Introduction to Pathology and Cell Injury

Question 1

What is disease?

Answer 1

A pathological condition of a body part, organ or system characterised by an identifiable group of signs and symptoms. Consequent morphological and functional disturbances.
Cell is central player.

Question 2

What is pathology and what are its disciplines?

Answer 2

Branch of medicine concerned with disease and understanding its processes - why patients experience symptoms, may often guide diagnoses.
Disciplines: chemical pathology, haematology, immunology, medical microbiology and cellular pathology (histopathology and cytopathology) - neuropathology, forensic & paediatric pathology.

Question 3

Give some examples of histology and cytology (how is sample taken)?

Answer 3

Histology - core biopsies, cancer resection specimens, excised skin legions.
Cytology - fine needle aspirates breast/thyroid, glands, urine, sputum etc.

Question 4

State some differences between histology and cytology.

Answer 4

Histology is often therapeutic as well as diagnostic, architecture is assessed as well as cell atypia, may differentiate invasive from in situ disease, provides information on completeness of excision, grading and staging, better for immunohistochemical and molecular testing, whereas Cytology is faster and cheaper, non/minimally invasive and safe, can be used for cells in fluids, sometimes preliminary to other investigations, higher inadequate and error rates, generally used to confirm/exclude cancer/dysplasia instead of diagnosing other conditions with accuracy.

Question 5

What might a histopathologist do to arrive at a diagnosis?

Answer 5

Pattern recognition, ask: is it normal? Inflammatory or neoplastic? Malignant or benign? Primary tumour or metastasis?

Question 6

If cancer is seen, what may be determined?

Answer 6

Type, stage, grade, completeness of excision or which margins are involved and likely efficacy of further treatments, all of which influence decisions on further management.

Question 7

How may a sample show if Herceptin is a worthwhile treatment?

Answer 7

Absence/presence of Her2 receptors.

Question 8

What do you need for microscopy?

Answer 8

Slices thin enough to see through, stained.

Question 9

What fixes the problem of autolysis?

Answer 9

Fixatives, usually formalin for 24-48hrs. Inactivate tissue enzymes and denature, prevent bacterial growth and harden tissue.

Question 10

When a pathologist chooses the sample, what does she do?

Answer 10

Cut up and put in a cassette.

Question 11

What is paraffin wax used for and how is it introduced?

Answer 11

It is used as a hardening agent.
First there is dehydration of the sample with alcohol in a vacuum, then the alcohol is replaced with xylene, which can mix with and is replaced by molten paraffin wax - even gets inside the cell.
It is embedded overnight with processors.

Question 12

What does the process of ‘blocking’ entail?

Answer 12

Getting tissue in a piece of wax that can be cut using metal trays.

Question 13

How thin are samples cut and by what?

Answer 13

A microtome cuts the tissue into 3-4 micron thin sections, which can float on water as a ribbon and then are picked up by microscope slides.

Question 14

Why is colouring of a sample necessary and what are popular stains?

Answer 14

Staining is used so structures are recognisable under a microscope.
Haematoxylin stains nuclei purple and eosin stains cytoplasm and connective tissue pink.

Question 15

How is a slice preserved and protected?

Answer 15

Mounting (mounting medium dries and hardens) and putting a coverslip on top.

Question 16

How does immunohistochemistry work?

Answer 16

Demonstrates substances by labelling any antigenic substance with specific antibodies joined to enzymes catalysing colour changing reactions.

Question 17

What are cytokeratins?

Answer 17

Cytokeratins are a family of intracellular fibrous proteins found Ian almost all epithelia. They give information about the primary site of carcinomas.

Question 18

What is molecular pathology?

Answer 18

How disease is caused by alterations in normal cell biology - may be to do with altered DNA, RNA or a protein, so FISH is an example of an investigative technique.
Sequencing of DNA from a tumour can show if a mutation is present in a particular gene.
mRNA ‘signatures’ may predict how a tumour is likely to behave.

Question 19

When might frozen sections be used and why? What are some faults?

Answer 19

Urgent histopathology as it’s a method or hardening tissue quickly. It may occur intraoperatively to establish the presence and nature of a lesion and influence the course of the operation.
It is not routinely used as the morphology is not as good as with paraffin wax sections - misinterpretation may be a problem, as well as absence of any diagnostic tissue in the frozen section.

Question 20

Cells have mechanisms to deal with mild changes in environmental conditions, but what may more severe changes lead to?

Answer 20

Cell adaptation, injury or death, the degree of which depends on the severity of the injury and the type of tissue.

Question 21

What may cell injury be caused by?

Answer 21

Hypoxia, toxins, physical agents (direct trauma, extreme temperatures, changes in pressure, electrical currents), radiation, microorganisms, immune mechanisms, dietary insufficiency or excess.

