This document discusses pathology and methods in general and clinical pathology. It provides definitions of disease and death as basic notions in pathology. It also classifies pathogenic factors and discusses mechanisms of cellular injury, including depletion of ATP, damage to mitochondria, influx of calcium, accumulation of reactive oxygen species, and defects in membrane permeability. Autopsy and biopsy methods used in pathology are described.

1. The subject of pathology.
Methods in the general and clinical
pathology.
Basic notions – disease/death.
Classification of the pathogenic
factors.
Radina Ivanova, MD, PhD
Associate Professor of PathologyAssociate Professor of Pathology
Medical University of SofiaMedical University of Sofia

2. Synopsis
 Subject of pathological anatomy. General and
clinical pathology.
 Main notions in pathology. Biopsy and
necropsy methods.
 Biopsy examination in different ways of
obtaining material and tissue processing.
 Cellular injuries – etiology.
 General mechanisms of cellular injury –
ischemia and hypoxia.
 General mechanisms of cellular injury - free
radicals injury.

3. PATHOLOGY
 Literally translated, pathology is the
study (logos) of suffering (pathos).
Pathos (suffering)
Logos (study)

4. PATHOLOGY
 Hartman (19th
century)
 theory of the diseases
 It involves the investigation of the causes
(etiology) of disease as well as the
underlying mechanisms (pathogenesis) that
result in the presenting signs and symptoms
of the patient
 to understand structural and functional changes
in cells, tissues and organs

5. Pathology
Clinical practice
Bridging Subject
Basic science
 It is a discipline that bridges
clinical practice and basic science
PATHOLOGY

6. Rudolph Virchow
1821-1902
The Father of
Modern Pathology
• “All diseases are the results of
visible cell abnormalities”, i.e.,
abnormal histology, i.e.,
histopathology.
History of pathology
• Morgagni, 1761- founder of pathological anatomy
• Bishat, 1800 – founder of pathological histology

7. PATHOLOGY
 GENERAL
 fundamental cellular and tissue responses
to pathologic stimuli.

E.g.. Inflammation, cancer, ageing.
 SYSTEMIC
 particular responses of specialized organs

E.g.. Goiter, pneumonia, breast cancer.

8. Basic notions in pathology
 Disease
 Death

9. What is Disease?
 dis + ease (not at ease…)
 Disease
 “Expression of discomfort due to structural or
functional abnormality”
 Health
 Complete physical, mental and social well being,
not merely absence of disease…

10. What is Disease?
 Disease
 A condition of impaired dynamic equilibrium of the
organism with the environment

Biologic processes in which the function and structure of the
organs are abnormal
 Health
 A condition of dynamic equilibrium of the organism with the
environment

Homeostasis – internal equilibrium of different organs and
systems

11. Disease Types:
 Inflammatory / Neoplastic / Degenerative
 Acute / Chronic
 Acute – short days to weeks.
 Chronic – long, months to years.
 Congenital / Familial / Acquired
 Genetic / Environmental
 Mild / Moderate / Severe

12. DISEASE
 Each disease - 4 aspects
 Etiology (“cause”)
 Pathogenesis (“insidious development”)
 Morphology (abnormal anatomy)
 Clinical expression

13. Etiology: What is the cause?
 Environmental agents
 Physical
 Chemical
 Nutritional
 Infections
 Immunological
 Psychological
 Genetic Factors
Multifactorial:Multifactorial:
Diabetes,Diabetes,
HypertensionHypertension
CancerCancer

14. One agentOne agent  One diseaseOne disease - Malaria- Malaria
Several agentsSeveral agents  One diseaseOne disease -- DiabetesDiabetes
One agentOne agent  Several diseasesSeveral diseases - Smoking- Smoking
Disease
Disease
Disease
DiseaseDisease

15. Pathogenesis
 Development (underlying mechanisms)
 “Sequence of events in the cells and tissues to a
stimulus/pathogen” starting from the initial
stimulus to the ultimate expression of disease.”

16. Morphology
 To render diagnoses, pathologists identify:
 changes in the gross or microscopic appearance
(morphology) of cells and tissues
 Other changes (molecular)

17. Clinical Significance
 Patient Signs and symptoms are related to
underlying pathology…
 Prognosis of disease depends on inside
pathology
 Is he going to recover or die soon?
 Guided therapy
 What treatment is suitable for this patient?

