms of G proteins, this permits the cAMP pathway
to be customized to specific tissue needs. When the appropriate ligand binds to a
stimulatory receptor, a Gs α subunit activates one of the adenylyl cyclases. Conversely,
when the appropriate ligand binds to the inhibitory receptor, a G i α subunit inhibits
adenylyl cyclase. The receptors are specific, responding at low threshold to only one or a
select group of related ligands. However, heterotrimeric G proteins mediate the
stimulatory and inhibitory effects produced by many different ligands. In addition, there
is cross-talk between the phospholipase C system and the adenylyl cyclase system, and
several of the isoforms of adenylyl cyclase are stimulated by calmodulin. Finally, the
effects of protein kinase A and protein kinase C are very widespread. Given this
complexity, how are specific responses to specific stimuli obtained? The answer lies in
part in tethering of the G proteins, adenylyl cyclase, and the protein kinases to the
cytoskeleton so that local microdomains are created. Some of this tethering is carried out
by lipid products (Figure 1-6).
Some cAMP escapes from cells upon stimulation by certain hormones, but the
amounts are small compared with the intracellular concentration, and only small
amounts of extracellular cAMP enter cells.
Two bacterial toxins have important effects on adenylyl cyclase that are mediated
by G proteins. The A subunit of cholera toxin catalyzes the transfer of ADP-ribose to
an arginine residue in the middle of the α subunit of G s. This inhibits its GTPase activity,
producing prolonged stimulation of adenylyl cyclase (see Chapter 25). Pertussis toxin
catalyzes ADP-ribosylation of a cysteine residue near the carboxyl terminal of the α
subunit of Gi. This inhibits the function of G i. In addition to the implications of these
alterations in disease, both toxins are used for fundamental research on G protein
function. The drug forskolin stimulates adenylyl cyclase activity by a direct action on the
enzyme.

Guanylyl Cyclase
Another cyclic nucleotide of physiologic importance is cyclic guanosine
monophosphate (cyclic GMP; cGMP). Cyclic GMP is important in vision. Light acts on
rhodopsin in rods; rhodopsin is linked to phosphodiesterase by Gtl; and activation of
phosphodiesterase accelerates conversion of cGMP to 5´-GMP (see Chapter 8). A similar
process occurs in the cones. In addition, there are cGMP-regulated ion channels, and
cGMP activates cGMP-dependent kinase (Table 1-7), producing a number of physiologic
effects.

Guanylyl cyclases are a family of enzymes that catalyze the formation of cGMP. They
exist in two forms (Figure 1-45). In one form, there is an extracellular amino terminal
domain that is a receptor, single transmembrane domain, and a cytoplasmic carboxyl
terminal that has a tyrosine kinase-like and a guanylyl cyclase catalytic domain. Three
such guanylyl cyclases have been characterized. Two are receptors for ANP (ANPR-A and
ANPR-B; see Chapter 24), and a third binds an Escherichia coli enterotoxin and the
gastrointestinal polypeptide guanylin (see Chapter 26). The other form of guanylyl
cyclase is soluble, contains heme, and is totally intracellular. There appear to be several
isoforms of the intracellular enzyme. They are activated by nitric oxide (NO) and NO-
containing compounds. NO is the endothelium-derived relaxing factor (EDRF), an
intercellular messenger formed from arginine, that plays an important role in regulating
the diameter of blood vessels (see Chapter 31). NO is also involved in many other
functions, including penile erection (see Chapter 23) and synaptic transmission in the
brain (see Chapter 4). Recent evidence indicates that in addition to its effects mediated
by cGMP, NO can activate Ca2+-dependent K+ channels by a direct action on the
channels.
Phosphatases
Numerous phosphatases that remove phosphate groups from proteins are found in cells.
Frequently these are closely associated with or coupled to tyrosine kinases and serine-
threonine kinases. Two examples are shown in Figure 1-45.

Growth Factors
Growth factors have become increasingly important in many different aspects of
physiology. They are polypeptides and proteins that are conveniently divided into three
groups. One group is made up of agents that foster the multiplication or development of
various types of cells; nerve growth factor (see Chapter 2), insulin-like growth factor I
(IGF-I; see Chapter 22), activins and inhibins (see Chapter 23), and epidermal growth
factor (EGF) are examples, and more than 20 have been described. The cytokines are a
second group. These factors are produced by macrophages and lymphocytes and are
important in regulation of the immune system (see Chapter 27). Again, more than 20
have been described. The third group is made up of the colony-stimulating factors that
regulate proliferation and maturation of red and white blood cells. They are also
discussed in Chapter 27.
Receptors for EGF, platelet-derived growth factor (PDGF), and many of the other
factors that foster cell multiplication and growth have a single membrane-spanning
domain with an intracellular tyrosine kinase domain (Figure 1-46). When ligand binds to
the receptor, the tyrosine kinase domain autophosphorylates itself. Some of the
receptors dimerize when they bind their ligands, and the intracellular tyrosine kinase
domains cross-phosphorylate each other. One of the pathways activated by
phosphorylation leads, through the product of the ras proto-oncogene and several MAP
kinases, directly to the production of transcription factors in the nucleus that alter gene
expression. This important direct path from the cell surface to the nucleus is shown
diagrammatically in Figure 1-46. Note that Ras is one of the small G proteins that
requires binding to GTP for activation.
Receptors for the cytokines and the colony-stimulating factors differ from the
other growth factors in that most of them do not have tyrosine kinase domains in their
cytoplasmic portions and some have little or no cytoplasmic tail. However, they initiate
tyrosine kinase activity in the cytoplasm. In some instances, this involves binding to the
associated transmembrane protein gp130 (see Chapter 27). In particular, they activate
the so-called Janus tyrosine kinases (JAKs) in the cytoplasm (Figure 1-47). These in
turn phosphorylate signal transducer and activator of transcription (STAT) proteins.
The phosphorylated STATs form homo- and heterodimers and move to the nucleus,
where they act as transcription factors. There are four known mammalian JAKs and
seven known STATs. The JAK-STAT pathway is also activated by growth hormone (see
Figure 22-4) and is another important direct path from the cell surface to the nucleus.
However, it should be emphasized that both the Ras and the JAK-STAT pathways are
complex and there is cross talk between them and the phospholipase C and cAMP
pathways.

Another family of receptors binds transforming growth factor β (TGFβ) and related
polypeptides. These receptors have serine-threonine kinase activity, and their effects are
mediated by SMADs, intracellular proteins that when phosphorylated move to the
nucleus, bind to DNA, and, with other factors, initiate transcription of various genes.

As noted above, integrins also initiate phosphorylation of proteins that enter the nucleus
and alter gene transcription.
 Note that a common theme is activation of transcription factors that without
activation are "locked" in the cytoplasm. Once activated, the transcription factor moves
to the nucleus and alters gene transcription. Additional examples include NF-AT (see
Figure 27-13) and NF-κB (see Chapters 20 and 33).

Receptor & G Protein Diseases

Many diseases are being traced to mutations of the genes for receptors. For
example, loss-of-function receptor mutations that cause disease have been reported for
the 1,25-dihydroxycholecalciferol receptor (see Chapter 21) and the insulin receptor (see
Chapter 19). Certain other diseases are caused by production of antibodies against
receptors. Thus, antibodies against TSH receptors cause Graves' disease (see Chapter
18), and antibodies against nicotinic acetylcholine receptors cause myasthenia gravis
(see Chapter 4).
An example of loss of function of a receptor is the type of nephrogenic
diabetes insipidus that is due to loss of the ability of mutated V 2 vasopressin receptors
to mediate concentration of the urine (see Chapters 14 and 38). Mutant receptors can
gain as well as lose function. A gain-in-function mutation of the Ca 2+ receptor (see
Chapter 21) causes excess inhibition of parathyroid hormone secretion and familial
hypercalciuric hypocalcemia. G proteins can also undergo loss-of-function or gain-
of-function mutations that cause disease (Table 1-9). In one form of
pseudohypoparathyroidism, a mutated Gs α fails to respond to parathyroid hormone,
producing the symptoms of hypoparathyroidism without any decline in circulating
parathyroid hormone. Testotoxicosis is an interesting disease that combines gain and
loss of function. In this condition, an activating mutation of G s α causes excess
testosterone secretion and prepubertal sexual maturation. However, this mutation is
temperature-sensitive and is active only at the relatively low temperature of the testes
(33 °C; see Chapter 23). At 37 °C, the normal temperature of the rest of the body, it is
replaced by loss of function, with the production of hypoparathyroidism and decreased
responsiveness to TSH. A different activating mutation in G s α is associated with the
rough-bordered areas of skin pigmentation and hypercortisolism in the McCune-
Albright syndrome. This mutation occurs during fetal development, creating a mosaic of
normal and abnormal cells. A third mutation in G s α reduces its intrinsic GTPase activity.
As a result, it is much more active than normal, and excess cAMP is produced. This
causes hyperplasia and eventually neoplasia in somatotrope cells of the anterior
pituitary. Forty percent of somatotrope tumors causing acromegaly (see Chapter 22)
have cells containing a somatic mutation of this type.

