Lecture 4Lecture 4

Morphological Alterations in Cell Morphological Alterations in Cell Injury and NecrosisInjury and Necrosis

Associate Professor Dr. Alexey Podcheko

Spring 2015

A routine H&E histologic section from an A routine H&E histologic section from an irregular white area within the anterior wall of the irregular white area within the anterior wall of the heart of a 54-year-old female who died secondary heart of a 54-year-old female who died secondary to ischemic heart disease reveals the myocytes to ischemic heart disease reveals the myocytes to be replaced by diffuse red material. This to be replaced by diffuse red material. This material stains blue with a trichrome stain. Which material stains blue with a trichrome stain. Which one of the listed statements correctly describes one of the listed statements correctly describes this material?this material?

It is secreted by fibroblasts and has a high content of glycine It is secreted by fibroblasts and has a high content of glycine and hydroxyprolineand hydroxyproline

It is secreted by endothelial cells and links macromolecules to It is secreted by endothelial cells and links macromolecules to integrins integrins

It is secreted by hepatocytes and is mainly responsible for It is secreted by hepatocytes and is mainly responsible for intravascular oncotic pressureintravascular oncotic pressure

It is secreted by monocytes and contains a core protein that is It is secreted by monocytes and contains a core protein that is linked to mucopolysaccharideslinked to mucopolysaccharides

INTENDED LEARNING INTENDED LEARNING OUTCOMESOUTCOMES

1.1. To know at least 4 morphological signs To know at least 4 morphological signs of reversible injuryof reversible injury

2.2. To know 3 morphological features of To know 3 morphological features of necrosisnecrosis

3.3. List 6 morphologic patterns of necrosis List 6 morphologic patterns of necrosis with clinical exampleswith clinical examples

Stages of the cellular response to stress and injurious stimuli

Common Mechanisms Leading to Cell Injury and Death

1 2 3 4 5

What are the signs of REVERSIBLE and IRREVERSIBLE injury?

Sequential development of biochemical and morphologic changes in cell injury

•There is a time lag between the stress and the morphologic changes of cell injury or death•Light microscopy changes of cell death visible only 4-12 hours after total ischemia

List of Reversible Injury List of Reversible Injury Associated Morphologic ChangesAssociated Morphologic Changes

1. Acute swelling of the cell and cellular 1. Acute swelling of the cell and cellular organelles (mitochondria, nucleus)organelles (mitochondria, nucleus)

a. Formation of membrane blebsa. Formation of membrane blebsb. Detachment of polyribosomes from dilated b. Detachment of polyribosomes from dilated

ERERc. Clumping of nuclear chromatin c. Clumping of nuclear chromatin

(desegregation)(desegregation)2. Fatty changes (lipid vacuoles)2. Fatty changes (lipid vacuoles)3. Loss of Glycogen3. Loss of Glycogen

Acute swelling of the cellAcute swelling of the cellAcute swelling is the result of failure of energy-dependent ion Acute swelling is the result of failure of energy-dependent ion pumps in the plasma membranepumps in the plasma membrane

Gross View of Cellular Swelling (Edema) at Gross View of Cellular Swelling (Edema) at the level of whole organthe level of whole organ

Pallor, turgor and increased weight

Microscopic changes induced by cellular Microscopic changes induced by cellular edema (swelling)edema (swelling)

1. Appearance of clear vacuoles (distended ER)- vacuolar degeneration

2. Increased eosinophilic staining

Plasma membrane alterations, such as blebbing, blunting, and loss of microvilli

Mitochondrial changes, including swelling , ER swelling

Nuclear alterations, with disaggregation of granular and fibrillar elements, clumping of nuclear content

Ultrastructural (EM) changes shown on this slide:

1.Plasma membrane alterations

2.Mitochondrial changes

3.Dilation of ER4.Nuclear

alterations5.Myelin figures6.Loss of

microvilli

Myelin Figures Myelin Figures Ultrastructural changes (shown on this slide):

1.Plasma membrane alterations

2.Mitochondrial changes

3.Dilation of ER4.Nuclear

alterations5.Myelin figures6.Loss of

microvilli

Ultrastructural changes (shown on this slide):

1.Plasma membrane alterations

2.Mitochondrial changes

3.Dilation of ER4.Nuclear

alterations5.Myelin figures6.Loss of

microvilli

normal

swelling

Fatty Changes (Steatosis)Fatty Changes (Steatosis)

Liver Steatosis (Fatty Liver)Normal Liver

Definition: abnormal accumulation of triglycerides within cells

Mechanisms of hepatic steatosis

Etiology of Hepatic Steatosis:1. Obesity2. Diabetes Mellitus 3. Toxins (CCl4)4. Protein Malnutrition5. Hypoxia

NAFLD (non-alcoholic fatty liver disease) represents a spectrum of fatty liver diseases:1. simple steatosis2. steatosis with inflammation NASH : non-alcoholic steatohepatitis3. fatty liver disease with inflammation and fibrosis (severe NASH)4. cirrhosis.

