PATHOLOGIC CALCIFICATION .pptx

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1. INTRACELLULAR ACCUMULATIONS Dr Ogbata Stanley Emeka Dip SLT, MBBS, FMCPath CONSULTANT ANATOMIC PATHOLOGIST.

2. INTRACELLULAR ACCUMULATIONS Definition/introduction Mechanisms/Pathogenesis Types Morphology Special stains Clinical correlate Further reading

3. INTRACELLULAR ACCUMULATIONS: INTRODUCTION One of the manifestations of metabolic derangements in cells is the intracellular accumulation of substances that may be harmless or cause further injury. These accumulations may be located in the cytoplasm, within organelles (typically lysosomes ), or in the nucleus, and they may be composed of substances that are synthesized by the affected cells or are produced elsewhere.

4. INTRACELLULAR ACCUMULATIONS : INTRODUCTION ct’d In many cases, if the overload can be controlled or stopped, the accumulation is reversible. In inherited storage diseases, accumulation is progressive and may cause cellular injury, leading in some instances to death of the tissue and the patient.

5. MECHANISMS OF INTRACELLULAR ACCUMULATIONS There are four main mechanisms leading to abnormal intracellular accumulations: Inadequate removal of a normal substance secondary to defects in packaging and transport, as in fatty change ( steatosis ) in the liver. Accumulation of an endogenous substance as a result of genetic or acquired defects in its folding, packaging, transport, or secretion , as with certain mutated forms of α1-antitrypsin.

6. MECHANISMS OF INTRACELLULAR ACCUMULATIONS ct’d Failure to degrade a metabolite due to inherited enzyme deficiencies, typically lysosomal enzymes. The resulting disorders are called lysosomal storage diseases.

7. MECHANISMS OF INTRACELLULAR ACCUMULATIONS ct’d Deposition and accumulation of an abnormal exogenous substance when the cell has neither the enzymatic machinery to degrade the substance nor the ability to transport it to other sites. Accumulation of carbon or silica particles is an example of this type of alteration.

8. MECHANISMS OF INTRACELLULAR ACCUMULATIONS ct’d

9. TYPES OF INTRACELLULAR ACCUMULATIONS Lipids All major classes of lipids can accumulate in cells: triglycerides, cholesterol/cholesterol esters, and phospholipids. Phospholipids are components of the myelin figures found in necrotic cells. In addition, abnormal complexes of lipids and carbohydrates accumulate in the lysosomal storage diseases

10. TYPES OF INTRACELLULAR ACCUMULATIONS ct’d Steatosis (Fatty Change): The terms steatosis and fatty change describe abnormal accumulations of triglycerides within parenchymal cells. Fatty change is often seen in the liver because it is the major organ involved in fat metabolism, but it also occurs in the heart, muscle, and kidney.

11. TYPES OF INTRACELLULAR ACCUMULATIONS ct’d The causes of steatosis include alcohol , toxins , protein malnutrition, diabetes mellitus, obesity, and anoxia. In higher-income nations, the most common causes of significant fatty change in the liver (fatty liver) are alcohol abuse and nonalcoholic fatty liver disease, which is often associated with diabetes and obesity.

12. MORPHOLOGY GROSS The liver is enlarged and yellow with tense, glistening capsule and rounded margins. The cut surface bulges slightly and is pale-yellow and greasy to touch. MICROSCOPY Fat in the cytoplasm of the hepatocytes is seen as clear area which may vary from minute droplets in the cytoplasm of a few hepatocytes ( microvesicules ) to distention of the entire cytoplasm of most cells by coalesced droplets ( macrovesicules ) pushing the nucleus to periphery of the cell.

13. MORPHOLOGY ct’d Occasionally, the adjacent cells containing fat rupture and produce fatty cysts. Infrequently, lipogranulomas may appear consisting of collection of macrophages, lymphocytes and multinucleate giant cells. Special stains such as Sudan III, Sudan IV, Sudan Black and Oil Red O can be employed to demonstrate fat in the tissue.