3. pertahanan sel :
 agen pertahanan kimiawi dan fisik

 antioksidan (primer, skunder, tersier), sitokin anti inflamasi

Antioksidan
adalah senyawa yang mempunyai struktur molekul yang dapat memberikan elektronnya
dengan cuma-cuma kapada molekul radikal bebas tanpa terganggu sama sekali dan
dapat memutus reaksi berantai dari radikal bebas.
Terdapat tiga macam antioksidan yaitu:
1). Antioksidan yang dibuat oleh tubuh kita sendiri yang berupa enzim antara lain
superoksida dismutase, glutathione peroxidase, peroxidasi dan katalase.
2). Antioksidan alami yang dapat diperoleh dari tanaman atau hewan yaitu tokoferol,
vitamin C, betakaroten, flavonoid dan senyawa fenolik.
3). Antioksidan sintetik, yang dibuat dari bahn-bahan kimia yaitu Butylated
Hroxyanisole (BHA), BHT, TBHQ, PG dan NDGA yang ditambahkan dalam makanan
untuk mencegah kerusakan lemak.

Atas dasar fungsinya antioksidan dapat dibedakan menjadi 4 (lima) yaitu :

A. Antioksidan Primer
Antioksidan ini berfungsi untuk mencegah terbentuknya radikal bebas baru karena ia
dapat merubah radikal bebas yang ada menjadi molekul ynag berkurang dampak
negatifnya yaitu sebelum sempat bereaksi. Antioksidan primer yang ada dalam tubuh
yang sangat terkenal adalah enzim superoksida dismutase. Enzim ini sangat penting
sekali karena dapat melinduhngi hancrnya sel-sel dalam tubuh akibat serangan radikal
bebas. Bekerjanya enzim ini sangat idpengaruhi oleh mineral-mineral seperti mangan,
seng, tembaga dan selenium yang harus terdapat dalam makanan dan minuman.

B. Antioksidan Sekunder
Antioksidan sekunder merupakan senyawa yang berfungsi menangkap radikal bebas
serta mencegah terjadinya reaksi berantai sehingga tidak terjadi keursakan yang lebih
besar. Contoh yang populer, antioksidan sekunder adalah vitamin E, vitamin C, dan
betakaroten yang dapat diperoleh dari buah-buahan.

C. Antioksidan Tersier
Antioksidan tersier merupakan senyawa yang memperbaiki sel-sel dan jaringan yang
rusak karena serangan radikal bebas. Biasanya yang termasuk kelompok ini adalah jenis
enzim misalnya metionin sulfoksidan reduktase yang dapat memperbaiki DNA dalam
inti sel. Enzim tersebut bermanfaat untuk perbaikan DNA pada penderita kanker.

D. Oxygen Scavanger
Antioksidan yang termasuk oxygen scavanger mengikat oksigen sehingga tidak
mendukung reaksi oksidasi, misalnya vitamin C.

E. Chelators/Sequesstrants
Mengikat logam yang mampu mengkatalisis reaksi oksidasi misalnya asam sitrat dan
asam amino.

Tubuh dapat menghasilkan antioksdan yang berupa enzim yang aktif bila didukung
oleh nutrisi pendukung atau mineral yang disebut juga ko-faktor. Antioksidan yang
dihasilkan oleh tubuh antara lain adalah :

1. Superoksida Dismutase
Antioksidan ini merupakan enzim yang bekerja bila ada pembantunya yaitu berupa
mineral-mineral seperti tembaga, mangan yang bersumber pada kacang-kacangan, padi-
padian. kekurangan mineral dapat dilakukan dengan meminum multivitamin dan
suplemen mineral. contoh nya brokoli, bayam, sawi dan juga hasil-hasil olahan seperti
tempe.

2. Glutathione Peroksidase
Adalah enzim yang berperan aktif dalam menghilangkan H2O2 dalam tubuh dan
mempergunakannya untuk merubah glutathione (GSH) menjadi glutathine teroksidasi
(GSSG) Enzim ini menjaga konsentrasi oksigen akhir agar stabil dan tidak berubah
menjadi pro-oksidan. Makanan yang kaya glutahione adalah kubis, brokoli, asparagus,
alpukat dan kenari. Glutathione sangat penting sekali melindungi selaput-selaput sel.
Senyawa ini merupakan tripeptida yang terdiri dari asam amino glisin, asam glutamat
dan sistein.

3. Katalase
Enzim katalase di samping mendukung aktivitas enzim SOD juga dapat mengkatalisa
perubahan berbagai macam peroksida dan radikal bebas menjadi oksigen dan air.
Enzim-enzim tersebut di atas dalam bekerjanya sengat membutuhkan mineral-mineral
penyusun sebagai berikut :
• Copper (Cu)
• Zinc (Zn)
• Selenium (Se)
• Manganese (Mn)
• Besi (Fe)

Mekanisme Kerja Antioksidan

Antioksidan adalah bahan tambahan yang digunakan untuk melindungi komponen-

komponen terutama lemak dan minyak. Meskipun demikian antioksidan dapat pula
digunakan untuk melindungi komponen lain seperti vitamin dan pigmen, yang juga
banyak mengandung ikatan rangkap di dalam strukturnya
Mekanisme kerja antioksidan secara umum adalah menghambat oksidasi
lemak yang akan di ubah menjadi radikal bebas. Dalam proses melumpuhkan radikal
bebas, vitamin E menjadi pelopor diikuti oleh vitamin C dan dengan bantuan
senyawa glutathion, betakaroten, seng, mangan dan selenium akan
memudahkan pelumpuhan radikal bebas
.
Contoh antioksidan yaitu vitamin E, vitamin C, kelompok karetonoid (beta karoten,
likopen, dan lutein), serta kelompok flavonoid. Sedangkan contoh mineral antioksidan
yaitu selenium dan seng. Secara alami, antioksidan dapat diperoleh dari sayur dan buah
yang kita konsumsi setiap hari. Namun, bagaimana mekanisme kerjanya?

Vitamin
Vitamin antioksidan yang cukup terkenal adalah vitamin C dan E. Vitamin C mencegah
oksidasi pada molekul yang berbasis cairan, misalnya plasma darah dan mata.
Sedangkan vitamin E yang larut dalam lemak bekerja pada sel lipid dan sirkulasi
kolesterol.

Sebuah studi yang diterbitkan dalam New England Journal of Medicine menunjukkan
bahwa vitamin E dapat memperlambat gejala Azheimer. Journal of the American
Medical Association juga menyatakan bahwa vitamin E dapat mencegah penyakit
jantung koroner. Sedangkan menurut Journal Ophtalmology, vitamin E dapat
menurunkan risiko terjadinya katarak.
Cara kerja vitamin E sebagai antioksidan adalah dengan menyumbangkan elektron
kepada radikal bebas. Karena itu, vitamin E yang kaku akan berubah menjadi vitamin E
yang radikal. Untuk menjinakkannya, diperlukan vitamin C yang akhirnya akan
membuat vitamin C juga menjadi radikal. Di sinilah, glutation akan muncul untuk
menetralkan vitamin C.

Mineral
Jika vitamin C dan E bertindak sebagai antioksidan langsung, mineral sendiri akan
berperan sebagai komponen antioksidan tubuh (endogen). Selenium, misalnya,
merupakan komponen penting glutation peroksidase. Selenium juga bekerja secara
sinergis dengan vitamin E. Sebuah studi yang diterbitkan dalam jurnal The Lancet
menyatakan bahwa mereka yang kekurangan selenium akan lebih berisiko menderita
kanker dibandingkan mereka yang berkecukupan selenium.