Signs of reversible damage on Signs of reversible damage on cardiomyocytescardiomyocytes

Myofibril relaxation is an Myofibril relaxation is an early sign of reversible injury early sign of reversible injury in cardiac myocytes, which in cardiac myocytes, which occurs within the first 30 occurs within the first 30 minutes of severe ischemia. minutes of severe ischemia. Myofibril relaxation Myofibril relaxation corresponds with intracellular corresponds with intracellular ATP depletion and lactate ATP depletion and lactate accumulation due to accumulation due to anaerobic glycolysis during anaerobic glycolysis during this period. this period.

Approximate Time of Onset of Key Events in Ischemic Cardiac

Myocytes

Irreversible Morphologic ChangesIrreversible Morphologic Changes

Stages of the cellular response to stress and injurious stimuli

List of Irreversible Morphologic List of Irreversible Morphologic ChangesChanges

1.1. Rapture of lysosomal membranes (autolysis)Rapture of lysosomal membranes (autolysis)2.2. Rapture of cell membranesRapture of cell membranes3.3. Mitochondria: The appearance of vacuoles and Mitochondria: The appearance of vacuoles and

phospholipid-containing amorphous densities, phospholipid-containing amorphous densities, aggregates of fluffy material probably due to aggregates of fluffy material probably due to denaturation of proteinsdenaturation of proteins

4.4. Nuclear changes: Nuclear changes: a. Piknosis – degeneration of nuclear chromatina. Piknosis – degeneration of nuclear chromatin b. Karyorrexis – nuclear fragmentationb. Karyorrexis – nuclear fragmentation c. Karyolysis – dissolution of the nucleusc. Karyolysis – dissolution of the nucleus

Morphologic changes of cells in reversible and irreversible cell injury

Normal kidney tubules with viable

epithelial cells

Early (reversible) ischemic injury

showing surface blebs, increased eosinophilia of cytoplasm, and

swelling of occasional cells

Irreversible injury of epithelial cells, with

loss of nuclei, fragmentation of

cells, and leakage of contents

Q3: A 35-year-old man infected with hepatitis B Q3: A 35-year-old man infected with hepatitis B experiences mild nausea for about 1 week and experiences mild nausea for about 1 week and develops very mild scleral icterus. On physical develops very mild scleral icterus. On physical examination, he has minimal right upper quadrant examination, he has minimal right upper quadrant tenderness. Laboratory findings include a serum tenderness. Laboratory findings include a serum AST of 68 U/L, ALT of 75 U/L, and total bilirubin of 5.1 AST of 68 U/L, ALT of 75 U/L, and total bilirubin of 5.1 mg/dL. The increase in this patient's serum enzyme mg/dL. The increase in this patient's serum enzyme levels most likely results from which of the following levels most likely results from which of the following changes in the hepatocytes?changes in the hepatocytes?

(A) Autophagy by lysosomes(A) Autophagy by lysosomes

(B) Clumping of nuclear chromatin(B) Clumping of nuclear chromatin

(C) Swelling of the mitochondria(C) Swelling of the mitochondria

(D) Dispersion of ribosomes(D) Dispersion of ribosomes

(E) Defects in the cell membrane(E) Defects in the cell membrane

• Pyknosis (pyknos, dense) - condensation of chromatin, shrunken dark nuclei• Karyorrhexis - (rhexis, tearing apart) - fragmentation of nuclear material• Karyolysis - lysis of chromatin due to the action of endonucleases (loss of nuclear staining, DNA breaking down and disappearing)

Dead cells show typical nuclear changes

Karyolysis – dissolution of the nucleusKaryolysis – dissolution of the nucleus

Nuclear changes during irreversible cell injuryNuclear changes during irreversible cell injury

PyknosisPyknosis – degeneration of nuclear – degeneration of nuclear chromatinchromatin

KaryorrhexisKaryorrhexis – nuclear fragmentation – nuclear fragmentation

Three types of IRREVERSIBLE INJURY

Necrosis Apoptosis Autophagy

Q1: It is important to be able to distinguish reversible from irreversible injury. Which one of the following morphologic changes is irreversible:A. Ischemic induced glycogen depletionB. Acute cellular swellingC. Formation of membrane blebs D. Karyolysis E. Clumping of nuclear chromatin

