Introduction to Anatomical Pathology

Krisna Murti
Anatomical Pathology, Faculty of Medicine, Sriwijaya University

Historical Periods of Pathology

1. Anatomical (before XIX century)
J.

Morgagni (1685-1777)
K. von Rokitansky (1804-1878) performed nearly 30,000 autopsies, wrote an
outstanding monograph on diseases of arteries and congenital heart defects

2. Microscopic (XIX century - mid-XX century)

R.

Virchow (1821-1902) father of cellular pathology

3.Ultramicroscopic (mid-XX century)

4.Modern - pathology of a living person

Definition
Pathology (Anatomical Pathology / Pathologic Anatomy)
is fundamental biomedical science that studies the structural
basis of pathological processes of human disease

Anatomical Pathology is a medical laboratory specialty, whose

objective is the diagnosis of diseases through the study of biological
material obtained from organs or tissues and that may consist of
cells or fluids.

The Scope of Pathology

1. The core of pathology:
The four aspects of a disease process that form the core of pathology:
Epidemiology
(1) Etiology: causes of the disease
(2) Pathogenesis: the evolution/mechanisms of its development
(3) Morphologic changes: the structural alteration induced in the cells and organs
of the body
(4) Clinical significance: the functional consequences of the morphologic changes
Management
Complications
Prevention

2. Classification:
Autopsy
(1) Human pathology
Biopsy
Cytology
(2) Experimental pathology

3. Position:
Its a bridging discipline involving both basic science and
clinical practice

4. Text of Pathology:
(1) General pathology:
concerned with the basic reaction of cells and tissues to abnormal
stimuli that underlie all diseases

(2) Systemic pathology :

describe the specific responses of specialized organs and tissues
to defined stimuli

Techniques of Pathology
1. Human pathology
(1) Autopsy
(2) Biopsy: surgical or diagnostic pathology
(3) Cytology: smear, fine needle aspiration
2. Experimental pathology
(1) Animal experiment: animal model
(2) Tissue and cell culture

Morphological Alteration
Structural alteration of cells or tissuesusually specific for
particular diseases
Clinical symptoms: alteration of function due to morphological
changes
Morphological alterations of tissues / organs cause malfunction
of the organs and then raise clinical symptoms

Observation and New Technique of Morphology

Gross appearance:
size, shape
weight
color
consistency
surface
edge of section

Histologic and Cytologic Observation

most common and basic formalin fixed
HE (hematoxylin and eosin) stained

Hemangioma of ventrical wall

Methods of Pathological Anatomy

Biopsy
Removal of tissue from a living subject to determine morphological
changes;

Autopsy
Post-mortem examination of a corpse to determine the cause and manner
of death and to evaluate any disease or injury that may be present

Experiment
Modelling of pathological process on animals and subsequent postmortem examination

Types of Biopsies
During surgery
Puncture (needle
biopsy)
Aspiration
Excisional
Incisional
Scrape

Anatomical Pathology
Histopathology
Cytology
Gynecology: papsmear,
Non gynecology: FNAC, sputum, pleural effusion, Transthoracal biopsy
(TTB), transthoracal needle aspiration (TTNA)

Vriescope (VC)
Histochemistry
Imunohistochemistry
Molecular

Levels of Study

Organismal
Organ
Tissue
Cellular
Ultrastructural
Molecular

Histopathology
Macroscopic/gross
Direct examination by eyes or by touching
Organs :
Sizes : enlargement / smaller
Consistency : soft, hard, solid, fragile
Color : pale, yellow, brownish

Microscopy
Using microscope
Helped by staining for histochemistry or
immunohistochemistry to show specific chemical contains in
cells or tissues
Observe structural alterations in cells / tissues due to
particular diseases

Histopathological Materials
Biopsy: excise, endoscopy, cystoscopy
Operation : appendices, ovary, breasts
Extirpation : Lymph nodes
Curettage : specific for abnormalities in endometrium, cervix
and utery
Eg. abortus, mola hidatidosa, hormonal abnormality,
malignancies

Procedures
a. Formalin 10% buffer
b. The whole tissues rinse in fixation (volume of formalin is
10 X tissues volume)
c. If the tissues are too large lamellation every 1 cm to
ensure fixation enters the tissues
d. Processingmachines

Processing
Removal of alcohol with
xylene that will be miscible
with the embedding medium
(paraffin)
Impregnating with paraffin.