14. FATTY LIVER : GROSS MORPHOLOGY Diaphragmatic surface sectioned slice of the liver shows pale yellow parenchyma with rounded borders.

15. Morphology - Normal liver:photomicrograph H & E

16. MORPHOLOGY ct’d schematic Photomicrograph: H & E

17. TYPES OF INTRACELLULAR ACCUMULATIONS ct’d Cholesterol and Cholesterol Esters The cellular metabolism of cholesterol is tightly regulated such that most cells use cholesterol for the synthesis of cell membranes without intracellular accumulation of cholesterol or cholesterol esters. Accumulations manifested histologically by intracellular vacuoles are seen in several pathologic processes.

18. TYPES OF INTRACELLULAR ACCUMULATIONS ct’d Atherosclerosis: In atherosclerotic plaques, smooth muscle cells and macrophages within the intimal layer of the aorta and large arteries are filled with lipid vacuoles, most of which contain cholesterol and cholesterol esters. Such cells have a foamy appearance (foam cells), and aggregates of them in the intima produce the yellow cholesterol-laden atheromas characteristic of this serious disorder.

19. TYPES OF INTRACELLULAR ACCUMULATIONS ct’d Some of these fat-laden cells may rupture, releasing cholesterol and cholesterol esters into the extracellular space, where they may form crystals. Some form long needles that produce distinct clefts in tissue sections, while other small crystals are phagocytosed by macrophages and activate the inflammasome , contributing to local inflammation.

20. Artery - Morphology: photomicrograph An arterial lumen is occluded by atheroembolic material that contains cholesterol clefts.

21. TYPES OF INTRACELLULAR ACCUMULATIONS ct’d Xanthomas : Intracellular accumulation of cholesterol within macrophages is also characteristic of acquired and hereditary hyperlipidemic states. Clusters of foamy cells are found in the subepithelial connective tissue of the skin and in tendons, producing tumorous masses known as xanthomas . Cholesterolosis : This refers to the focal accumulations of cholesterol-laden macrophages in the lamina propria of the gallbladder.

22. MORPHOLOGY - gross Xanthelasma Tuberous xanthoma

23. MORPHOLOGY Plane xanthoma with yellow palmar creases. Cholesterolosis : photomicrograph – H & E

24. TYPES OF INTRACELLULAR ACCUMULATIONS ct’d Niemann -Pick disease, type C: This lysosomal storage disease is caused by mutations affecting an enzyme involved in cholesterol trafficking, resulting in cholesterol accumulation in multiple organs

25. TYPES OF INTRACELLULAR ACCUMULATIONS ct’d Proteins Intracellular accumulations of proteins usually appear as rounded, eosinophilic droplets, vacuoles, or aggregates in the cytoplasm. By electron microscopy, they can be amorphous, fibrillar , or crystalline in appearance.

26. TYPES OF INTRACELLULAR ACCUMULATIONS ct’d Excesses of proteins within the cells sufficient to cause morphologically visible accumulation have diverse causes: Reabsorption droplets in proximal renal tubules are seen in renal diseases associated with protein loss in the urine ( proteinuria ). In the kidney, small amounts of protein filtered through the glomerulus are normally reabsorbed by pinocytosis in the proximal tubule.

27. TYPES OF INTRACELLULAR ACCUMULATIONS ct’d In disorders with heavy protein leakage across the glomerular filter, there is increased reabsorption of the protein into vesicles, and the protein appears as pink hyaline droplets within the cytoplasm of the tubular cell. The process is reversible; if the proteinuria diminishes, the protein droplets are metabolized and disappear.

28. Photomicrograph – H & E: Protein reabsorption droplets in the renal tubular epithelium.

29. TYPES OF INTRACELLULAR ACCUMULATIONS ct’d The proteins that accumulate may be normal secreted proteins that are produced in excessive amounts and accumulate within the ER, as occurs in certain plasma cells engaged in active synthesis of immunoglobulins . The ER becomes hugely distended, producing large, homogeneous eosinophilic inclusions called Russell bodies.

