APOPTOSIS, INTRACELULLAR

ACUMULATION, PHATOLOGIC
CALCIFICATION, CELULAR
AGING
 APOPTOSIS
Programmed cell death

Orderly cellular self destruction

Process: as crucial for survival of multi-cellular

organisms as cell division

MULTIPLE FORMS???
 Forms of cell death

"Classic"
Necrosis Apoptosis Mitotic catastrophe

Passive Active Passive

Pathological Physiological or Pathological

pathological

Swelling, lysis Condensation, Swelling, lysis

cross-linking

Dissipates Phagocytosed Dissipates

Inflammation No inflammation Inflammation

Externally induced Internally or Internally induced

externally induced
 APOPTOSIS

Evolutionarily conserved

•Occurs in all multicellular animals studies (plants too!)

•Stages and genes conserved from nematodes (worms)

and flies to mice and humans
STAGES OF CLASSIC APOPTOSIS
Healthy cell

DEATH SIGNAL (extrinsic or intrinsic)

Commitment to die (reversible)

EXECUTION (irreversible)

Dead cell (condensed, crosslinked)

ENGULFMENT (macrophages, neighboring cells)

DEGRADATION
 STAGES OF CLASSIC APOPTOSIS

Genetically controlled: Caenorhabditis elegans

soil nematode (worm)

ces2 ces1 ced9 ced3,4

Healthy cell Committed cell Dead cell

BCL2 Caspases
(proteases)

C. elegans genes == mammalian genes

Cells are balanced between life and death
DAMAGE Physiological death signals

DEATH SIGNAL

PROAPOPTOTIC ANTIAPOPTOTIC
PROTEINS PROTEINS
(dozens!) (dozens!)

DEATH
 APOPTOSIS: important in embryogenesis

Morphogenesis (eliminates excess cells):

Selection (eliminates non-functional cells):

 APOPTOSIS: important in embryogenesis

Immunity (eliminates dangerous cells):

Self antigen
recognizing cell

Organ size (eliminates excess cells):

 APOPTOSIS: important in adults
Tissue remodeling (eliminates cells no longer needed):

Apoptosis
Virgin mammary gland Late pregnancy, lactation Involution
(non-pregnant, non-lactating)

- Testosterone
Apoptosis

Prostate gland
 APOPTOSIS: important in adults
Tissue remodeling (eliminates cells no longer needed):

Apoptosis

Resting lymphocytes + antigen (e.g. infection) - antigen (e.g. recovery)

Steroid immunosuppressants: kill

lymphocytes by apoptosis

Lymphocytes poised to die by apoptosis

 APOPTOSIS: important in adults
Maintains organ size and function:

Apoptosis
X
+ cell division

Cells lost by apoptosis are replaced by cell division

(remember limited replicative potential of normal cells

restricts how many times this can occur before
tissue renewal declines)
 APOPTOSIS: control
Receptor pathway (physiological):
FAS ligand TNF

Death receptors:
(FAS, TNF-R, etc)
Death
domains

Adaptor proteins

Pro-caspase 8 (inactive) Caspase 8 (active)

Pro-execution caspase (inactive)

Execution caspase (active)

MITOCHONDRIA Death
 APOPTOSIS: control
Intrinsic pathway (damage):

Mitochondria

BAX Cytochrome c release BCL-2

BAK BCL-XL
BOK BCL-W
BCL-Xs Pro-caspase 9 cleavage MCL1
BAD BFL1
BID DIVA
B IK NR-13
BIM Pro-execution caspase (3) cleavage Several
NIP3 viral
BNIP3 proteins

Caspase (3) cleavage of cellular proteins,

nuclease activation,
etc.

Death
 APOPTOSIS: control

Physiological Intrinsic
receptor pathway damage pathway

MITOCHONDRIAL SIGNALS

Caspase cleavage cascade

Orderly cleavage of proteins and DNA

CROSSLINKING OF CELL CORPSES; ENGULFMENT

(no inflammation)
APOPTOSIS: Role in Disease

TOO MUCH: Tissue atrophy

Neurodegeneration
Thin skin
etc

TOO LITTLE: Hyperplasia

Cancer
Athersclerosis
etc
 APOPTOSIS: Role in Disease
Neurodegeneration

Neurons are post-mitotic (cannot replace themselves;

neuronal stem cell replacement is inefficient)