Question 22

What are the 4 causes of hypoxia (decreased oxygen supply)?

Answer 22

1. Hypoxaemic hypoxia - arterial content of oxygen is too low.
2. Anaemia hypoxia - decreased ability of haemoglobin to carry oxygen.
3. Ischaemic hypoxia - interruption of blood supply.
4. Histiocytic hypoxia - inability to utilise oxygen in cells due to disabled oxidative phosphorylation enzymes.

Question 23

What determines how long hypoxia takes to damage cells?

Answer 23

The type of tissue e.g. Neurones may only take a few minutes whereas fibroblasts can last for a few hours.

Question 24

How may the immune system damage the body’s cells?

Answer 24

Hypersensitivity reactions or autoimmune reactions.

Question 25

Which cell components are most susceptible to injury?

Answer 25

Cell membranes (plasma membrane and those around organelles), nucleus (DNA), proteins (structural and enzymes) and mitochondria.

Question 26

What happens in an ischaemic cell to cause a loss of microvilli, blebs, ER swelling and myelin figures?

Answer 26

Lack of blood supply means there’s no oxygen for oxidative phosphorylation, so less ATP and the NaKATPase stops working, so there’s cellular swelling resulting in the above.

Question 27

What affect on the cell does increased glycolysis (as a result of hypoxia), decreased pH and glycogen have on the cell?

Answer 27

Clumping on nuclear chromatin, also detachment of ribosomes from RER, so reduced protein synthesis and lipid deposition.

Question 28

What makes prolonged hypoxia irreversible?

Answer 28

The introduction of Ca2+ in to the cell.

Question 29

Which 2 insults to the cell, other than hypoxia, attack membranes primarily?

Answer 29

Frost bite and free radicals.

Question 30

What are free radicals?

Answer 30

Reactive oxygen species with single unpaired electrons in their outer shells. The unstable configuration reacts with other molecules to produce more free radicals.

Question 31

Which 3 free radicals have particular cellular, biological significance?

Answer 31

OH. hydroxyl, most dangerous.
H2O2. Hydrogen peroxide
O2- superoxide

Question 32

How are free radicals produced?

Answer 32

As part of normal metabolic reactions, inflammation for oxidative bursts of neutrophils, by radiation, contact with unbound metals in the body (Fe in haemochromatosis or Cu in Wilson’s disease) or drug metabolism e.g. that of paracetamol or carbon tetrafluoride by CYP450s in the liver.

Question 33

What affect does the antioxidant system have on free radicals? What has a similar role?

Answer 33

Vitamins A, C and E donate electrons to free radicals.
Other control mechanisms include metal carrier and storage proteins sequestering Fe and Cu and some enzymes that neutralise free radicals.

Question 34

What is oxidative imbalance?

Answer 34

If the number of free radicals overpowers the antioxidant system, they will target lipids in cell membranes, taking an electron and causing an autocatalytic chain reaction. Free radicals may also oxidise proteins, carbohydrates and DNA - it becomes bent out of shape/broken/cross linked, so mutagenic and therefore carcinogenic.

Question 35

How do heat shock proteins in the cell aim to protect itself?

Answer 35

Response aims to ‘mend’ misfolded proteins to maintain cell viability. (Unfoldases or chaperonins e.g. Ubiquitin)

Question 36

How do hypoxic cells appear under a light microscope?

Answer 36

Injured cells are pale and swollen with water, but when dead appear very pink with eosin, as denatured proteins have coagulated. Pyknosis (condensation of chromatin), Karyorrhexia (destructive fragmentation of nucleus) or Karyolysis may be seen.

Question 37

How do hypoxic cells appear under an electron microscope?

Answer 37

Blebs are seen as the cytoskeleton is broken down by proteases- calcium has acted as a coenzyme, chromatin clumps, mitochondria and EAR swell, ribosomes ping off, myelin figures collect under the cell membrane and lysosomes membranes leak then completely ruptures- cell digests itself.

Question 38

How may a testing function diagnose cell death?

Answer 38

Add dye and see if cell lets it inside.

Question 39

What is oncosis?

Answer 39

Cell death with swelling, the spectrum of changes that occur in injured cells prior to death.

Question 40

What is necrosis?

Answer 40

In a living organism, the morphological changes that occur after a cell has been dead for some time. (Seen 12-24hrs later)

Question 41

What are the 2 main types and the 2 special types of necrosis?

Answer 41

Main: coagulative and liquefactive necrosis.
Special: caseous and fat necrosis.

Question 42

What causes coagulative necrosis and how might it be spotted when?

