1. 2nd Year Pathology 2010 Extracellular Pathology Amyloidosis Pathological calcification Pathological calcification Ageing Ageing

2. Amyloid and Amyloidosis A Set of Disorders: The main feature is the extracellular deposition of proteins arranged in the form of a Beta-pleated sheet –“Amylum” Latin for starch –Term first used By Virchow in 1854 –Protein….Friedreich and Kekule. 5 years later

3. Amyloidosis Amyloidosis is a clinical disorder caused by extracellular deposition of insoluble abnormal fibrils that injure tissue. The fibrils are formed by the aggregation of misfolded, normally soluble proteins. In humans, about 23 different unrelated proteins are known to form amyloid fibrils.

4. Amyloidosis All types of amyloid consist of a major fibrillar protein that defines the type of amyloid (approximately 90%) plus various minor components. Although each type of fibril may be associated with a distinct clinical picture, all share certain physical and pathologic properties

5. Organs affected by amyloid: Larger, Paler, Firmer How do we identify Amyloid in tissues? Macroscopically (Grossly) Waxy, Sharper cut edges, Lugol’s iodine (black). Microscopically: Light Microscopy- Amyloid is eosinophilic and stains with Congo Red Polarized light: Apple Green Birefringence. Electron Microscopy: Rigid non branching fibrils (7.5 nm to 10 nm in diameter)

6. Kidney

7. Heart, Lugol’s iodine

8. H&E Stain

9. Congo Red Stain.

10. Apple Green Birefringence

11. Electron Microscope

12. The Beta- Pleated sheet is the unifying feature of the amyloidoses Basis of 1) Congo Red reaction 2) Fibrillar ultrastructure 3) Resistance to proteolytic digestion

13. Why? Protein Misfolding –Intrinsic tendency to misfold (Transthyretin) –Misfolding and aggregation at high concentration (Beta Microglobulin) –Point mutations (Hereditary amyloidosis) Review: Molecular mechanisms of amyloidosis. Merlini G, Bellotti V. NEJM August 2003 349:583-96

14. Classification Older classifications: 1° and 2° More useful to classify on the basis of: 1) Nature of protein involved 2) Anatomical distribution 3) Inherited or acquired

15. Acquired Systemic Amyloidosis 1) Amyloidosis of immune origin. AL type  Acquired, Systemic form  Protein precursor is usually immunoglobulin light chain.  Occurs in association with a monoclonal (i.e. neoplastic) proliferation of B lymphocytes or plasma cells e.g.  Myeloma (Lambda chains)  Waldenstrom’s macroglobulinaemia (LPL)

16. AL amyloid proteins : Intact immunoglobulin light chain or the amino terminal fragment of a chain or both. (mostly lambda chains) 90% of these patients have Bence Jones proteins Amyloid accumulation: Two step process 1) Secretion of excess amounts of monoclonal light chain 2) Conversion to Beta pleated form.

17. Clinical features Middle to old age M>F 90% have a Bence Jones protein Bone marrow = excess plasma cells Manifestations Neuropathy, Restrictive Cardiomyopathy, Skin manifestations, Polyarthropathy, Macroglossia, Carpal Tunnel Syndrome (Heart, CNS, Joints, Skin, Tongue, Soft Tissues)

18. Immunofluorescence microscopy with antibody to the lambda light chain.

19. Cardiac 2 nd Year Pathology 2009

21. Acquired Systemic Amyloidosis 2) Haemodialysis associated Amyloid  Acquired, systemic  Protein is Beta-2-microglobulin.  Normally broken down by kidneys  A naturally occurring “amyloidogenic” protein not filtered by dialysis membranes  After 7 years on dialysis 30% of patients get Carpal Tunnel Syndrome (after 10 years 50%)  Joints, synovium, tendon sheaths

22. Acquired Systemic Amyloidosis 3) Reactive Systemic Amyloidosis  Also known as secondary amyloid or AA type amyloid  Acquired, systemic form  Protein precursor is serum amyloid A (SAA)  Acute phase reactant which markedly increases with tissue injury or inflammation under the influence of IL1, TNF and IL-6  AA is a cleavage product of SAA.  Distribution of amyloid: kidney, liver, spleen, adrenals, thyroid + many other tissues.  Diagnosis: Rectal Biopsy

23. Disease Associations: 1) Chronic infection: T.B., Leprosy, Syphilis, Chronic osteomyelitis, Bronchiectasis 2) Chronic inflammation Reiter’s disease, Whipple’s disease 3) Chronic autoimmune disease: R.A., I.B.D., Connective Tissue Disease 4) Long standing paraplegia. (UTI) 5) Neoplasms: Renal adenocarcinoma and Hodgkin’s disease

24. Liver

25. Adrenal

26. Hereditary Systemic Amyloidosis Rare. Most common: Familial Mediterranean Fever. Less common: Three types: 1)Neuropathic. 2)Cardiopathic. 3) Nephropathic. The Amyloid protein is usually Transthyretin (Prealbumin). Transthyretin is also associated with a form of Amyloid known as Systemic Senile Amyloid, where amyloid is systemically deposited, mainly in the heart in elderly individuals.