Neuronal death caused by loss of proper connections,

loss of proper growth factors (e.g. NGF), and/or
damage (especially oxidative damage)

Neuronal dysfunction or damage results in loss of synapses

or loss of cell bodies
(synaptosis, can be reversible; apopsosis, irreversible)

PARKINSON'S DISEASE
ALZHEIMER'S DISEASE
HUNTINGTON'S DISEASE etc.
 APOPTOSIS: Role in Disease
Cancer

Apoptosis eliminates damaged cells

(damage => mutations => cancer

Tumor suppressor p53 controls senescence

and apoptosis responses to damage

Most cancer cells are defective in apoptotic response

(damaged, mutant cells survive)

High levels of anti-apoptotic proteins

or
Low levels of pro-apoptotic proteins
===> CANCER
 APOPTOSIS: Role in Disease
AGING

Aging --> both too much and too little apoptosis

(evidence for both)

Too much (accumulated oxidative damage?)

---> tissue degeneration

Too little (defective sensors, signals?

---> dysfunctional cells accumulate
hyperplasia (precancerous lesions)
OPTIMAL FUNCTION (HEALTH)

APOPTOSIS

AGING

APOPTOSIS
Neurodegeneration, cancer, …..
INTRACELLULAR
ACCUMULATIONS
 Objectives
To study:
• Overview of intracellular accumulations
• Accumulation of Lipids
• Accumulation of Cholesterol
• Accumulation of Proteins
• Accumulation of Glycogen
• Accumulation of Pigments
• Pathologic Calcification
 Overview
• Under some circumstances cells may
accumulate abnormal amounts of
various substances,.
• They may be harmless or associated
with varying degrees of injury .
 Overview
• May be found:
– in the cytoplasm
– within organelles (typically lysosomes)
– in the nucleus
• Came to the cell through:
– Synthesis by affected cells
– Produced elsewhere
 Pathways
1. Normal or increased rate of production of a
normal substance, but metabolic rate is
inadequate to remove it (e.g. fatty change in
liver)
 Pathways
2. A normal or an abnormal endogenous
substance accumulates because of genetic
or acquired defects in its folding, packaging,
transport, or secretion.
e.g. In α-1antitrypsin deficiency,
α1at accumulates in the liver causing cirrhosis)
α-1antitrypsin deficiency
 Pathways
3. An inherited defect in an enzyme may result
in failure to degrade a metabolite.
The resulting disorders are called storage
diseases.
 Pathways
4. An abnormal exogenous substance is
deposited and accumulates because the cell
has neither the enzymatic machinery to
degrade the substance nor the ability to
transport it to other sites.
(e.g. Accumulations of carbon or silica particles)
 Lecture will include
• Overview of intracellular accumulations

• Accumulation of Lipids
• Accumulation of Cholesterol
• Accumulation of Proteins
• Accumulation of Glycogen
• Accumulation of Pigments
• Pathologic Calcification
Fatty Change
(Steatosis)
 Fatty Change
• Fatty change refers to any abnormal
accumulation of triglycerides within
parenchymal cells.
• Site: liver, most common site
– it may also occur in heart, skeletal muscle,
kidney, and other organs.
 Causes of Fatty Change
• Toxins (most importantly: Alcohol
abuse)
• diabetes mellitus
• Protein malnutrition (starvation)
• Obesity
• Anoxia
Starvation will
increase this Hepatotoxins (e.g. alcohol) by
disrupting mitochondria and
SER ; anoxia

•Defects in any of the

steps of uptake,
catabolism, or
secretion can lead to
lipid accumulation.

CCl4 and protein

malnutrition
The significance of fatty change

• Depends on the cause and severity of the

accumulation.
– Mild it may have no effect on cellular function.
– Severe fatty change may transiently impair cellular
function

– In the severe form, fatty change may precede cell

death, and may be an early lesion in a serious liver
disease called nonalcoholic steatohepatitis
 Morphology of fatty change

• Most common site: the liver and the heart.

• With increasing accumulation, the organ

enlarges and becomes progressively yellow,
soft, and greasy.
 Light of microscopy fatty change
• Early: small fat vacuoles in the
cytoplasm around the nucleus.