Answer 42

Protein denaturalising is the cause in e.g. ischaemia of solid organs (with lots of CT support).
Cellular architecture is somewhat preserved, so it may be seen as a ‘ghost outline’ of cells e.g. After an MI.

Question 43

What causes liquefactive necrosis and how? What may be seen?

Answer 43

Liquefactive necrosis is caused by enzyme release e.g. Ischaemia in loose tissues leading to it’s enzymatic digestion.
Lots of neutrophils will be present.

Question 44

What type of necrosis causes amorphous/structureless debris and is associated with infections (particularly which one)?

Answer 44

Caseous necrosis, often associated with tuberculosis.

Question 45

Name 2 causes of fat necrosis.

Answer 45

In breast tissue it may be secondary to trauma and cause a non-cancerous tumour.
In pancreatitis, digestive enzymes leak out and attack fats in the abdominal cavity. Calcium reacts with fatty acids to produce calcium soaps.

Question 46

What is gangrene?

Answer 46

Appearance of necrosis visible to the naked eye.

Question 47

What is infarction?

Answer 47

Necrosis caused by reduction in arterial blood flow.

Question 48

What is an infarct?

Answer 48

An area of necrotic tissue resulting from an infarction - an area of ischaemic necrosis.

Question 49

What is the difference between dry and wet gangrene and which type does gas gangrene belong to?

Answer 49

Dry gangrene is necrosis modified by exposure to air (coagulative necrosis) whereas wet gangrene is necrosis modified by infection (liquefactive necrosis) - a type is gas gangrene, where infection is with anaerobic bacteria that produce gas (often found in soil).

Question 50

What may cause infarction?

Answer 50

The commonest causes of infarction are thrombosis and embolism, but tissue may also become infarcted by external pressure, such as in the instance of testicular torsion.

Question 51

Why are some infarcts white and some infarcts red?

Answer 51

White infarcts occur in solid organs where there is occlusion of an end artery; often a wedge shaped area of coagulative necrosis.
Red infarcts occur in loose tissue, which may have a dual blood supply, like the lungs or have numerous anastomoses, prior coagulation, raised venous pressure or reperfusion.

Question 52

The consequence of infarction can be none to death, which factors determine this?

Answer 52

Alternative blood supply, speed of ischaemia, tissue involved and oxygen content of blood.

Question 53

What is ischaemia reperfusion injury?

Answer 53

Damaged, not yet necrotic tissue, sustains more damage if blood flow returns, because of an increased production of oxygen free radicals with reoxygenation, increased neutrophils leading to inflammation and tissue injury, delivery of complement proteins and the activation of the complement pathway.

Question 54

As well as entering damaged cells, molecules may leak out with local and systemic effects, what are they?

Answer 54

Local inflammation, general toxic effect.

Question 55

Which molecules, after leaking out of a damaged cell, will appear in higher concentrations in the blood and so aid diagnosis? What other affects will they have?

Answer 55

Potassium leaks out (with tumour lysis from chemotherapy or severe burns) and causes cardiac arrest.
Enzymes eg creating kinase, which used to diagnose MI.
Myoglobin leaks out in RHABDOMYOLYSIS when there’s damage to skeletal muscle, with the myoglobin possibly causing kidney failure seen with myoglobinuria (brown urine).

Question 56

What is apoptosis?

Answer 56

Cell death with shrinkage, induced by a regulated intracellular program where a cell activates enzymes that degrade its own nuclear DNA and proteins.
Equal and opposite force to mitosis.
Active process.

Question 57

State some differences between the cell in apoptosis and in necrosis/oncosis.

Answer 57

In apoptosis the DNA breakdown is not random - intenucleosomal cleavage of DNA requires energy. The membrane integrity is maintained as lysosomal enzymes are not involved. It may be pathological or physiological. Budding of apoptosis bodies instead of blebbing - shrinking not bloating.
No adjacent inflammation in apoptosis - single cells targeted.

Question 58

When does apoptosis occur physiologically and when pathologically?

Answer 58

Physiologically to maintain steady state, hormone controlled involution (e.g. ovary shrinkage after menopause), embryogenesis (e.g. Sculpting of paddles).
Pathologically in the cytotoxic T cell killing of a virus inflected or neoplastic cell, when a cell has irreparable DNA damage, in Graft vs. Host disease after a bone marrow transplant.

Question 59

Intrinsic and extrinsic pathways can activate what type of enzymes, which control and mediate apoptosis by causing cleavage of DNA and proteins making up the cytoskeleton?

Answer 59

Caspases

Question 60

How may apoptosis be triggered intrinsically?

Answer 60

Irreparable DNA damage or withdrawal of growth factors/hormones activates the p53 protein (guardian of the genome), the outer mitochondrial membrane becomes leaky, cytochrome C is released causing activation of caspases.