18. Outcome of disease
 Restitutio ad integrum (recovery)
 Substitutio (not fully recovery,
chronic disease)
 Death

19. What is death?
 Clinical death-minutes
 medical term for cessation of blood
circulation and breathing

cardiopulmonary resuscutation
 Biological death
 irreversible cessation of circulatory and
respiratory functions and of all functions of
the entire brain (areflexia)

20. Late signs of death
 Algor mortis -cooling of the body after death
 Depends on body temperature at the time of death and
environmental temperature
 Livor mortis –discoloration
 the red-purple discoloration caused by the settling of the
blood in the dependent portions of the body due to gravity
 Rigor mortis
 the stiffening of the muscles after death due to chemical
changes in the muscle fibers (depletion of ATP, lowered pH)
 The decomposition of a body – several stages
 2-3 days -green staining on the right side of the abdomen,
smell

21. Methods in the general and
clinical pathology
 Autopsy
 hospital (pathologo-anathomic)

performed by pathologists in hospitalized patients who
died of natural causes
 forensic

investigate the death in cases of violence or presumed
violence
 Biopsy
 histological
 cytological

22. Autopsy
 Aims
 to identify the cause of death
 to clarify cases without clinical diagnosis or in those in
which the patient death was unexpected
 recognition of the effect of the treatment in the evolution
of the disease.
 recognition of new diseases and of new lesion patterns
 source of information, allowing the making of precise
statistics on the most frequent diseases
 material for the residents, students and staff learning
 material for scientific research.

23. Autopsy-performance
 Stages
 External examination
 Dissection -instrumentarium

internal examination (necropsies)
 Pathology protocol

Macroscopic diagnosis

Final pathologcal diagnosis

24. Biopsy
 Histological investigation-pieces of tissue
 Surgery

Frozen sections (gefrieren)
 Endoscopic

Bronchoscopy, gasto-, colonoscopy, etc
 Large needle biopsy

Liver, ren, pulmo, prostate
 Cytological investigation – isolated cells, smears
 Body fluids – ascites, urine, pleural effusions
 Fine needle biopsy of thyroid, prostate, lymph nodes
 Vaginal smears (PAP)

25. Processing of biopsies
 Histological tissue processing
 Formalin-10%
 Paraffin embedding-block
 Microtome cutting – 5 µ
 Staining of sections

HE

Special (histochemical) stainings
 Van Gieson, Gomori, Sudan, PAS, Perls,
congo-rot
 Cytological smears
 Drying on air/alcohol fixation
 Staining

MGG, Papanicolau

26. Other methods in pathology
 Electron microscopy
 Immunofluoresce
 Immunohistochemistry
 Morphometry/flow
cytometry
 DNA
 Molecular pathological
methods
 In situ hybridization
 PCR

27. Pathogenic factors
Classification
1. Oxygen deprivation (hypoxia, ischemia)
2. Chemical agents and drugs
3. Physical agents
4. Infectious agents
5. Immunologic reactions
6. Nutritional imbalances
7. Genetic derangements
8. Aging

28. 1. Hypoxia
 = or oxygen deficiency
 an extremely important and common cause of cell
injury and death.
 should be distinguished from ischemia ( a loss of
blood supply in a tissue due to impeded arterial flow
or reduced venous drainage).
 Reasons for oxygen deprivation
 Ischemia
 inadequate oxygenation of the blood (pneumonia)
 reduction in the oxygen-carrying capacity of the
blood (anemia)
 carbon monoxide (CO) poisoning (CO forms a stable
complex with hemoglobin that prevents oxygen
binding.)

29. 2. Chemical Agents
 An enormous number of chemical substances can
injure cells
 Concentrated glucose or salt
 Oxygen at sufficiently high partial pressures
 Poisons and potentially toxic agents (air pollutants,
insecticides, CO, ethanol)
 Therapeutic drugs
 if used excessively or inappropriately

30. Pathogenic factors
3. Physical Agents
 Trauma, extremes of temperatures, radiation, electric
shock, sudden changes in atmospheric pressure
4. Infectious Agents
 Viruses, tapeworms, rickettsiaes, bacteria, fungi,
protozoas.
5. Immunologic Reactions
 autoimmune reactions against one's own tissues
 allergic reactions against environmental substances in
genetically susceptible individuals

31. Pathogenic factors
6. Nutritional Imbalances
 Nutritional deficiencies - protein-calorie insufficiency,
specific vitamin deficiencies
 Excesses of nutrition
obesity –DM
diets rich in animal fat - atherosclerosis
7. Genetic Defects
 congenital malformations associated with Down
syndrome, sickle cell anemia
 deficiency of functional proteins -enzymes in inborn
errors of metabolism
8. Aging
 alterations in replicative and repair abilities of individual
cells and tissues

32. Cellular responses to stress and
pathogenic factors
 Normal cells have a
fairly narrow range of
function or steady state
 Homeostasis
 Excess physiologic or
pathologic stress may
force the cell to a new
steady state
 Adaptation
 Too much stress
exceeds the cell’s
adaptive capacity
 Injury