External Anti-Inflammatory Signals

1. Anti-Inflammatory Cytokines
Endothelial Cells receive signals from humoral factors, inflammatory mediators,
and physical forces from both the circulation and the tissue. Several potential triggers
capable of inducing proinflammatory and prothrombotic cellular responses
have been identified; these include modified lipoproteins, proinflammatory
cytokines, chemokines, vasoactive peptides, neuropeptides, hyperglycemia
and advanced glycosylated end products, smoking, oxidative stress, and
others. Smooth Muscle Cells also are targets of these triggers. Yet the vascular
inflammatory responses and their temporal patterns can be regulated by anti-
inflammatory cytokines. Anti-inflammatory cytokines that exert inhibitory
effects on vascular cells include transforming growth factor-b (TGF-b),
interleukin-10 (IL-10), and IL-1 receptor antagonist (IL-1ra).

TGF-b
TGF-b family members are secreted in inactive complexes with a latency-associated
peptide (LAP), a protein derived from the N-terminal region of the TGF-b gene product.
Extracellular activation of these complexes is a critical step in the regulation of TGF-b
function in vivo. Cytokine activation of ECs increases TGF-b1 synthesis and activation of
latent TGF-b by the plasminogen/plasmin system.2 Active TGF-b is produced by ECs in
vitro when they are cocultured with pericytes or SMCs.3 Production of active TGF-b has
also been found in human arterial SMCs in culture.4 Active TGF-b is detectable in the
aortic wall of mice and is decreased in transgenic mice expressing apo(a) as a
consequence of apo(a) inhibition of the plasminogen/plasmin system.2 TGF-b was first
reported to be a deactivating factor of macrophages capable, for example, of suppressing
inducible nitric oxide synthase (iNOS) protein expression in macrophages.5 TGF-b also
has potent anti-inflammatory effects on vascular cells. TGF-b1 downregulates cytokine-
induced expression of E-selectin and vascular cellular adhesion molecule-1 (VCAM-1) in
ECs6,7 as well as VCAM-1 in SMCs.8 TGF-b1 significantly decreases monocyte
chemotactic protein-1 (MCP-1) expression in human umbilical vein ECs (HUVECs)
stimulated with tumor necrosis factor-a (TNF-a) or IL-1b but not with interferon-g (IFN-
g).9 The expression of TNF-a receptors seems to be downmodulated by TGF-b1.9
Furthermore, TGF-b1 inhibits the elaboration of IL-8 by TNF-activated ECs10 and
inhibits the IL-8–dependent migration of neutrophils through the activated endothelial
monolayer.10 TGF-b is able to restore endothelialdependent vasodilation impaired by
TNF-a.11 In addition, TGF-b suppresses iNOS induction in the vascular wall, leading to
the prevention of lipopolysaccharide (LPS) shock in the rat.12 TGF-b expressed by
vascular cells may also operate as a paracrine anti-inflammatory factor: glomerular
mesangial cells express TGF-b in an active form that inhibits the production of
proinflammatory cytokines by emigrated macrophages.13 Such cross-communications
between vascular cells and infiltrating macrophages may play an important role in the
recovery from the inflammatory process. The pleiotropic effects of TGF-b are mediated
from membrane to nucleus through distinct combinations of three types of cell-surface
receptors (types I, II, and III), types I and II being serine and threonine kinases and their
downstream effectors, known as Smad proteins.14 Smad-mediated effects result from a
competitive interaction between Smad proteins activated by TGF-b1 and NF-kB proteins
activated by proinflammatory stimuli. Smad proteins interact with the limited amount of
cAMP response element– binding protein (CREB)-binding protein (CBP) present in ECs,
therefore blocking the association of CBP with p65/NF-kB6 that is required for maximal
transcriptional NF-kB activity (Figure 1). This type of signaling mechanism may play an
important role in the immunomodulatory actions of this cytokine/ growth factor in the
cardiovascular system.6 Most of the anti-inflammatory effects of TGF-b on vascular cells
were documented in vitro. However, the relevance of in vitro findings to in vivo
conditions is substantiated by the observation that TGF-b1–deficient mice die in utero or
in the perinatal period because of widespread uncontrolled inflammation. 15 The TGF-b1
knockout mice have multifocal inflammatory disease in many tissues, but the heart and
lungs are most severely affected. Increased adhesion of leukocytes to the endothelium of
pulmonary veins and increased expression of major histocompatibility complex (MHC)
class I and II proteins are seen in pulmonary vascular endothelium as early as day 8.16
Mice heterozygous for the deletion of the TGF-b1 gene (TGF-b11/2 mice) show higher
levels of endothelial intercellular adhesion molecule-1 (ICAM-1) and VCAM-1 and
enhanced macrophage infiltration than the wild-type mice after 12 weeks of cholesterol-
enriched diet.17 These findings suggest that the presence of endogenous TGF-b1 in the
vessel wall is protective against vascular inflammation.

Pro- and Anti-Inflammatory Signals With Activities on Vascular Cells

IL-10
IL-10 is a pleiotropic cytokine produced by Th2-type T cells, B cells, monocytes, and
macrophages that inhibits a broad array of immune parameters, including Th1
lymphocyte cytokine production, antigen presentation, and antigenspecific T-cell
proliferation. IL-10 also has potent anti-inflammatory properties on macrophages. In
vitro experiments showed that the expression of IL-10 in LPS-stimulated monocytes is
delayed relative to that of other proinflammatory cytokines (TNF-a and IL-1) and
coincides with their downregulation. Moreover, in vivo studies showed that plasma TNF-
a levels are higher and remained elevated for a much longer period of time in IL-10–
deficient (IL-102/2) mice injected with LPS than in IL-101/1 mice.18 It therefore seems
that IL-10 acts in a feedback loop to inhibit continued proinflammatory cytokine
production. In vitro cell culture systems have yielded conflicting insights into the
modulatory actions of IL-10 on ECs and SMCs. Most of the studies failed to demonstrate
any direct anti-inflammatory effect on the expression by ECs of adhesion molecules,19
chemokines,20 colony-stimulating factors,21 IL-6 production,22 or IFN-g induction of
class II MHC surface antigen.23 Similarly, IL-10 had no effect on IL-8 and MCP-1 release
by human aortic SMCs in response to IL-1a or TNF-a.24 Lack of direct effects of IL-10 on
vascular cells in vitro might be attributable to the lack of IL-10 receptor or to an
impairment of the complex intracellular IL-10 signaling pathway. Moreover, the effects
of IL-10 on vascular cells may vary according to the origin of the cells and the signaling
pathways induced by the proinflammatory stimuli. Accordingly, in other studies it was
found that IL-10 downregulates the expression of ICAM-1 and VCAM-1 on IL-1–
activated HUVECs,25 decreases both IL-8 and IL-6 production by irradiated HUVECs,26
inhibits TNF-induced or fibroblast growth factor-2 (FGF-2)–induced human aortic SMC
proliferation, 27 and partially antagonizes IFN-g–induced expression
of the secretory nonpancreatic phospholipase A2 in human SMCs.28 Moreover,
pretreatment of human aortic ECs with recombinant IL-10 as well as transfection with
an adenovirus expressing viral IL-10 causes a significant decrease in minimally modified
LDL-induced monocyte binding. 29 Interestingly, these latter observations are
supported by in vivo findings in mice showing that electrotransfer of IL-10 cDNA results
in significant decrease in endothelial NF-kB activation and in expression of VCAM-1 and
ICAM-1 after 10 days on a high-fat diet.30 IL-10 exerts its biological effects on cells by
interacting with a specific cell-surface receptor. Functionally active IL-10 receptor is
composed of two distinct subunits. Both subunits belong to the class II cytokine receptor
family. The IL-10 receptor a chain (or IL-10R1) plays the dominant role in mediating
high-affinity ligand binding and signal transduction. 31 The IL-10 receptor b subunit (IL-
10R2, also known as the orphan receptor CRF2-4) serves as an accessory chain essential
for the active IL-10 receptor complex and to initiate IL-10 –induced signal transduction
events.32 Studies using macrophages from mice with disrupted genes for Jak1, Stat1, or
Stat3 have revealed an obligate role for Jak1and Stat3 in mediating the anti-
inflammatory actions of IL-10.33 In addition to the Janus kinase/signal transducers and
activators of transcription (JAK-STAT) pathway, the presence of a carboxyl-terminal
sequence containing at least one functionally critical serine on the intracellular domain
of the IL-10 receptor a chain is required for expression of the anti-inflammatory actions
of IL-10.33 IL-10 functions to block NF-kB activity through both the suppression of IkB
kinase activity, preventing IkBa degradation, and the suppression of NF-kB DNA-
binding activity34 (Figure 1). IL-10 also affects signaling through extracellular signal–
regulated kinase (ERK) 1 and ERK2 and other mitogen-activated protein kinase (MAPK)
pathways that are potentially important for chemokine and cytokine induction35 and
destabilizes the mRNA of proinflammatory genes with clustered AU-rich elements
motifs.36 So far, nothing is known concerning the expression of IL-10 receptor a and b
chains and the complex IL-10 signaling pathway in vascular cells. In vivo, IL-10 most
likely exerts its anti-inflammatory effects on the vascular system through inhibition of
leukocyte-EC interactions37–40 and inhibition of proinflammatory cytokine and
chemokine production by macrophages or lymphocytes.41–43 In an acute lung injury
model, IL-10 significantly decreased lung injury and ICAM-1 levels through decrease in
TNF-a levels.37 LPS-induced expression of ICAM-1 and VCAM-1 in the vasculature of
the small intestine and leukocyte adhesion in mesenteric venules are markedly increased
in IL-10–deficient mice compared with wild-type animals, underlining the inhibitory
role of endogenous IL-10 in the control of intestinal vascular inflammation. 38,40
Transfected viral IL-10 decreases leukocyte vein extravasation through a decrease in
endothelial expression of P- and E-selectin and ICAM-1.40 IL-10 similarly blunts
inflammation secondary to myocardial ischemia/reperfusion through an ICAM-1–
dependent mechanism44 and reduces liver injury and mortality in a mouse septic shock
model through decreased neutrophil margination and ICAM-1 and VCAM-1
expression.45 Endogenous IL-10 might also be produced during myocardial ischemia-
reperfusion by lymphocytes infiltrating the reperfused myocardium and could limit
myocardial macrophage activation.46 The protective effect of IL-10 against the
development of diet-induced atherosclerosis could also be attributed to inflammatory
cell deactivation. Expression of IL-10 in the atherosclerotic lesion42,47 is associated with
low iNOS expression by macrophages and low levels of cell death.42 In addition,
atherosclerotic lesions of IL-102/2 mice fed an atherogenic diet are characterized by
increased infiltration of inflammatory cells, particularly activated T cells, and by
increased production of proinflammatory cytokines, underscoring the anti-inflammatory
actions of IL-10 produced within the atherosclerotic plaque.43 In a model of balloon
angioplasty or stent implantation in hypercholesterolemic rabbits, treatment with
recombinant human IL-10 markedly reduced macrophage infiltration and intimal
hyperplasia.48 Additional mechanisms of vascular protection by endogenous IL-10
include decreased superoxide anion production in blood vessels in response to LPS,
which prevents the impairment of endothelium-dependent relaxation.49 It is noteworthy
that IL-10 most likely exerts its anti-inflammatory effect when produced locally in the
vascular wall. Chronic production of high levels of IL-10 in the systemic circulation may
instead lead to immunostimulatory effects.