30. TYPES OF INTRACELLULAR ACCUMULATIONS ct’d Defective intracellular transport and secretion of critical proteins . In α1-antitrypsin deficiency, mutations in the protein significantly slow folding, resulting in the buildup of partially folded intermediates, which aggregate in the ER of the hepatocyte and are not secreted.

31. TYPES OF INTRACELLULAR ACCUMULATIONS ct’d The resultant deficiency of the circulating enzyme where it is needed in the lung causes emphysema. In many of these protein-folding diseases, the pathology results not only from loss of protein function but also ER stress caused by the misfolded proteins, which initiates the unfolded protein response and culminates in cell death by apoptosis.

32. TYPES OF INTRACELLULAR ACCUMULATIONS ct’d Accumulation of cytoskeletal proteins. There are several types of cytoskeletal proteins, including microtubules (20 to 25 nm in diameter), thin actin filaments (6 to 8 nm), thick myosin filaments (15 nm), and intermediate filaments (10 nm).

33. TYPES OF INTRACELLULAR ACCUMULATIONS ct’d Intermediate filaments, which provide a flexible intracellular scaffold that organizes the cytoplasm and resists forces applied to the cell, are divided into five classes: keratin filaments (characteristic of epithelial cells), neurofilaments ( neurons ), desmin filaments (muscle cells ), vimentin filaments (connective tissue cells), and Glial filaments ( astrocytes ). Accumulations of keratin filaments and neurofilaments are associated with certain types of cell injury.

34. TYPES OF INTRACELLULAR ACCUMULATIONS ct’d Alcoholic hyaline is an eosinophilic cytoplasmic inclusion in liver cells that is characteristic of alcoholic liver disease and is composed predominantly of keratin intermediate filaments. The neurofibrillary tangle found in the brain in Alzheimer disease contains neurofilaments and other proteins.

35. TYPES OF INTRACELLULAR ACCUMULATIONS ct’d Aggregation of abnormal proteins. Abnormal or misfolded proteins may deposit in tissues and interfere with normal functions. The deposits can be intracellular, extracellular, or both, and the aggregates may either directly or indirectly cause the pathologic changes. Certain forms of amyloidosis fall in this category of diseases. These disorders are sometimes called proteinopathies or protein-aggregation diseases.

36. TYPES OF INTRACELLULAR ACCUMULATIONS ct’d Hyaline Change The term hyaline usually refers to an alteration within cells or in the extracellular space that gives a homogeneous, glassy, pink appearance in routine histologic sections stained with H&E. It is widely used as a descriptive histologic term rather than a specific marker for cell injury. This morphologic change is produced by a variety of alterations and does not represent a specific pattern of accumulation.

37. TYPES OF INTRACELLULAR ACCUMULATIONS ct’d Intracellular hyaline accumulations of protein include: reabsorption droplets, Russell bodies, and alcoholic hyaline. Extracellular hyaline has been more difficult to analyze.

38. TYPES OF INTRACELLULAR ACCUMULATIONS ct’d Collagenous fibers in old scars may appear hyalinized , but the biochemical basis of this change is not clear. In long-standing hypertension and diabetes mellitus, the walls of arterioles, especially in the kidney, become hyalinized , resulting from extravasated plasma protein and deposition of basement membrane material.

39. Examples of hyaline change

40. Examples of hyaline change Mallory hyaline/body observed in: Alcoholic hepatitis 2. Indian childhood cirrhosis (ICC) 3. Primary biliary cirrhosis 4. Wilson disease 5. Hepatocellular carcinoma 6. Focal nodular hyperplasia. Russell body: Excessive accumulation of immunoglobulins in the plasma cells. Crooke’s hyaline body: Present in basophil cells of pituitary gland in Cushing syndrome.

41. MORPHOLOGY - MICROSCOPY Extracellular hyaline Intracellular hyaline

42. TYPES OF INTRACELLULAR ACCUMULATIONS ct’d Glycogen Excessive intracellular deposits of glycogen are seen in patients with an abnormality in either glucose or glycogen metabolism. Glycogen is a readily available source of glucose stored in the cytoplasm of healthy cells.