Q2: A 54-year-old Caucasian male comes to the Q2: A 54-year-old Caucasian male comes to the emergency room with retrosternal chest pain of 30 emergency room with retrosternal chest pain of 30 minutes duration. The patient also complains of minutes duration. The patient also complains of sweating and mild dyspnea. A single tablet of sweating and mild dyspnea. A single tablet of nitroglycerin is delivered sublingually, and the patient’s nitroglycerin is delivered sublingually, and the patient’s pain decreases significantly. The patient has pain decreases significantly. The patient has experienced several similar episodes of pain over the experienced several similar episodes of pain over the last 12 hours, all of which resolved spontaneously. last 12 hours, all of which resolved spontaneously. Which of the following ultra structural changes would Which of the following ultra structural changes would most likely indicate irreversible myocardial cell injury in most likely indicate irreversible myocardial cell injury in this patient? this patient?

A. Myofibril relaxation A. Myofibril relaxation B. Disaggregation of polysomes B. Disaggregation of polysomes C. Mitochondrial vacuolization C. Mitochondrial vacuolization D. Disaggregation of nuclear granules D. Disaggregation of nuclear granules E. Triglyceride droplet accumulation E. Triglyceride droplet accumulation

Necrosis

• Necrosis=Denaturation of intracellular proteins +

enzymatic digestion of cell components/cell

•Occur in living cells

•Autolysis vs. Heterolysis

•Cytoplasmic Eosinophilia

• Nuclear Pyknosis, Karyolysis and Karyorrhexis

Distinctive Morphologic Patterns of Necrosis

1.Coagulative 2.Liquefactive 3.Caseous 4.Fat 5.Fibrinoid6.Gangrenous

Coagulative Necrosis

1. It is the most common form of necrosis (affect

all organs except brain)

2. Characteristic of ischemia

3. Infarct is a localized area of coagulative

necrosis

Coagulative Necrosis, Main Features

A. Architecture of dead tissues is preserved for at least several days due to the denaturing and coagulation of proteins within cytoplasm

B. Ghost outlines of cells but loss of nucleusC. Firm texture of affected tissuesD. Necrotic cells eventually are removed by

phagocytosis by infiltrating leukocytes

1 day 3-4 days 7 days

Miocardial Infarction

anterior wallTransmural MI

MI: pale myocardial infarctionMI: pale myocardial infarction

MI morphologyMI morphology-The infracted myocardial fibers are eosinophilic-there is no nuclear staining; -neutrophilic granulocytes accumulated atthe margins of infarction (vital sign)

Coagulative Necrosis

Spleen Infarct

Coagulative Necrosis

Spleen Infarct

Examples of Coagulative NecrosisExamples of Coagulative Necrosis

MIMI

Spleen InfarctSpleen Infarct

Kidney InfarctKidney Infarct

Liquefactive Necrosis

1.Characteristic of bacterially induced necrosis

2.Presence of large numbers of inflammatory cells with complete digestion of cells

3.Cellular destruction by hydrolytic enzymes4.Examples: abscesses in various organs and

tissues, brain and pancreas infarcts

Liquefactive Necrosis - Gross

MORE LIQUID MORE LIQUID MORE MORE WATER WATER MORE PROTONS MORE PROTONS

T2 weighted MRI images emphasize water density but some anatomic resolution is lost

Liquefactive Necrosis

Puss

Liver

Abscessing bronchopneumonia

Q4: A 68-year-old woman suddenly lost Q4: A 68-year-old woman suddenly lost consciousness; on awakening 1 hour later, she consciousness; on awakening 1 hour later, she could not speak or move her right arm and leg. could not speak or move her right arm and leg. Two months later, a head CT scan showed a Two months later, a head CT scan showed a large cystic area in the left parietal lobe. Which large cystic area in the left parietal lobe. Which of the following pathologic processes has most of the following pathologic processes has most likely occurred in the brain?likely occurred in the brain?

(A) Fat necrosis(A) Fat necrosis

(B) Coagulative necrosis(B) Coagulative necrosis

(C) Apoptosis(C) Apoptosis

(D) Liquefactive necrosis(D) Liquefactive necrosis

(E) Karyolysis(E) Karyolysis

Caseous Necrosis

1.Combination of coagulation and liquefaction necrosis

2.Gross: soft and “cottage-cheese-like” appearance

3.Characteristic of Lung Tuberculosis

Caseous Necrosis=TB

Caseous Necrosis

Q5: A chest radiograph of a 26-year-old man showed a Q5: A chest radiograph of a 26-year-old man showed a 4-cm nodule in the upper lobe of the left lung. The 4-cm nodule in the upper lobe of the left lung. The nodule was excised with a pulmonary wedge nodule was excised with a pulmonary wedge resection, and sectioning showed the nodule to be resection, and sectioning showed the nodule to be sharply circumscribed with a soft, white center. sharply circumscribed with a soft, white center. Culture of tissue from the nodule grew Culture of tissue from the nodule grew Mycobacterium tuberculosis. Which of the following Mycobacterium tuberculosis. Which of the following pathologic processes has most likely occurred in this pathologic processes has most likely occurred in this nodule?nodule?