IL-1 Receptor Antagonist

IL-1 is one of the most potent proinflammatory cytokines acting on both ECs and SMCs.1
Processed mature IL-1 signals via the type I IL-1 receptor but also binds to a
nonsignaling receptor (IL-1 receptor type II). The IL-1 receptor antagonist (IL-1ra) is an
endogenous secreted protein that binds to IL-1 type I and II receptors without signaling.
An intracellular form of IL-1ra is expressed by human ECs and SMCs,50,51 but its role
remains unclear. However, in vivo studies reveal that IL-1ra does have vascular
protective effects. Treatment with recombinant IL- 1ra inhibits fatty streak formation in
apoE2/2 mice.52 More importantly, IL-1ra knockout mice develop lethal chronic
inflammation of the arterial wall, associated with massive transmural infiltration of
neutrophils, macrophages, and CD41 lymphocytes in branch points and flexures of the
aorta and in its primary and secondary branches.53 Additional support for a vascular
role of IL-1ra is provided by the recent observation of an association between IL-1ra gene
polymorphism and coronary artery disease.

Th2 Anti-Inflammatory Cytokines IL-4 and IL-13

Th2 cytokines IL-4 and IL-13 suppress the production of inflammatory cytokines by
macrophages and monocytes and are considered anti-inflammatory cytokines. However,
IL-4 and IL-13 selectively induce VCAM-1 and P-selectin expression on ECs with no
effect on ICAM-1 or E-selectin.55–57 IL-13 markedly enhances IL-8 and MCP-1 release
by cytokinestimulated human SMCs24 but inhibits NOS II expression in LPS-activated
rat SMCs.58 In vivo studies indicate that IL-4 and IL-13 are capable of promoting
angiogenesis57 and that IL-4 plays a role in the progression of early inflammatory
atherosclerotic lesions driven by immunization against heat shock protein 65.59 This
may be consistent with the recently reported switch from Th1 to Th2 responses during
atherosclerosis progression in severely hypercholesterolemic apoE2/2 mice.60 However,
it remains unknown to what extent this switch might affect lesion progression. In
general, it seems that the macrophages deactivating cytokines IL-4 and IL-13 display
proinflammatory activities in the vascular system.

High-Density Lipoprotein
There is abundant evidence from epidemiological studies that
HDL plasma concentration is inversely correlated with the
occurrence of coronary artery disease. Besides the effects of
HDL on the promotion of cholesterol efflux and protection
against lipid peroxidation, it exerts potent anti-inflammatory
activities on ECs. HDL inhibits cytokine-induced expression
of E-selectin, ICAM-1, and VCAM-1 on ECs at the transcriptional
level.61–64 The effects of HDL seem to be related to its
phospholipid content.63 The anti-inflammatory effects of
HDL on ECs could involve the sphingosine kinase (SphK)
pathway through the generation of sphingosine 1 phosphate
(S1P).65 HDLs inhibit the TNF-induced SphK activity and
S1P generation and are expected to subsequently reduce the
activation of ERK and NF-kB signal cascades (Figure 1).
However, other studies reported that the anti-inflammatory
effects of HDL are not mediated by a direct inhibition of the
NF-kB pathway, because HDLs do not inhibit IkBa degradation
or the nuclear translocation of NF-kB.62 Furthermore,
HDLs inhibit E-selectin expression in response to proinflammatory
cytokines but have no effect on the expression of
NF-kB–dependent genes, such as granulocyte-macrophage
colony-stimulating factor (GM-CSF) and COX-2. Interestingly,
HDLs also stimulate COX-2 expression in SMCs,66
which suggests that the anti-inflammatory effect of HDL
might be restricted to a specific set of genes.
Whatever the molecular mechanisms of the antiinflammatory
effects of HDL, the pathophysiological relevance
of the ability of HDL to exert these activities is
substantiated by several studies showing that HDLs inhibit
endothelial cell adhesion molecule expression in vivo. Elevation
of the circulating levels of HDL inhibits E-selectin
expression by microvascular ECs in a porcine model of acute

Angiogenic and Growth Factors

VEGF is a potent factor in increasing permeability of endothelial
cells that leads to the passage of plasma components
and leukocytes from the blood vessel into the tissues and may
therefore contribute to the inflammatory response. Yet VEGF
might exert anti-inflammatory protective functions by stimulating
endothelial NO production,68 which may inhibit
leukocyte recruitment (see below). The receptor mediating
VEGF-induced NO production in ECs is likely to be VEGF
receptor-2.69 This effect is mediated by a signaling cascade
initiated by flk-1/KDR activation of c-Src, leading to phospholipase
Cg1 activation, inositol 1,4,5-trisphosphate formation,
release of intracellular Ca21, and NOS activation.69
Angiopoietin-1 (Ang-1), the ligand of the endotheliumspecific
tyrosine kinase receptor Tie-2, has been shown to be
an anti-inflammatory agent in vitro.70 Ang-1 pretreatment of
ECs abolished TNF-a–induced transmigration. This effect
likely results from enhanced platelet EC adhesion molecule-1
(PECAM-1) localization to the EC junctions.70 Interestingly,
hypoxia and inflammatory cytokines upregulate Tie2 receptor
in HUVECs and human microvascular EC-1.71 In this context,
enhancement of PECAM-1 or PECAM-1 engagement
mediated by Ang-1 might promote a noninflammatory phenotype
of ECs.
Growth factors not only modulate vascular cell survival
and growth but also may act as modulators of inflammatory
responses. FGF-2 can inhibit endothelial expression of tissue
factor and other inflammatory genes, including tissue plasminogen
activator, plasminogen activator inhibitor-2, and
IL-8, in response to phorbol myristate acetate.72 Exposure of
ECs to FGF-2 or FGF-1/heparin also inhibits cytokinemediated
expression of ICAM-1, VCAM-1, E-selectin, and
COX.73,74 However, this anti-inflammatory effect of FGF-1
and FGF-2 seems to be EC-specific, because FGF-2 induces
tissue factor expression in fibroblasts, monocytes, and
SMCs.75 In SMCs, growth factors may have ambiguous
effects. Platelet-derived growth factor induces MCP-176 and
ICAM-1,77 whereas it inhibits cytokine-stimulated expression
of NOS II and NO release.