33. Cell injury
 Reversible/Irreversible
 Reversibility depends
on the type, severity
and duration of injury
 Cell death is the result
of irreversible injury
Responses of the heart to different types of
stress:
• Hypertension – hypertrophy (adaptation)
• Ischemia
•incomplete occluded coronary artery – cell injury
•complete or prolonged occlusion – cell death

34. Mechanisms of cell injury
1. Depletion of ATP
2. Damage to Mitochondria
3. Influx of Calcium
4. Accumulation of Oxygen-Derived Free Radicals (Oxidative Stress)
5. Defects in Membrane Permeability
5. Damage to DNA and Proteins

35. 1. Depletion of ATP
 ATP - the energy store of cells
 required for almost all
synthetic and degradative
processes within the cell

membrane transport, protein
synthesis, lipogenesis
 The major causes of ATP
depletion
 reduced supply of oxygen and
nutrients,
 mitochondrial damage,
 toxins (e.g., cyanide).

36. 2. Damage to Mitochondria
 Mitochondria -critical players in cell
injury and death.
 Mitochondria can be damaged by
 ↑ cytosolic Ca2+,
 reactive oxygen species (ROS)
 oxygen deprivation (hypoxia, toxins).
 Two major consequences of
mitochondrial damage:
 loss of mitochondrial membrane potential and
pH changes, resulting in failure of oxidative
phosphorylation and progressive depletion of
ATP
 leakage of cytochrome c and other proteins
into the cytosol and death by apoptosis.

37. 3. Influx of Calcium
 Increased cytosolic Ca++
 activates a number of enzymes,
with potentially deleterious cellular
effects
 induction of apoptosis by direct
activation of caspases and by
increasing mitochondrial
permeability
 Increase in cytosolic Ca++
by:
 increased influx across the
plasma membrane
 ischemia and certain toxins
 release of Ca2+ from the
intracellular stores

38. 4. Accumulation of Oxygen-Derived Free
Radicals (Oxidative Stress)
 Free radicals have an unpaired electron in their outer orbit
 cause autocatalytic reactions (molecules that react with free radicals
are in turn converted into free radicals, thus propagating the chain of
damage)
 Oxidative stress – a condition called in ↑ production of
ROS or ↓ degradation - an excess of free radicals
 Generated by:
 Absorption of radiant energy
 Oxidation of endogenous and exogenous compounds
 Oxidation of exogenous compounds

39. 4. Accumulation of Oxygen-Derived Free
Radicals (Oxidative Stress)
 Reactions relevant for
cell injury by ROS
 Lipid peroxidation 
damage to cellular and
organellar membranes
 Protein cross-linking and
fragmentation due to
oxidative modification of
amino acids and proteins
 DNA damage due to
reactions of free radicals
with thymine

40. 5. Defects in Membrane Permeability
 The plasma membrane can be
damaged by:
 ischemia,
 various microbial toxins,
 lytic complement components,
 physical and chemical agents.
 The most important sites of
membrane damage during cell injury
are:
 Mitochondrial membrane damage
 Plasma membrane damage
 leads to loss of osmotic balance and influx
of fluids and ions, as well as loss of cellular
contents.
 Injury to lysosomal membranes
 leads to leakage of their enzymes into the
cytoplasm and enzymatic digestion of cell
components, necrosis.

41. 6. Damage to DNA and Proteins
 Damage to DNA and
proteins in:
 radiation injury
 oxidative stress,
 inherited mutations
 Cells have mechanisms
that repair damage to DNA
 if this damage is too
severe to be corrected the
cell initiates its suicide
program and dies by
apoptosis.

42. INJURY MECHANISMS IN
REVERSIBLE CHANGES
 Decreased ATP
 Mitochondrial damage
 Increased intracellular calcium
 Increased free radicals
 Increased cell membrane permeability
(cellular swelling)

43. INJURY MECHANISMS IN
IRREVERSIBLE CHANGES
 Mitochondrial irreversibility
 Irreversible membrane defects
 Lysosomal digestion

44. General mechanism of cellular injury
in hypoxia and ischemia
 ↓ intracellular generation of ATP
 failure of many energy-
dependent cellular systems

ion pumps

depletion of glycogen stores,
with accumulation of lactic acid
and lowering the intracellular
pH

reduction in protein synthesis
 If hypoxia continues, worsening
ATP depletion causes further
deterioration
 If oxygen is restored, all of these
disturbances are reversible.
 If ischemia persists, irreversible
injury and necrosis ensue.