Haematoxylin-Eosin Staining (Routine)

Machine
Manual

Cytology
Cytos (cells) and logos (knowledge)
Exfoliative cytology: knowledge of cellsimportant for early
diagnosis of diseases / malignancies
Spontan exfoliation occur due to mature cells changed by
younger cells the normal behavior of superficial tissues /
organs

A. Structure of cells
. A cell is an important structure
. Different forms depending of the functions
. The cell from different tissues / organs has specific behavior

B. Structure of tissues

. Tissues consist of collection of cells

. Pap smearmany types of cells originated from different tissue types
of female genital tracts

Cytology
Cytology: to observe and examine structural alteration of every found cell
To detect malignancies: pap smear/pap test/ papanicolau test/cervical smear
early detection of cervical cancer
Genetic disorders
Hormonal disorders

a. Gynecologic: swab of lateral vaginal wall, cervix and endometrium

b. Non gynecologic: sputum (lung), ascites, pleural effusion

Gynecologic
1. Hormonal evaluation
2. Early detection of cancer
3. Inflammation detection

Procedures:
-. Lateral wall of vagina (1/3 inner wall)
-. Cervix
-. Endometrium

Pap Smear/Cervical Smear

The examination to detect abnormalities of cervix in particular
cervical cancer
Taken from fornix posterior/squamo-columnar junction
fixation : alcohol 95%
Staining : Papanicolau

Representative if Endocervical cells are present

Not representative/inadequate if :

Too little
Swab is too thick
Too much blood
Inadequate fixation
Endocervical cells are not present

Pap Smear

Non Gynekologic
1. Respiratory tracts
- sputum
- Bronchial washing
- TTB (transthoracic biopsy)
- TTNA
2. Digestive tracts endoscopy for stomach
3. Urinary tracts urine
4. Pleural cavity, pericard, sinovial, abdomen (ascites),
cerebrospinal fluid

Fine Needle Aspiration Cytology (FNAC)

Staining : PAP, HE, MGG

Aplications : superfisial and deep organs

Superfisial : lien, breasts, thyroid, peripheral lung, pleura, peritoneal,

sinovial, tumor
Deep organs: deep lung lesion, liver, kidenys, prostate
Superficial

Needle Biopsy
Deep

Procedures
Pap smear Fixation: alcohol 95%
The swab as soon as possible rinse into alcohol 95% for minimal 30 min
Urine/fluid from body cavities (pleural effusion, ascites)
- Can be sent as swab of sediment after centrifugation
- Fixation: alcohol 95%
- Or urine/pleural effusion/ascites about 100-200 cc fix with alcohol 50% aa
Sputum (swab like pap smear)
- Fixation: alcohol 95%, should be done for 3 x in 3 consecutive days

3. FNAC (fine needle aspiration cytology)

Dry with no fixation: dry the swab on the air without fixation then send
to the lab
Wet Fixation: swab rinses into alcohol 95% for minimal 30 min then
send to the lab

Frozen Section/Vries Coupe/VC

Examination of
histopathology while
the patient is in the
operation room
To ensure the
malignancy is
present or not then
therapy

Procedures in sending the specimens

1. Fill in forms completely
2. Send the specimens in adequate fixation
(formalin 10% buffer)

Histochemistry and Cytochemistry

PAS staining

Autopsy
Clinical autopsy is part of pathologists duties
Identify new diseases
Identify etiologies of death
Examination of the correctness of the diagnosis and treatment
Establish the cause of death
Research
Teaching students and physicians

Judicial Autopsy (Autopsi kehakiman)

Forensic
Criminal cases

Legal Authority of Autopsy

Autopsy is performed mandatory:
Suspicion of violent death
Death less then 1 day after admission to hospital
Death during surgery, diagnostic manipulations and/or
anesthesia
Death from infection
Suspicion of overdose or drug intolerance
Pregnant women, women in/after childbirth
Children under 1 year
Autopsy can be not performed on religious grounds