2. Intracellular Anti-Inflammatory Mediators

Regulators of NF-kB Signaling Pathway

The expression of inducible genes leading to the synthesis of cytokines, chemokines,
adhesion molecules, and autacoids relies on transcription factors. Among the primary
transcription factors, NF-kB plays a central role in the regulation of inflammatory
mediators.79 Events leading to the activation of NF-kB rely on the phosphorylation of
IkBa followed by its ubiquitination and proteolytic degradation into the proteasome.
Phosphorylation of IkBa depends on a IkB kinase (IKK) complex containing two kinases,
IKKa and IKKb, and the regulatory protein NEMO (NF-kB essential modifier, also
named IKKg).80 Moreover, NF-kB itself induces the synthesis of IkBa to regenerate an
inactive form of NF-kB and ensures the transiency of NF-kB activation. The intracellular
redox status of the cell is extremely important in the regulation of NF-kB/IkB by
preventing the activation of IkB kinase.81 Antioxidants, aspirin, N-acetyl-L-cysteine
(NAC), and flavonoids may inhibit the activation of NF-kB. We have seen that TGF-b and
IL-10 negatively regulate the NF-kB pathway. Glucocorticoids enhance the formation of
IkB, and several constitutive or inducible cytoprotective genes have been shown to
inhibit NF-kB activity in ECs (see below). NO also inhibits NF-kB activity in ECs through
the induction and stabilization of IkBa (see below). The activation of NF-kB can also be
attenuated by inhibiting the proteolytic degradation of IkB in the proteasome. A
naturally occurring antibacterial peptide, PR39, which reversibly binds the 26S
proteasome and blocks the degradation of IkBa by the ubiquitinpreoteasome pathway,
suppresses VCAM-1 and ICAM-1 gene expression in TNF-a–activated human ECs and
reduces the size of myocardial infarction in an in vivo mouse model of coronary
ligature.82 Another possibility to interrupt the NF-kB pathway is to block the interaction
between NEMO and IKKb.83 A cell-permeable NEMO-binding domain (NBD) peptide
able to disrupt the IKKb-NEMO interaction efficiently reduces E-selectin expression in
TNF-a–treated HUVECs.83 These anti-inflammatory agents (PR39 and NBD peptide)
that directly interact intracellularly with the NF-kB pathway (Figure 1) might prove to be
useful tools to block proinflammatory activation in the vascular wall.
Protective Genes
Endogenous protective genes can be expressed by vascular cells to limit the
inflammatory process and injury. Indirect arguments suggest that both ECs and VSMCs
may develop an autoprotective phenotype during inflammation.
Cytoprotective Genes
In addition to protecting ECs from apoptosis, several antiapoptotic genes have been
shown to possess potent anti-inflammatory properties (Figure 2). As a consequence, they
have been named cytoprotective genes. These include members of the Bcl-2 family (Bcl-
2, Bcl-xL, and A1), A20, and heme oxygenase-1 (HO-1). A1 and A20 are induced in
response to inflammatory stimuli to protect ECs from unfettered activation and from
undergoing apoptosis even when NF-kB is blocked.84 Overexpression of Bcl-2, Bcl-xL,
A1, or A20 inhibits VCAM-1, E-selectin, and IL-8 expression in ECs by inhibiting NF-kB
activation.85,86 In vivo experiments underscore the importance of these protective
genes in organ xenografts. ECs in hamster hearts that accommodate themselves
in rats express certain genes, such as A20 and bcl-2,whereas hearts that are rejected do
not express these genes.87
In addition, vessels of rejected hearts show florid transplant
arteriosclerosis, whereas those of accommodated hearts do
not. Moreover, studies in mice deficient for A20 confirm the
critical role of this protective gene for limiting TNF-a–
dependent NF-kB activation and inflammation.88 A20 knockout
mice develop severe inflammation and cachexia, are
hypersensitive to both LPS and TNF, and die prematurely.
Taken together, these data suggest that A1 and A20 offer the
mean of achieving an anti-inflammatory effect in the vascular
wall.
HOs also belong to the family of cytoprotective genes.
HOs catalyze the rate-limiting step in the degradation of
heme to yield equimolar amounts of biliverdin, carbon
monoxide, and iron. Besides their antiapoptotic effect, there
is a growing body of evidence that ascribes an antiinflammatory
role for the products of the inducible form of
HO (HO-1).89 HO-1 can be upregulated in human ECs by
TNF and IL-1.90 In particular, exogenous administration of
HO-1 by gene transfer protects the rat lung against hyperoxiainduced
neutrophil infiltration and tissue injury.91 Moreover,
HO-1 deficiency in humans is associated with the presence of
severe and persistent endothelial damage.92
Fas and Fas Ligand
Fas ligand (FasL) is a death factor that induces apoptosis in
Fas-bearing cells. FasL is constitutively expressed on ECs but
not in SMCs.93 Local administration of TNF-a to arteries
downregulates endothelial FasL expression and induces
mononuclear cell infiltration, whereas FasL overexpression
markedly attenuates TNF-a–induced cell infiltration. Moreover,
adherent mononuclear cells undergo apoptosis rather
than diapedesis under these conditions as a result of Fas-FasL
ligation. These data suggest that endothelial FasL plays an
active role in inhibiting leukocyte extravasation and vascular
inflammation. Recent experiments in FasL-deficient mice
additionally support this contention.94 In a model of flow
restriction in the common carotid artery, vascular T lymphocyte
and macrophage infiltration after flow restriction is
notably enhanced in FasL knockout mice compared with
wild-type mice. Moreover, the flow-restricted common carotid
arteries develop greater neointima formation in FasL
knockout mice than in wild-type mice.
Serpine Proteinase Inhibitor 9
SMCs and ECs express the 33-kDa precursors of both
IL-1a and IL-1b as cell-associated proteins, but SMCs
neither contain mature IL-1b nor are able to process recombinant
IL-1b precursor into its mature 17-kDa form. Despite
this failure, SMCs express IL-1– converting enzyme but
possess in their cell membrane compartment an inhibitory
factor of IL-1b processing, recently identified as the serpine
proteinase inhibitor 9 (PI-9).95 PI-9 is homogenously expressed
in the normal arterial wall, and its expression inversely
correlates with immunoreactive IL-1b in the atherosclerotic
plaque, suggesting a potential role for PI-9 in this
inflammatory disease.
Nitric Oxide
Besides its action on vasomotor tone regulation, endothelium-
derived NO has been recognized to be an antiinflammatory
molecule. Endogenous NO synthesis inhibits
leukocyte rolling and adhesion as well as cytokine-induced
expression of ICAM-1 and VCAM-1.96,97 NO inhibits M-CSF
synthesis in ECs.98 Furthermore, inhibition of basal NO
production by NG-nitro-L-arginine in human ECs upregulates
and exogenous addition of NO decreases MCP-1 expression.
99 NO donors inhibit the expression of MCP-1 in SMCs
exposed to LPS or oxidized LDL100 and diminish VCAM-1
expression induced by IFN-g.101 The anti-inflammatory effects
of NO are attributable, at least in part, to inhibition of
NF-kB activation through increased expression and nuclear
translocation of IkBa97,102 (Figure 3). The crucial role of NO
as an endogenous anti-inflammatory mediator was later
substantiated by in vivo experiments of chronic inhibition of
NO synthesis. Administration of the NO synthesis inhibitor
Nv-nitro-L-arginine methyl ester (L-NAME) induces vascular
monocyte infiltration, MCP-1, IL-6, M-CSF, ICAM-1, and
VCAM-1 expression as well as NF-kB activation.103–106 In
vivo transfection of cis element decoy oligodeoxynucleotides
against NF-kB prevents the L-NAME–induced early inflammation,
suggesting that the NF-kB pathway is essential in this
process.104 Along with inflammatory changes, vascular superoxide
anion production is also increased after chronic NO
blockade, and the antioxidant NAC prevents all of these
Figure 3. Scheme explaining anti-inflammatory mechanisms of
shear stress. Low or oscillatory shear stress, as well as proinflammatory
signals, increases intracellular oxidative stress that
activates the NF-kB pathway. Physiological laminar shear stress
stimulates NO production, which induces IkBa expression,
resulting in inhibition of NF-kB activation and acceleration of
p50/p65 nuclear deactivation. Physiological shear stress can
also stimulate superoxide dismutase expression that reduces
oxidative stress and can block the JNK pathway that is activated
in response to proinflammatory stimuli.
Therefore, inhibition of NO synthesis increases
vascular oxidative stress leading to inflammatory
responses. Interestingly, in L-NAME–induced vascular inflammation,
treatment with an angiotensin II type 1 receptor
antagonist also prevents NF-kB activity and the consequent
inflammatory changes.103,107 Taken together, these data suggest
that endogenous endothelial NO decreases proinflammatory
oxidative stress-sensitive signals by suppressing localized
activity of angiotensin II in blood vessels.
Peroxisome Proliferator–Activated Receptors