Question 61

How may apoptosis be triggered extrinsically?

Answer 61

Cells that are a danger (part of tumour/virally infected) recognised, TNFalpha signal released by killer T cells, binds to plasma membrane (at ‘death receptors’), leading to caspase activation.

Question 62

Why do apoptotic bodies express proteins on their surface?

Answer 62

So that they can be recognised by phagocytes or neighbouring cells within which final degradation can occur.

Question 63

Why may a cell have abnormal accumulations?

Answer 63

If a cell can’t metabolise something it will remain inside. It may be derived from the cell’s own metabolism, extracellular space or the outer environment.

Question 64

What are the five main groups of abnormal cellular accumulations?

Answer 64

Water and electrolytes
Lipids
Carbohydrates
Proteins
Pigments

Question 65

What is hydropic swelling?

Answer 65

For instance in hypoxia when energy supplies are cut off, sodium and water flood in, which is a particular problem in the brain.
A sign of severe cellular distress - fluid accumulates.

Question 66

What is steatosis? Where and why?

Answer 66

Accumulation of triglycerides often seen in the liver (major organ of fat metabolism), asymptomatic if mild. May be caused by alcohol, diabetes, obesity and toxins. May get big, greasy, yellow liver.
Cholesterol can’t be broken down and is insoluble - only eliminated by liver, with excess stored in vesicles. Atherosclerotic plaques in smooth muscle cells and macrophages and it macrophages in the skin and at tendons - Hereditary hyperlipidaemias.

Question 67

How are proteins seen in the cytoplasm, for instance after Alcoholic liver disease?

Answer 67

Eosinphilic droplets - bright pink damaged keratin filaments.

Question 68

What is Mallory’s hyaline?

Answer 68

Alpha1-antitrypsin deficiency - liver produced it incorrectly, so the folding means it can’t be packaged correctly, so it’s not secreted, systematic, so proteins in lungs unchecked - emphysema.

Question 69

When pigments accumulate more in cells from urban air pollutants (soot, coal dust, carbon), what happens to them?

Answer 69

Ingested and phagocytosis by alveolar macrophages causing anthracosis (blackening of lungs) and blackened peribronchial lymph nodes.
Harmless until large amounts then fibrosis and emphysema.

Question 70

Describe an endogenous pigment.

Answer 70

Haemosiderin (yellow/brown) is an Fe storage molecule, derived from ahh which forms when there’s excess Fe - bruising or if systematic haemosiderosis - maybe result of haemolytic anaemia or blood transfusions.

Question 71

What is hereditary haemochromatosis?

Answer 71

Increased intestinal absorption of iron, with the treatment of repeated bleeding. ‘Bronze diabetes’ as haemosiderin forms. Iron deposited in skin, liver, pancreas, heart and endocrine organs causing heart dysfunction, liver damage and endocrine failures.

Question 72

What is jaundice?

Answer 72

Accumulation of bilirubin (check for bright yellow sclera), which is formed in all cells (breakdown product of haem), but must be eliminated in bile (taken from tissues by albumin, then conjugates and is secreted). Deposited in tissues extracellularly or in macrophages if bile flow obstructed/overwhelmed.

Question 73

Describe some mechanisms of intracellular accumulations.

Answer 73

Abnormal metabolism, alterations in protein folding and transport, deficiency of critical enzymes, inability to degrade phagocytosed particles.

Question 74

What happens in calcification of tissues?

Answer 74

Abnormal depositions of calcium salts.

Question 75

Is dystrophic (local) or metastatic calcification more common and where may it occur?

Answer 75

Dystrophic - occurs in area of dying tissue, atherosclerotic plaques, ageing/damaged heart valves, tuberculous lymph nodes, some malignancies.

Question 76

Metastatic calcification may be due to hypercalcaemia, secondary to disturbances in calcium metabolism, with crystal deposition in all tissues. It’s usually asymptomatic, but may be lethal (regress if cause corrected). What might the hypercalcaemia be caused by?

Answer 76

Increased secretion of PTH (parathyroid hormone) resulting in bone resorption (primary, secondary or ectopic) or destruction of bone tissue (e.g. from immobilisation or Paget’s). Either may be caused by a tumour.

Question 77

What may trigger local calcification?

Answer 77

A local change or disturbance favours nucleation of hydroxyapatate crystals, which may cause organ dysfunction.

Question 78

What is replicative senescence?

Answer 78

After a certain number of divisions, telomeres reach critical length.

Question 79

Which type of cells contain telomerase and why?

Answer 79

Germ cells and stem cells use it to avoid explication senescence, by retaining the original length of their telomeres, so they may continue to replicate indefinitely.
Many cancer cells produce it.