(A) Apoptosis(A) Apoptosis(B) Caseous necrosis(B) Caseous necrosis(C) Coagulative necrosis(C) Coagulative necrosis(D) Fat necrosis(D) Fat necrosis(E) Fatty change(E) Fatty change

Fat Necrosis

• Distinct morphologic appearance and occurs in a distinct set of clinical circumstances – chalky white appearance• Action of pancreatic lipases on surrounding fatty tissues•Examples: 1. Acute pancreatitis 2. Trauma of fat tissue (breast)

Fat Necrosis

The areas of white chalky deposits represent foci of fat necrosis with calcium soap formation - saponification

Enzymatic Fat NecrosisEnzymatic Fat Necrosis

Damage to cells releases tryglycerides . The triglycerides are broken down by action of lipases to fatty acids. Fatty acids associate with calcium and then form calcium soaps (saponification)

Breast Fat NecrosisBreast Fat Necrosis

Calcification on mammography mimic Calcification on mammography mimic breast cancerbreast cancer

Fibrinoid NecrosisFibrinoid NecrosisNecrotic tissue that histologically resembles fibrinNecrotic tissue that histologically resembles fibrin

Micro: has an eosinophilic pink homogenous Micro: has an eosinophilic pink homogenous appearanceappearance

Fibrinoid necrosis in an artery (AI Vasculitis)

Gangrenous NecrosisGangrenous Necrosis

Gross term used to describe dead tissueGross term used to describe dead tissue

Common sites: lower limbs, gallbladder, Common sites: lower limbs, gallbladder, GI tract, testesGI tract, testes

Two types: Two types:

-Wet gangrene resembles liquefactive -Wet gangrene resembles liquefactive necrosis; necrosis;

-Dry gangrene resembles coagulative -Dry gangrene resembles coagulative necrosisnecrosis

Wet Gangrene (Diabetic angiopathy)

Gangrene = ischemic necrosis with bacterial superinfection

DRY GANGRENEDRY GANGRENE

1. To know at least 4 morphological signs of 1. To know at least 4 morphological signs of reversible injuryreversible injury

Objectives Review:

1.1. Acute swelling of the cell and mitochondriaAcute swelling of the cell and mitochondria2.2. Loss of microvilliLoss of microvilli3.3. Formation of membrane blebs, myelin figuresFormation of membrane blebs, myelin figures4.4. Detachment of ribosomes from dilated ERDetachment of ribosomes from dilated ER5.5. Clumping of nuclear chromatin (desegregation)Clumping of nuclear chromatin (desegregation)6.6. Fatty changes (lipid vacuoles)Fatty changes (lipid vacuoles)7.7. Loss of glycogenLoss of glycogen

2. To know 4 morphological features of 2. To know 4 morphological features of necrosisnecrosis

Objectives Review:

1.1. Rapture of lysosomal membranes (autolysis)Rapture of lysosomal membranes (autolysis)2.2. Rapture of cell membranesRapture of cell membranes3.3. Mitochondria: The appearance of vacuoles and Mitochondria: The appearance of vacuoles and

phospholipid-containing amorphous densities, phospholipid-containing amorphous densities, aggregates of fluffy material probably due to aggregates of fluffy material probably due to denaturation of proteinsdenaturation of proteins

4.4. Nuclear changes: Nuclear changes: a. Piknosis – degeneration of nuclear chromatina. Piknosis – degeneration of nuclear chromatin b. Karyorrexis – nuclear fragmentationb. Karyorrexis – nuclear fragmentation c. Karyolysis – dissolution of the nucleusc. Karyolysis – dissolution of the nucleus

3. List 6 morphologic patterns of necrosis 3. List 6 morphologic patterns of necrosis with clinical exampleswith clinical examples

Objectives Review:

Necrosis Pattern Example

1 Coagulative Myocardial Infarction (MI)

2 Liquefactive Brain Abscess

3 Caseous Lung Tuberculosis

4 Fat Acute Pancreatitis

5 Fibrinoid Autoimmune Vasculitis

6 Gangrenous Diabetic angiopathy