Peroxisome proliferator–activated receptors (PPARs) are transcription
factors belonging to the nuclear receptor superfamily,
of which three different PPAR subtypes have been identified,
PPARa, PPARb/d, and PPARg. PPARs regulate gene expression
by binding with the retinoid receptor RXR as a heterodimeric
partner to specific DNA sequence elements termed
PPAR-responsive elements. Fatty acid derivatives and eicosanoids
have been identified as natural ligands for PPARs.108
Furthermore, fibrates are synthetic ligands for PPARa that
mediate the lipid-lowering activity of these drugs, and the
antidiabetic thiazoldinediones are synthetic ligands for PPARg.
PPARa and PPARg have been found to be expressed in both
ECs109,110 and SMCs111–113 in vitro and in vivo in the human
atherosclerotic plaque.114–116 Anti-inflammatory actions of
PPARs were first reported in monocytes and macrophages for
PPARg.117,118 Thereafter, PPARa activation, but not PPARg,
was shown to repress cytokine-induced activation of COX-2 and
IL-6 in human SMCs111 and VCAM-1 in human ECs, resulting
in reduced functional adhesion of monocytes.119 These in vitro
findings are in good agreement with an earlier study showing
that PPARa knockout mice have increased acute inflammatory
responses.120 Moreover, recent findings clearly indicate that
PPARa has anti-inflammatory properties in the vascular wall;
aortic explants isolated from PPARa knockout mice display an
exacerbated response to inflammatory stimuli, resulting in increased
IL-6 production compared with wild-type mice.113
Furthermore, fibrate treatment represses IL-6 mRNA levels in
LPS-stimulated aortas of wild-type mice but not of PPARa
knockout mice.
In addition to regulating gene transcription via PPAR responsive
elements, PPARs have recently been shown to modulate
gene transcription by interfering with other transcription factor
pathways in a DNA binding–independent manner.111,113,117,119
PPARs have been shown to downregulate inflammatory response
genes by negatively interfering with the STAT, AP-1,
and NF-kB transcriptional pathways.109,111,113,117,119 For example,
direct protein-protein interactions between PPARa and
AP-1 and NF-kB proteins have been invoked as mechanisms of
transrepression.113 In addition, by regulating antioxidant enzyme
activities, such as catalase,121 PPARa activators reduce the
oxidative stress and, as a result, may inhibit NF-kB activation.
Finally, PPARa activators may antagonize NF-kB activation
through the expression of the inhibitory protein IkBa, as shown
in IL-1b–stimulated human aortic SMCs in the presence of
fibrates122 (Figure 1).
Several years ago, the n-3 fatty acid docosahexaenoic acid
docosahexaenoic acid was reported to limit cytokine-induced
expression of VCAM-1 and other proinflammatory mediators in
human ECs.123 We now know that this is likely attributable to
the anti-inflammatory activities of PPARa, which is a target for
various long-chain fatty acids, including n-3 fatty acids.124
The anti-inflammatory activities of PPARa take on particular
significance in view of the findings that fibrate treatment
decreases plasma concentrations of inflammatory cytokines
in patients with angiographically established atherosclerosis.
111 Furthermore, the recent Veteran’s Administration HDL
Intervention Trial showed a beneficial effect of the fibrate
gemfibrozil on atherosclerotic events that could not be
accounted for by reductions in LDL concentrations.125
Anti-Inflammatory Effects of Shear Stress
Apart from humoral stimuli, ECs are constantly exposed to a
spectrum of hemodynamic forces generated by pulsatile
blood flow, hydrostatic pressure, cyclic strains, and wall
shear stresses.126 Biomechanical forces induce endothelial
structural changes and modulate gene expression.127 Most in
vitro experiments using ECs suggest that physiological levels
of shear stress are essentially anti-inflammatory and antiadhesive.
Prolonged exposure of ECs to laminar flow, as occurs
in vivo, results in downregulation of ICAM-1 and VCAM-
1,128 whereas prolonged low or oscillatory shear-stress conditions
enhance monocyte adhesion and VCAM-1, ICAM-1,
and E-selectin expression.129,130 Anti-inflammatory effects of
laminar shear stress may also prevail in conditions of activated
ECs. Chronic laminar shear stress suppresses VCAM-1
expression by IL-1b–stimulated ECs, whereas oscillatory
shear stress has no effect.130 Similarly, monocyte adhesion to
ECs in the presence of oxidized lipids is markedly reduced by
physiological pulsatile unidirectional flow, whereas oscillatory
flow promotes monocyte adhesion.131 These in vitro
studies are in agreement with in vivo experiments showing
that low or oscillatory shear stress provides a proinflammatory
stimulus to ECs.30,132,133 Constitutive NF-kB activation
and VCAM-1 expression are seen in ECs located in aortic
regions of high probability for atherosclerotic lesion development,
which are known to be regions of altered hemodynamics
forces.30,133 Moreover, chronically decreased blood
flow in rabbits stimulates VCAM-1 expression and enhances
monocyte adhesion.132
Molecular mechanisms of anti-inflammatory actions of
shear stress involve protection against oxidative stress and
inhibition of NF-kB and jun-N-terminal kinase (JNK)–AP-1
pathways (Figure 3). Laminar shear stress induces Cu/Zn
superoxide dismutase,134,135 suggesting that absence or decrease
of shear stress results in increased production of
superoxide radicals. In contrast, low or oscillatory flow
patterns induce a sustained activation of pro-oxidant processes,
resulting in redox-sensitive gene expression.129,136
The antioxidant pyrrolidine dithiocarbamate, but not NAC,
strongly inhibits low shear–induced NF-kB activation, expression
of VCAM-1, and monocyte adhesion.129 Because
NAC seems to have no effect on superoxide radical,137 it is
tempting to hypothesize that low shear stress allows O2
2-
dependent activation of NF-kB and subsequently VCAM-1.
However, both pyrrolidine dithiocarbamate and NAC inhibit
NF-kB activation and VCAM-1 expression induced by oscillatory
shear stress or cytokines, suggesting that other pro-
Tedgui and Mallat Anti-Inflammatory Mechanisms in the Vascular Wall 883
Downloaded from http://circres.ahajournals.org/ by guest on November 1, 2011
oxidant pathways may be involved (H2O2/OH z-sensitive
mechanisms).129,130 Taken together, these data also indicate
that ECs discriminate between various types of flow and
between flow and cytokine stimulation.
Other mechanisms may account for the anti-inflammatory
actions of shear stress. The MAPK JNK is activated by exposure
of cells to cytokines or environmental stress and contributes to
inflammatory responses.138 Laminar shear stress specifically
inhibits cytokine-induced JNK activity but has no effect on the
other MAPKs.139 Shear stress also abrogates the complementinduced
IL-8 and MCP-1 expression in ECs through upregulation
of the complement-inhibitory protein clustering.140 In addition,
shear stress upregulates the expression of the inhibitory
adapter protein tumor necrosis receptor-associated factor
(TRAF)-3, and transfection of a dominant-negative TRAF3
mutant reverses the inhibitory effect of shear stress on CD40-
induced MCP-1 expression.141 Finally, shear stress is known to
be the physiological activator/inducer of NOS III, and NO has
anti-inflammatory actions by scavenging O2
2 and through inhibition
of NF-kB pathway (see above).
Conclusions
The involvement of vascular inflammation in several pathological
conditions, including atherosclerosis, ischemia/reperfusion,
hypertension, restenosis, angiogenesis, septic shock,
and cerebral malaria, has only recently been fully revealed. In
the context of atherosclerosis, the inflammatory response
seems to determine the development of the disease from even
the site and extent of endothelial activation to lesion growth
and lesion complications. This is consistent with clinical
studies in humans showing a major prognostic and independent
value for markers of inflammation in predicting the
occurrence of ischemic vascular events.142 Great progress has
been made during the last decade in the understanding of the
molecular mechanisms that mediate the inflammatory responses
in vascular cells. The identification of protective
anti-inflammatory mechanisms is much more recent. Interestingly,
several effects of some well-known antiatherogenic
and vasculoprotective agents, including HDL, statins, fibrates,
NO, shear stress, antioxidants, and n3-fatty acids,
might in fact be attributable, at least in part, to their
anti-inflammatory properties. More importantly, recent data
suggest that vascular cells may develop genetically determined
intrinsic anti-inflammatory mechanisms.143 Delineating
these anti-inflammatory pathways will be one of the
challenging future objectives in vascular biology, which
could yield promising novel therapeutic possibilities. Yet
such anti-inflammatory strategies should be specifically targeted
at the diseased tissue, particularly when long-term
therapy is necessary, to limit potential side effects.