43. TYPES OF INTRACELLULAR ACCUMULATIONS ct’d Whatever the clinical setting, the glycogen masses appear as clear vacuoles within the cytoplasm because glycogen dissolves in aqueous fixatives; thus, it is most readily identified when tissues are fixed in absolute alcohol. Staining with Best carmine or the PAS reaction imparts a rose-to-violet color to the glycogen, but can also stain protein-bound carbohydrates. Diastase digestion of a parallel section that demonstrates loss of staining due to glycogen hydrolysis is therefore an important validation.

44. TYPES OF INTRACELLULAR ACCUMULATIONS ct’d Diabetes mellitus is the prime example of a disorder of glucose metabolism. In this disease, glycogen is found in renal tubular epithelial cells, as well as within liver cells, β cells of the islets of Langerhans within the pancreas, and heart muscle cells. Glycogen accumulates within select cells in a group of related genetic disorders that are collectively referred to as the glycogen storage diseases, or glycogenoses . In these diseases, enzymatic defects in the synthesis or breakdown of glycogen result in massive accumulation, causing cell injury and cell death.

45. TYPES OF INTRACELLULAR ACCUMULATIONS ct’d Pigments Pigments are colored substances, some of which are normal constituents of cells (e.g., melanin), whereas others are abnormal and accumulate in cells under special circumstances. Pigments can be exogenous, coming from outside the body, or endogenous, synthesized within the body itself.

46. Different types of pigments

47. TYPES OF INTRACELLULAR ACCUMULATIONS ct’d Exogenous Pigments The most common exogenous pigment is carbon (coal dust), a ubiquitous air pollutant in urban areas. When inhaled, it is picked up by macrophages within the alveoli and then transported through lymphatic channels to lymph nodes in the tracheobronchial region. Accumulations of this pigment blacken the tissues of the lungs ( anthracosis ) and the involved lymph nodes.

48. TYPES OF INTRACELLULAR ACCUMULATIONS ct’d In coal miners, the aggregates of carbon dust may induce a fibroblastic reaction or even emphysema, and thus cause a serious lung disease known as coal worker’s pneumoconiosis. Tattooing is a form of localized, exogenous pigmentation of the skin. The pigments inoculated are phagocytosed by dermal macrophages, in which they reside for the remainder of the life of the embellished. The pigments do not usually evoke any inflammatory response.

49. TYPES OF INTRACELLULAR ACCUMULATIONS ct’d Endogenous Pigments Lipofuscin is an insoluble pigment, also known as lipochrome or wear-and-tear pigment. Lipofuscin is composed of polymers of lipids and phospholipids in complex with protein, suggesting that it is derived through lipid peroxidation of polyunsaturated lipids of intracellular membranes.

50. TYPES OF INTRACELLULAR ACCUMULATIONS ct’d Lipofuscin is not injurious to the cell or its functions. Its importance lies in its being a telltale sign of free radical injury and lipid peroxidation . The term is derived from the Latin ( fuscus , brown), referring to brown lipid.

51. TYPES OF INTRACELLULAR ACCUMULATIONS ct’d In tissue sections, it appears as a yellow-brown, finely granular cytoplasmic , often perinuclear , pigment. It is seen in cells undergoing slow, regressive changes and is particularly prominent in the liver and heart of aging patients or patients with severe malnutrition and cancer cachexia .

52. Lipofuscin granules in cardiac myocytes

53. TYPES OF INTRACELLULAR ACCUMULATIONS ct’d Melanin Derived from the Greek ( melas , black), is an endogenous, brown-black, pigment formed when the enzyme tyrosinase catalyzes the oxidation of tyrosine to dihydroxyphenylalanine in melanocytes . For practical purposes, melanin is the only endogenous brown-black pigment. The only other that could be considered in this category is homogentisic acid, a black pigment that occurs in patients with alkaptonuria , a rare metabolic disease.