27. Familial Mediterranean Fever  Commonest form of hereditary systemic amyloidosis  Autosomal Recessive, Gene on Chromosome 16  AA type amyloid  Two manifestations of the disease: Short febrile attacks with pain mimicking pleurisy, peritonitis or synovitis Amyloidosis: Manifest early in life  Death before 40 without Renal Transplant  Treated with Colchicine (Stabilization of inflammatory cells)

28. Localized amyloidosis Endocrine associated: Pituitary - age related Islets of Langerhans - NIDDM related Medullary Carcinoma of Thyroid (Calcitonin) Intacerebral: Alzheimer’s, Spongiform Encephalopathy

29. Islets

30. Amyloid angiopathy 2 nd Year Pathology 2009

31. Alzheimer’s disease

32. Medullary Thyroid Carcinoma

33. Pathological Calcification Dystrophic Calcification Metastatic Calcification Idiopathic Calcification

34. Dystrophic Calcification Normal serum calcium/phosphate Nonviable or dying tissues Occurs in atherosclerosis, damaged valves, necrosis (coagulative, caseous, liquefactive), Leiomyomas Monckeberg’s medial sclerosis Intracellular / Extracellular Two phases: Initiation and Propagation

35. Dystrophic Calcification Tricuspid valve Stomach

36. Metastatic Calcification Must be associated with elevated Calcium. Occurs in vital tissues Aetiology: –Increased PTH »Hyperparathyroidism –Bone destruction »Tumour (MM, Leukaemia), Skeletal Mets (Breast Ca.) » Increased bone turnover (Pagets), Immobilisation. –Vitamin D related disorders »Excess Vitamin D »Sarcoidosis –Renal Failure »Retention of Phosphate, (secondary HyperPTH)

37. Metastatic Calcification Kidney: Around tubules - Renal Failure. Lung: Alveolar walls. Stomach: Fundal glands. Identification of Calcium: Von Kossa 2 nd Year Pathology 2009

39. Hyperparathyroidism Primary, Secondary, Tertiary 1° = Increased bone re-absorbtion and mobilization of calcium from bone Increased renal tubule re-absorption. Increased Vit. D activity “ Bones, stones, (psychic moans) and abdominal groans” 2 nd Year Pathology 2009

40. Aetiology: 1) Parathyroid adenoma. 75% 2) Hyperplasia. 10-15% 3) Parathyroid carcinoma <5% (MEN 1 or MEN2a) Secondary: Renal Failure, Osteomalacia.

44. Ageing Aging is NOT a disease The changes that occur with aging make aged persons more susceptible to disease Aging is probably multifactorial involving both internal (Genetically programmed) and external (Tissue damage) factors that combine to exert their effects

45. l Theories of Aging –Cellular Changes. There is cumulative free radical damage to DNA and to proteins (perhaps due to lack of antioxidants). –Genetics Another theory suggests that aging is determined by genetic programming and malfunction. 1962 The Hayflick phenomenon. Telomeres and Telomerase Progeria (Werner’s syndrome). DNA Helicase –Loss of Homeostasis –Environmental Stress –Neuroendocrine Dysfunction –Nutrition

46. Organ Systems Central Nervous System –Stroke. –Alzheimer’s. –Neuronal loss. Eye –Cataracts. –Presbyopia. Ear –Presbycusis. 2 nd Year Pathology 2009

47. Cardiovascular System –Atheroma –Calcification (Senile calcific aortic sclerosis) –Senile Amyloid. Urinary Tract –Decreased GFR. –Increased UTI in women. Musculoskeletal System –Decreased bone mass. –Osteoarthritis.

48. Genital Tract –Menopause leads to atrophy of ovaries, uterus, and breasts. The epithelium of the vagina and vulva also become thinner. –Prostatic Hyperplasia. Skin –Decreased elasticity. –Senile lentigines 2 nd Year Pathology 2009

49. Telomere

50. Timing of Aging: Telomeres Telomerase stabilizes telomere lengths –Active in germ cells, absent in most somatic cells –May be reactivated in cancer cells When cells replicate, small portion of telomere not duplicated Telomere shortening – growth checkpoint = signal for cell to become senescent Mechanism for cells to count their divisions

51. Genetic Damage and Aging: Werner’s Syndrome DNA helicases are known to be involved in the repair, replication and expression of the genetic material. Defects in DNA helicase lead to premature aging Rapid accumulation of genetic damage, mimicking the aging process Premature aging also seen in Cockayne syndrome and ataxia telangiectasia, other inherited defects in DNA repair.

52. Summary Ageing Effects a combination of –Intrinsic factors: telomere lengths –Extrinsic factors: progressive DNA (and other cellular) damage –The latter may be accelerated by inherited defects in DNA repair.

53. Summary Amyloid. –Appearance / identification –Types of acquired and inherited amyloidosis Pathological calcification. –Dystrophic / Metastatic / Idiopathic –Causes and Consequences Ageing –Theories of Aging