4. agen2 yang melemahkan sel :

 Burns
 Chemical irritants
 Frostbite
 Toxins
 Infection by pathogens
 Necrosis
 Physical injury, blunt or penetrating
 Immune reactions due to hypersensitivity
 Ionizing radiation
 Foreign bodies, including splinters and dir
5. mekanisme pertahanan sel melalui keradangan (inflamasi) : definisi,
jenis, proses, faktor2 pemicu, peran/kegunaan, mekanisme trjadinya
tanda2 radang , upaya pencegahan, penghambatan dan penanganan

Inflammation

is the complex biological response of vascular tissues to harmful stimuli, such as

pathogens, damaged cells, or irritants. It is a protective attempt by the organism to
remove the injurious stimuli as well as initiate the healing process for the tissue.
Inflammation is not a synonym for infection. Even in cases where inflammation is
caused by infection it is incorrect to use the terms as synonyms: infection is caused by an
exogenous pathogen, while inflammation is the response of the organism to the
pathogen.
In the absence of inflammation, wounds and infections would never heal and progressive
destruction of the tissue would compromise the survival of the organism.
Inflammation can be classified as either acute or chronic.
Acute inflammation is the initial response of the body to harmful stimuli and is achieved
by the increased movement of plasma and leukocytes from the blood into the injured
tissues. A cascade of biochemical events propagates and matures the inflammatory
response, involving the local vascular system, the immune system, and various cells
within the injured tissue.
Prolonged inflammation, known as chronic inflammation, leads to a progressive shift in
the type of cells which are present at the site of inflammation and is characterised by
simultaneous destruction and healing of the tissue from the inflammatory process.
Types of Inflamation

Comparison between acute and chronic inflammation:

Acute Chronic
Persistent acute inflammation due to
Pathogens, injured
Causative agent non-degradable pathogens, persistent
tissues
foreign bodies, or autoimmune reactions
Mononuclear cells (monocytes,
Major cells involved Neutrophils macrophages, lymphocytes, plasma
cells), fibroblasts
IFN-γ and other cytokines, growth
Vasoactive amines,
Primary mediators factors, reactive oxygen species,
eicosanoids
hydrolytic enzymes
Onset Immediate Delayed
Duration Few days Up to many months, or years
Healing, abscess
Outcomes formation, chronic Tissue destruction, fibrosis
inflammation

Sign and Simptoms of Inflamation

The classic signs and symptoms of acute inflammation:

English Latin
Redness Rubor
Heat Calor
Swelling Edema
Pain Dolor
Loss of function Functio laesa
Acute inflammation is a short-term process which is characterised by the classic signs of
inflammation - swelling, redness, pain, heat, and loss of function - due to the infiltration
of the tissues by plasma and leukocytes. It occurs as long as the injurious stimulus is
present and ceases once the stimulus has been removed, broken down, or walled off by
scarring (fibrosis). The first four characteristics have been known since ancient times
and are attributed to Celsus. Loss of function was added to the definition of
inflammation by Rudolf Virchow in the 19th century[1].

Penyebab Inflamasi

 Burns
 Chemical irritants
 Frostbite
 Toxins
 Infection by pathogens
 Necrosis
 Physical injury, blunt or penetrating
 Immune reactions due to hypersensitivity
 Ionizing radiation
 Foreign bodies, including splinters and dirt

Mekanisme dan Proses Inflamasi

The process of acute inflammation is initiated by the blood vessels local to the injured
tissue, which alter to allow the exudation of plasma proteins and leukocytes into the
surrounding tissue.
The increased flow of fluid into the tissue causes the characteristic swelling associated
with inflammation, and the increased blood flow to the area causes the reddened colour
and increased heat.
The blood vessels also alter to permit the extravasation of leukocytes through the
endothelium and basement membrane constituting the blood vessel. Once in the tissue,
the cells migrate along a chemotactic gradient to reach the site of injury, where they can
attempt to remove the stimulus and repair the tissue.
Meanwhile, several biochemical cascade systems, consisting of chemicals known as
plasma-derived inflammatory mediators, act in parallel to propagate and mature the
inflammatory response.
These include the complement system, coagulation system and fibrinolysis system.
Finally, down-regulation of the inflammatory response concludes acute inflammation.
Removal of the injurious stimuli halts the response of the inflammatory mechanisms,
which require constant stimulation to propagate the process. Additionally, many
inflammatory mediators have short half lives and are quickly degraded in the tissue,
helping to quickly cease the inflammatory response once the stimulus has been
removed[1].

Plasma cascade systems

 The complement system, when activated, results in the increased removal of
pathogens via opsonisation and phagocytosis.
 The kinin system generates proteins capable of sustaining vasodilation and other
physical inflammatory effects.
 The coagulation system or clotting cascade which forms a protective protein
mesh over sites of injury.
  The fibrinolysis system, which acts in opposition to the coagulation system, to
counterbalance clotting and generate several other inflammatory mediators.

Plasma derived mediators

* non-exhaustive list
Produced
Name Description
by
A vasoactive protein which is able to induce vasodilation,
Bradykinin Kinin system increase vascular permeability, cause smooth muscle
contraction, and induce pain.
Cleaves to produce C3a and C3b. C3a stimulates
histamine release by mast cells, thereby producing
Complement
C3 vasodilation. C3b is able to bind to bacterial cell walls and
system
act as an opsonin, which marks the invader as a target for
phagocytosis.
Stimulates histamine release by mast cells, thereby
Complement producing vasodilation. It is also able to act as a
C5a
system chemoattractant to direct cells via chemotaxis to the site
of inflammation.
A protein which circulates inactively, until activated by
collagen, platelets, or exposed basement membranes via
Factor XII
conformational change. When activated, it in turn is able
(Hageman Liver
to activate three plasma systems involved in
Factor)
inflammation: the kinin system, fibrinolysis system, and
coagulation system.
A complex of the complement proteins C5b, C6, C7, C8,
and multiple units of C9. The combination and activation
Membrane
Complement of this range of complement proteins forms the
attack
system membrane attack complex, which is able to insert into
complex
bacterial cell walls and causes cell lysis with ensuing
death.
Fibrinolysis Able to break down fibrin clots, cleave complement
Plasmin
system protein C3, and activate Factor XII.
 Cleaves the soluble plasma protein fibrinogen to produce
insoluble fibrin, which aggregates to form a blood clot.
Coagulation
Thrombin Thrombin can also bind to cells via the PAR1 receptor to
system
trigger several other inflammatory responses, such as
production of chemokines and nitric oxide.