• Later stages: the vacuoles coalesce

to create cleared spaces that
displace the nucleus to the cell
periphery

• Occasionally contiguous cells

rupture (fatty cysts)
 Is Fatty liver reversible?
• Fatty change is reversible
except if some vital intracellular process
is irreversibly impaired (e.g., in CCl4
poisoning),
 Prognosis of Fatty liver
• Mild: benign natural history (approximately
3% will develop cirrhosis

• Moderate to sever: inflammation,

degeneration in hepatocytes, +/- fibrosis
(30% develop cirrhosis)

• 5 to 10 year survival:67% and 59%

 Lecture will include
• Overview of intracellular accumulations
• Accumulation of Lipids
• Accumulation of Cholesterol
• Accumulation of Proteins
• Accumulation of Glycogen
• Accumulation of Pigments
• Pathologic Calcification
Cholesterol and Cholesteryl
Esters
• Cellular cholesterol metabolism is tightly
regulated to ensure normal cell
membrane synthesis without significant
intracellular accumulation
Conditions associated with Cholesterol and
Cholesteryl Esters accumulation
Several different pathologic processes:

1. Macrophages in contact with the lipid debris

of necrotic cells or abnormal (e.g.,
oxidized) forms of lipoproteins
 1. Macrophages in contact with the
lipid debris
These macrophages may
be filled with minute,
membrane-bound
vacuoles of lipid,
imparting a foamy
appearance to their
cytoplasm (foam cells).
Foam cells
2. Atherosclerosis:
smooth muscle cells and macrophages are
filled with lipid vacuoles composed of
cholesterol and cholesteryl esters
 Atherosclerosis:
• These give
atherosclerotic
plaques their
characteristic yellow
color and contribute
to the pathogenesis
of the lesion
 Conditions associated with Cholesterol
and Cholesteryl Esters accumulation

3. In hereditary and acquired

hyperlipidemic syndromes,
macrophages accumulate intracellular
cholesterol

4. Xanthomas: clusters of foamy

macrophages present in the
subepithelial connective tissue of skin or
in tendons
 Objctives
To study:
• Overview of intracellular accumulations
• Accumulation of Lipids
• Accumulation of Cholesterol

• Accumulation of Proteins
• Accumulation of Glycogen
• Accumulation of Pigments
• Pathologic Calcification
Proteins
• Morphologically visible protein
accumulations are much less common
than lipid accumulations
• They may occur because excesses are
presented to the cells or because the
cells synthesize excessive amounts
 Protein accumulations
Example:
1. Nephrotic syndrome:

• In the kidney trace amounts of albumin

filtered through the glomerulus are normally
reabsorbed by pinocytosis in the proximal
convoluted tubules

• After heavy protein leakage, pinocytic

vesicles containing this protein fuse with
lysosomes, resulting in the histologic
appearance of pink, hyaline cytoplasmic
droplets
The process is reversible; if the proteinuria abates, the protein
droplets are metabolized and disappear.
 Protein accumulations
Example:
2. marked accumulation of newly
synthesized immunoglobulins that may
occur in the RER of some plasma cells,
forming rounded, eosinophilic Russell
bodies.
 Protein accumulations
Example:
3. Mallory body, or "alcoholic hyalin," is an
eosinophilic cytoplasmic inclusion in
liver cells that is highly characteristic of
alcoholic liver disease
These inclusions are
composed predominantly
of aggregated
intermediate filaments
 Protein accumulations
Example:
4. The neurofibrillary tangle found in the
brain in Alzheimer disease is an
aggregated protein inclusion that
contains microtubule-associated
proteins
 Lecture will include
• Overview of intracellular accumulations
• Accumulation of Lipids
• Accumulation of Cholesterol
• Accumulation of Proteins

• Accumulation of Glycogen
• Accumulation of Pigments
• Pathologic Calcification
Glycogen
 Glycogen
• Associated with abnormalities in the
metabolism of either glucose or glycogen.
• Examples:
1. In poorly controlled diabetes mellitus, glycogen
accumulates in renal tubular epithelium, cardiac
myocytes, and β cells of the islets of Langerhans.