54. TYPES OF INTRACELLULAR ACCUMULATIONS ct’d Here the pigment is deposited in the skin, connective tissue, and cartilage, and the pigmentation is known as ochronosis . Hemosiderin A hemoglobin-derived, golden yellow to- brown, granular, or crystalline pigment is one of the major storage forms of iron.

55. morphology –melanin pigmentation: gross

56. MORPHOLOGY - MICROSCOPY SCHEMATIC PHOTOMICROGRAPH: H & E

57. TYPES OF INTRACELLULAR ACCUMULATIONS ct’d Iron is normally carried by a specific transport protein called transferrin . In cells, it is stored in association with a protein, apoferritin , to form ferritin micelles. Ferritin is a constituent of most cell types. When there is a local or systemic excess of iron, ferritin forms hemosiderin granules, which are easily seen with the light microscope.

58. TYPES OF INTRACELLULAR ACCUMULATIONS ct’d Hemosiderin pigment represents aggregates of ferritin micelles. Under normal conditions, small amounts of hemosiderin can be seen in the mononuclear phagocytes of the bone marrow, spleen, and liver, which are responsible for recycling of iron derived from hemoglobin during the breakdown of effete red blood cells.

59. Hereditary hemochromatosis - photomicrograph: prussian blue stain Prussian blue staining marks the intraparenchymal deposition of hemosiderin .

60. TYPES OF INTRACELLULAR ACCUMULATIONS ct’d Local or systemic excesses of iron cause hemosiderin to accumulate within cells. Local excesses result from hemorrhages in tissues. The best example of localized hemosiderosis is the common bruise.

61. TYPES OF INTRACELLULAR ACCUMULATIONS ct’d Extravasated red blood cells at the site of injury are phagocytosed over several days by macrophages, which break down the hemoglobin and recover the iron. After removal of iron, the heme moiety is converted first to biliverdin (“green bile”) and then to bilirubin (“red bile”).

62. TYPES OF INTRACELLULAR ACCUMULATIONS ct’d In parallel, the iron released from heme is incorporated into ferritin and eventually hemosiderin . These conversions account for the often dramatic play of colors seen in a healing bruise, which typically changes from red-blue to green-blue to golden-yellow before it is resolved.

63. TYPES OF INTRACELLULAR ACCUMULATIONS ct’d When there is systemic iron overload, hemosiderin may be deposited in many organs and tissues, a condition called hemosiderosis . The main causes of hemosiderosis are:

64. TYPES OF INTRACELLULAR ACCUMULATIONS ct’d increased absorption of dietary iron due to an inborn error of metabolism called hemochromatosis , hemolytic anemias , in which excessive lysis of red blood cells leads to release of abnormal quantities of iron, and repeated blood transfusions, because transfused red blood cells constitute an exogenous iron load.

65. Commonly used special stains in histopathology

66. Commonly used special stains in histopathology

67. Exercise 1 A 45-year-old man is referred because of a recent diagnosis of hereditary hemochromatosis . Which of the following is a correct statement about this disorder? (A) Damage to organs results from abnormal deposition of lead. (B) It can progress to liver cirrhosis, diabetes mellitus, and skin pigmentation. (C) Most cases are due to spontaneous mutations. (D) Skin hyperpigmentation is due to bilirubin accumulation. (E) The TIBC is characteristically increased.

68. Exercise 2 The illustration is from a liver biopsy of a 34-year-old woman with a long history of alcoholism. Which of the following is the best explanation for the changes shown here? (A) Accumulation of triglycerides within hepatocytes . (B) Apoptosis with replacement of damaged cells by lipid-laden macrophages. (C) Bilirubin accumulation with mobilization of fat by bile salts. (D) Enzymatic fat necrosis with digestion of liver parenchyma by released enzymes. (E) Irreversible damage to mitochondria.

69. Exercise 3 A 20-year-old man presents with yellowing of the sclerae , skin, and oral mucosa. Which of the following accumulations underlies these findings? (A) Bilirubin (B) Hemosiderin (C) Lead (D) Melanin (E) Silver

70. PATHOLOGIC CALCIFICATION Definition/introduction Types Pathogenesis Morphology Special stains for calcium Clinical correlate Further reading

71. Definition/introduction Pathologic calcification is the abnormal tissue deposition of calcium salts, together with smaller amounts of iron, magnesium , and other mineral salts . The deposition can be intracellular, extracellular, or in both locations .