Terminologies
Alteration of epithelia in malignancy
Variation in nuclear size and forms (pleomorphism)
Alteration in nuclear chromatin: hyperchromatic, vesicular
Increase NC ratio
Nucleoli can be seen
: Metaplasia
Alteration of a mature cell type to another type
Eg: changes of columnar epithelial-squamous

Cont
Dyplasia: alteration of mature cell type to disorientation
Undifferentiated cell: not differentiate not growth to specialized types

Cont
Hyperchromatism: an increase in the histological staining, usually in the
nucleus
Hyperplasia: an increase in the amount of organic tissue that results from
cell proliferation
Hypertrophy: An enlargement of organs due to the increase of cell size
Physiology: athletes muscles, pregnant uteri
Pathology: hypertensioncardiac enlargement

Cont
Karyolysis: The complete dissolution of the chromatin of a dying cell due to the
enzymatic degradation by endonucleases
Karyorrhexis: the destructive fragmentation of the nucleus of a dying cell
whereby its chromatin is distributed irregularly throughout the cytoplasm
Carcinoma: Malignant neoplasm originated from epithelial
Pyknosis : the irreversible condensation of chromatin in the nucleus of cell
undergoing necrosis or apoptosis
Polymorphism : occur when two or more clearly different phenotypes exist in
the same population of a species the occurrence of more than one form or
morph or multiple alleles in a single gene
Pleomorphism : Occurring in various distinct forms. In terms of cells, having
variation in the size and shape of cells or their nuclei.

Flow Cytometry (FCM)

1. One kind of cellsquantitative analysis
2. DNA ploidy analysis
3. Protein nucleus acidquantitative analysis
4. Selection of collection of cells

Image analysis (IA)

Nuclei: diameter; circumference; area; volume; morphology

Laser scanning confocal microscope (LSCM)

Aliving cellobservation in situ or development or quantitative

Modern Methods in Morphology

Immunohistochemistry (IHC)
Electron microscopy
In situ hybridization (ISH)
Polymerase chain reaction (PCR)

Immunohistochemical techniques
Immunohistochemistry
It is based on specific interaction of tissue and cellular
antigens with a specially derived antibodies bearing the different
labels.
Immunohistochemistry (IHC)
Immunofluorescence (IF)

Opportunities IHC
Determination of cells belonging to a particular tissue;
Identification of individual products (e. g. abnormal
proteins), routes of cellular and intercellular signals,
synthesis of certain proteins, glyco-and lipoproteins

Immunohistochemistry
1. Ag-Ab specific reaction
2. Applications
(a) Location analysis
cytokeratincell membrane
(b) Clinical diagnosis and distinguishing
diagnosis of tumor histogenesis

Immunofluorescence (IF)

Ultrastructure Observation
Used to study the details of cell structure, detection of viruses, bacteria, immune complex
deposits
Examples of use:
Oncology - Birbeck granules in histiocytosis X
Oncology - Z-discs in rhabdomyosarcoma cells
Nephrology - diagnosis of glomerulonephritis
TEM (transmitting electron microscope)

SEM (scanning electron microscope)

In Situ Hybridization (ISH)

In situ hybridization (ISH)
Method for localizing / detection of specific sequences (of DNA or RNA in situ
/ directly in tissue specimens) in morphologically preserved tissue sections
or cell preparations by hybridizing the complementary strand of a nucleotide
probe against the sequence of interest.
It is based on principle of complementary interaction of DNA or RNA in
specimen with labelled nucleotide sequence (probe)
If nucleic acids are preserved in a histological specimen, then it can be
detected by using a complementary probe

Is used for:
Detection

of viral genomes
Detection of mutant genes
Detection of active protein synthesis
(unlike IHC which allows to determine the presence of a protein)
IHC data validation

Polymerase Chain Reaction (PCR)

Method for detection of specific sequences of DNA or RNA in any
biological sample
PCR is in vitro amplification (i. e. increase in the number of
copies) of nucleic acids triggered by synthetic oligonucleotide
primers
Differences from the in situ hybridization:
Because of amplification it is more sensitive (about 1
million times)
Usually not combined with morphology

Molecular Biology Technique

1. Polymerase chain reaction (PCR)
2. DNA sequencing
3. Biochip technique
(1) Gene chip (DNA chip)
(2) Protein chip (protein microarray)
(3) Tissue chip (tissue microarray)

Thank you