2. Glycogen accumulates within cells in a group of

closely related genetic disorders collectively referred
to as glycogen storage diseases, or glycogenoses
• In these diseases, enzymatic defects in
the synthesis or breakdown of glycogen
result in massive stockpiling, with
secondary injury and cell death.
Pigments
• Pigments are colored substances that
are either:
– exogenous, coming from outside the body,
or
– endogenous, synthesized within the body
itself.
 Exogenous pigment
• The most common is carbon
• When inhaled, it is phagocytosed by
alveolar macrophages and transported
through lymphatic channels to the
regional tracheobronchial lymph nodes.
 Exogenous pigment
• Aggregates of the pigment blacken the draining
lymph nodes and pulmonary parenchyma
(anthracosis).
• Heavy accumulations may
induce emphysema or a
fibroblastic reaction that can
result in a serious lung
disease ( coal workers'
pneumoconiosis)
Asbestos bodies

Asbestos bodies
 Objectives
To study:
• Overview of intracellular accumulations
• Accumulation of Lipids
• Accumulation of Cholesterol
• Accumulation of Proteins
• Accumulation of Glycogen
• Accumulation of Pigments
• Pathologic Calcification
• Endogenous pigments include
lipofuscin, melanin, and certain
derivatives of hemoglobin.
 Lipofuscin
• "wear-and-tear pigment" is an insoluble brownish-
yellow granular intracellular material that seen in a
variety of tissues (the heart, liver, and brain) as a
function of age or atrophy.

• Consists of complexes of lipid and protein that derive

from the free radical-catalyzed peroxidation of
polyunsaturated lipids of subcellular membranes.

• It is not injurious to the cell but is important as a marker

of past free-radical injury.

• The brown pigment when present in large amounts,

imparts an appearance to the tissue that is called
brown atrophy.
 Lipofuscin

• By electron microscopy, the pigment appears

as perinuclear electron-dense granules
 Melanin
• is an endogenous, brown-black pigment
produced in melanocytes

• Although melanocytes are the only source of

melanin, adjacent basal keratinocytes in the
skin can accumulate the pigment
(dermal macrophages)
Melanin
 Hemosiderin
• is a hemoglobin-derived granular pigment
that is golden yellow to brown and
accumulates in tissues when there is a
local or systemic excess of iron.

• Hemosiderin pigment represents large

aggregates of ferritin micelles, can be
seen by light and electron microscopy
 Hemosiderin
• Although hemosiderin accumulation is
usually pathologic, small amounts of
this pigment are normal in the
mononuclear phagocytes of the bone
marrow, spleen, and liver, where there
is extensive red cell breakdown
 Hemosiderin
• Local excesses of iron, and consequently of
hemosiderin, result from hemorrhage.

• Bruise: The original red-blue color of

hemoglobin is transformed to varying shades
of green-blue by the local formation of
biliverdin (green bile) and bilirubin (red bile)
from the heme
 Hemosiderin
The iron ions of hemoglobin accumulate as golden-yellow
hemosiderin.

The iron can be unambiguously identified by the Prussian

blue histochemical reaction
 Hemosiderosis
• is systemic overload of iron, hemosiderin is
deposited in many organs and tissues

• It is found at first in the mononuclear

phagocytes of the liver, bone marrow, spleen,
and lymph nodes and in scattered
macrophages throughout other organs.

• With progressive accumulation, parenchymal

cells throughout the body (principally the liver,
pancreas, heart, and endocrine organs) will be
affected
 Hemosiderosis
• Hemosiderosis occurs in the setting of:

1.increased absorption of dietary iron

2.impaired utilization of iron
3.hemolytic anemias
4.transfusions (the transfused red cells
constitute an exogenous load of iron).
.
 Effect of hemosiderosis
• In most instances of systemic
hemosiderosis, the iron pigment does
not damage the parenchymal cells

• However, more extensive

accumulations of iron are seen in
hereditary hemochromatosis with tissue
injury including liver fibrosis, heart
failure, and diabetes mellitus
Cell Aging
• Aging is generally characterized by the declining
ability to respond to stress, increasing
homeostatic imbalance and increased risk of
aging-associated diseases.
• Death is the ultimate consequence of aging.
• Differences in maximum life span between
species correspond to different "rates of aging".
For example, inherited differences in the rate
of aging make a mouse elderly at 3 years
and a human elderly at 90 years.
• Genetic differences affect physiological
processes like efficiency of DNA repair,
antioxidant enzymes, rates of free radical
production etc
 Senescence
The process by which a cell looses its
ability to divide, grow, and function. This
loss of function ultimately ends in death.

• A degenerative process, only.