Cellular component
The cellular component involves leukocytes, which normally reside in blood and must
move into the inflamed tissue via extravasation to aid in inflammation. Some act as
phagocytes, ingesting bacteria, viruses, and cellular debris. Others release enzymatic
granules which damage pathogenic invaders. Leukocytes also release inflammatory
mediators which develop and maintain the inflammatory response. Generally speaking,
acute inflammation is mediated by granulocytes, while chronic inflammation is mediated
by mononuclear cells such as monocytes and lymphocytes.
Leukocyte extravasation
Neutrophils migrate from blood vessels to the inflamed tissue via chemotaxis, where
they remove pathogens through phagocytosis and degranulation.
Various leukocytes are critically involved in the initiation and maintenance of
inflammation. These cells must be able to get to the site of injury from their usual
location in the blood, therefore mechanisms exist to recruit and direct leukocytes to the
appropriate place. The process of leukocyte movement from the blood to the tissues
through the blood vessels is known as extravasation, and can be divided up into a
number of broad steps:
1. Leukocyte localisation and recruitment to the endothelium local to
the site of inflammation – involving margination and adhesion to the
endothelial cells: Recruitment of leukocytes is receptor-mediated. The products of
inflammation, such as histamine, promote the immediate expression of P-selectin on
endothelial cell surfaces. This receptor binds weakly to carbohydrate ligands on
leukocyte surfaces and causes them to "roll" along the endothelial surface as bonds
are made and broken. Cytokines from injured cells induce the expression of E-
selectin on endothelial cells, which functions similarly to P-selectin. Cytokines also
induce the expression of integrin ligands on endothelial cells, which further slow
leukocytes down. These weakly bound leukocytes are free to detach if not activated
by chemokines produced in injured tissue. Activation increases the affinity of bound
integrin receptors for ligands on the endothelial cell surface, firmly binding the
leukocytes to the endothelium.
2. Migration across the endothelium, known as transmigration, via the
process of diapedesis: Chemokine gradients stimulate the adhered leukocytes to
move between endothelial cells and pass the basement membrane into the tissues.
3. Movement of leukocytes within the tissue via chemotaxis: Leukocytes
reaching the tissue interstitium bind to extracellular matrix proteins via expressed
integrins and CD44 to prevent their loss from the site. Chemoattractants cause the
leukocytes to move along a chemotactic gradient towards the source of inflammation.
Cell derived mediators

Name Type Source Description

These cells contain a large variety of
enzymes which perform a number of
functions. Granules can be classified as
either specific or azurophilic depending
Lysosome
Enzymes Granulocytes upon the contents, and are able to break
granules
down a number of substances, some of
which may be plasma-derived proteins
which allow these enzymes to act as
inflammatory mediators.
Stored in preformed granules, histamine
Mast cells,
Vasoactive is released in response to a number of
Histamine basophils,
amine stimuli. It causes arteriole dilation and
platelets
increased venous permeability.
Antiviral, immunoregulatory, and anti-
tumour properties. This interferon was
IFN-γ Cytokine T-cells, NK cells originally called macrophage-activating
factor, and is especially important in the
maintenance of chronic inflammation.
Activation and chemoattraction of
Primarily
IL-8 Chemokine neutrophils, with a weak effect on
macrophages
monocytes and eosinophils.
Able to mediate leukocyte adhesion and
activation, allowing them to bind to the
endothelium and migrate across it. In
neutrophils, it is also a potent
Leukotriene B4 Eicosanoid Leukocytes
chemoattractant & is able to induce the
formation of reactive oxygen species and
the release of lysosome enzymes by these
cells.
Nitric oxide Soluble gas Macrophages, Potent vasodilator, relaxes smooth
endothelial muscle, reduces platelet aggregation, aids

Prostaglandins Eicosanoid
TNF-α and IL-1 Cytokines
macrophages leukocyte adherence, activation of
in leukocyte recruitment, direct
cells, some
antimicrobial activity in high
neurons
concentrations.
A group of lipids which can cause
Mast cells
vasodilation, fever, and pain.
Both affect a wide variety of cells to
induce many similar inflammatory
reactions: fever, production of cytokines,
Primarily endothelial gene regulation, chemotaxis,
fibroblasts. Responsible for the systemic
effects of inflammation, such as loss of
appetite and increased heart rate.

Peran Inflamasi
Keuntungan Inflamasi

It is a protective attempt by the organism to remove the injurious stimuli as well as

initiate the healing process for the tissue. In the absence of inflammation, wounds and
infections would never heal and progressive destruction of the tissue would compromise
the survival of the organism.

Kerugian Inflamasi

However, inflammation which runs unchecked can also lead to a host of diseases, such as
hay fever, atherosclerosis, and rheumatoid arthritis. It is for this reason that
inflammation is normally tightly regulated by the body.

Inflammatory disorders
Abnormalities associated with inflammation comprise a large, unrelated group of
disorders which underly a variety of human diseases. The immune system is often
involved with inflammatory disorders, demonstrated in both allergic reactions and some
myopathies, with many immune system disorders resulting in abnormal inflammation.
Non-immune diseases with aetiological origins in inflammatory processes are thought to
include cancer, atherosclerosis, and ischaemic heart disease.[1]
A large variety of proteins are involved in inflammation, and any one of them is open to a
genetic mutation which impairs or otherwise dysregulates the normal function and
expression of that protein.
Examples of disorders associated with inflammation include:
 Asthma
 Autoimmune diseases
 Chronic inflammation
 Chronic prostatitis
 Glomerulonephritis
 Hypersensitivities
 Inflammatory bowel diseases
 Pelvic inflammatory disease
 Reperfusion injury
 Rheumatoid arthritis
 Transplant rejection
  Vasculitis

Allergies
An allergic reaction, formally known as type 1 hypersensitivity, is the result of an
inappropriate immune response triggering inflammation. A common example is hay
fever, which is caused by a hypersensitive response by skin mast cells to allergens. Pre-
sensitised mast cells respond by degranulating, releasing vasoactive chemicals such as
histamine. These chemicals propagate an excessive inflammatory response characterised
by blood vessel dilation, production of pro-inflammatory molecules, cytokine release,
and recruitment of leukocytes.[1] Severe inflammatory response may mature into a
systemic response known as anaphylaxis.
Other hypersensitivity reactions (type 2 and type 3) are mediated by antibody reactions
and induce inflammation by attracting leukocytes which damage surrounding tissue. [1]
Myopathies
Inflammatory myopathies are caused by the immune system inappropriately attacking
components of muscle, leading to signs of muscle inflammation. They may occur in
conjunction with other immune disorders, such as systemic sclerosis, and include
dermatomyositis, polymyositis, and inclusion body myositis.[1]

Leukocyte defects
Due to the central role of leukocytes in the development and propagation of
inflammation, defects in leukocyte function often result in a decreased capacity for
inflammatory defence with subsequent vulnerability to infection[1]. Dysfunctional
leukocytes may be unable to correctly bind to blood vessels due to surface receptor
mutations, digest bacteria (Chediak-Higashi syndrome), or produce microbicides
(chronic granulomatous disease). Additionally, diseases affecting the bone marrow may
result in abnormal or few leukocytes.

Upaya Menghentikan Proses Inflamasi

Pharmacological
Certain drugs or chemical compounds are known to affect inflammation. Vitamin A
deficiency causes an increase in inflammatory responses [2], and anti-inflammatory drugs
(NSAID & SAID) work specifically by inhibiting normal inflammatory components.
Termination
The inflammatory response must be actively terminated when no longer needed to
prevent unnecessary "bystander" damage to tissues.[1] Failure to do so results in chronic
inflammation, cellular destruction, and attempts to heal the inflamed tissue. One
intrinsic mechanism employed to terminate inflammation is the short half-life of
inflammatory mediators in vivo. They have a limited time frame to affect their target
before breaking down into non-functional components, therefore constant inflammatory
stimulation is needed to propagate their effects.
Active mechanisms which serve to terminate inflammation include [1]:
 TGF-β from macrophages
 Anti-inflammatory lipoxins
 Inhibition of pro-inflammatory molecules, such as leukotrienes
6. proses terjadinya keadaan pada kasus  LIHAT DI BRAIN STORMING
DAN MAPPING KASUS
7. terapi herbal untuk keradangan (penanganan pada kasus)  LIHAT
JURNAL PIPER BETLE LINN di brain storming