• Has no positive features.
• Traditionally aging was explained two theories

• Programmed theories imply that aging is

regulated by biological clocks operating
throughout the life span. This regulation would
depend on changes in gene expression that affect
the systems responsible for maintenance, repair
and defense responses.

• Stochastic theories blame environmental

impacts on living organisms that induce
cumulative damage at various levels as the cause
of aging, examples which range from damage to
deoxyribonucleic acid (DNA), damage to tissues
and cells by oxygen radicals (free radicals), and
cross-linking.
More recently theories of aging are categorized as
• Molecular Gene Theory
– Codon restriction
– Somatic mutation
– Gene regulation
• Cellular theory
– Free radical theory
– Wear and tear theory
– Apoptosis
– Senescence
• telomere loss (replicative senescence)
• cellular stress (cellular senescence).
• System theory
– Rate of living theory
– Neuro endocrine theory
– Immunologic theory
• Evolutionary theory
– Disposable soma
– Antagonistic pleiotropy
– Mutation accumulation
Molecular Gene Theories

• Codon restriction - Fidelity/accuracy of mRNA

translation is impaired due to inability to decode codons
in mRNA.
• Error catastrophe - Fidelity of gene expression
declines with age, resulting in increased fraction of
abnormal proteins.
• Somatic mutation - Accumulation of molecular
damage, primarily to DNA/genetic material.
• Dys-differentiation - Gradual accumulation of random
molecular damage impairs regulation of gene expression.
• Gene regulation - Aging caused by changes in gene
expression regulating both aging and development. Gene
expression protein folding and activity
Cellular Theories

• Free radical - Oxidative metabolism produces highly reactive

free radicals that subsequently damage protein and DNA.
Mitochondrial DNA Damage

• Wear and tear - Accumulation of normal injury – Glyco-

oxidation Theory of Aging (products from glucose with proteins
+ oxidation; AGE (advanced glycation endproducts
– Inflammation Theory of Aging

• Apoptosis - Programmed cell death resulting from intrinsic

damage and genetically determined events or genome crisis.

• Senescence - Phenotypes of aging are caused by an increase

in frequency of senescent cells. Senescence may be the result of
telomere loss (replicative senescence) or cell stress (cellular
senescence).
 Telomere shortening causes cell senescence

• Somatic cells usually lack telomerase activity,

which means that telomeres shorten with each
cell division.

• Cultured cells may go into crisis as the result

of reaching zero telomere length.

• Reactivation of telomerase enables cells to

survive crisis and to become immortal.
System Theories

• Rate-of-living - Assumes a fixed amount of metabolic

potential for every living organism (live fast, die young).

• Neuroendocrine - Alterations in neuroendocrine

control of homeostasis results in age-related
physiological changes also referred as
Neuroendocrine Theories of Aging

• Immunologic – decline of immune function with age

results in increased incidence of disease also referred
as Immunological Theory of Aging
Evolutionary Theories

• Disposable Soma - Somatic cells are maintained only

to ensure continued reproductive success, following
reproduction the soma is disposable. (life span theory)

• Antagonistic Pleiotropy - Genes that are beneficial at

younger ages are deleterious at older ages.

• Mutation Accumulation - Mutations that affect health at

older ages are not selected against
The Free Radical Theory of Aging
Oxygen free radicals generated cause
cumulative oxidative damage, resulting in
structural degeneration, (apoptosis),
functional decline, and age-related
diseases.

Some believe that oxidative stress is the

predominant cause of age-associated
degenerative change.
Oxidative Stress and Aging

• What happens when oxidant production

is greater then antioxidant defenses?

– Oxidative Stress
• DNA damage
• Protein damage
• Lipid Damage
Free Radical Theory
• - Oxidative metabolism produces highly reactive free
radicals that subsequently damage protein and DNA.
• Evidence from model organisms...
• - Superoxide dismutase (SOD) transgenes can extend the
life
span of Drosophila.
• - Chemicals that mimic catalase (peroxidase) activity can
extend
C. elegans life span.
• - Long-lived mutants are typically stress resistant, including
resistant to drugs (i.e. paraquat stress, which induces
increases
in free radicals)
• - Life span extension by insulin-like signaling mutants in C.
elegans requires catalase activity.
 p53 may affect aging

Key Concepts

• Shortening of telomeres below a critical length is

associated with reduced longevity.

• Increase of p53 above wild-type levels can decrease

tumor formation, but also decreases longevity.