72. TYPES PATHOLOGIC CALCIFICATION There are 2 types: Dystrophic calcification Metastatic calcification

73. Dystrophic calcification This is the deposition of calcium salts locally in dead/dying tissues. It occurs despite normal serum levels of calcium and in the absence of derangements in calcium metabolism . Hypercalcemia however accentuates dystrophic calcification.

74. Dystrophic calcification ct’d Dystrophic calcification may simply be a telltale sign of previous cell injury. It is often a cause of organ dysfunction

75. Dystrophic calcification cont’d Dystrophic calcification is encountered in: atheromas of advanced atherosclerosis. aging or damaged heart valves. tuberculous lymph node. some types of papillary cancers e.g. papillary carcinoma of the thyroid. Monckeberg medial calcific sclerosis.

76. Metastatic Calcification This is the deposition of calcium salts in otherwise normal tissues. It almost always results from hypercalcemia secondary to some disturbance in calcium metabolism . Metastatic calcification is reversible upon correction of underlying metabolic disorder.

77. Metastatic Calcification ct’d Metastatic calcification may occur widely throughout the body but principally affects the interstitial tissues of the: gastric mucosa kidneys lungs systemic arteries and pulmonary veins .

78. Metastatic Calcification ct’d Metastatic calcification is encountered in: hyperparathyroidism due to parathyroid tumors, and ectopic secretion of PTH-related protein by malignant tumors. resorption of bone tissue secondary to primary tumors of bone marrow as seen in multiple myeloma and leukemia, or diffuse skeletal metastasis as seen in breast cancer, accelerated bone turnover as in Paget disease, or immobilization.

79. Metastatic Calcification cont’d vitamin D–related disorders including vitamin D intoxication, sarcoidosis (in which macrophages activate a vitamin D precursor), and idiopathic hypercalcemia of infancy ( Williams syndrome ). renal failure, which causes retention of phosphate, leading to secondary hyperparathyroidism . Less common causes include aluminum intoxication, which occurs in patients on chronic renal dialysis, and milk-alkali syndrome, which is due to excessive ingestion of calcium and absorbable antacids such as milk or calcium carbonate.

80. PATHOGENESIS PATHOLOGIC CALCIFICATION Dystrophic calcification : It involves 2 phases: Initiation: Following cell injury (i.e. degeneration or necrosis), there is membrane damage and release of membrane phospholipids. Phosphatases associated with phospholipids generate phosphate ions. There is also excess uptake of calcium by injured mitochondria in degeneration and necrosis.

81. PATHOGENESIS PATHOLOGIC CALCIFICATION ct’d Thus, calcium and phosphate so generated from these mechanisms form precipitates of calcium phosphate. Propagation : Simultaneously, some structural changes occur in calcium and phosphate groups which result in further deposition and form mineral crystals.

82. PATHOGENESIS PATHOLOGIC CALCIFICATION ct’d Metastatic calcification: Metastatic calcification occurs due to excessive binding of inorganic phosphate ions with elevated calcium ions due to underlying metabolic derangement. This leads to precipitates of calcium phosphate at the preferential sites, due to presence of acid secretions or rapid changes in pH levels at these sites.

83. Differences between dystrophic and metastatic calcification.

84. Morphology Macroscopy : Fine , white granules or clumps, often felt as gritty deposits. Microscopy: Basophilic , amorphous irregular granules, sometimes clumped appearance on H&E.

85. Morphology ct’d Dystrophic calcification of the aortic valve - gross. Calcification in Monckeberg medial calcific sclerosis – photomicro , H & E.

86. Morphology ct’d : photomicrograph Metastatic pulmonary Calcification appearing as basophilic deposits along the alveolar septa. H & E. Metastatic calcification of the liver. von Kossa’s staining for calcium

87. Special stains for calcium Von- Kossa - silver impregnation method. - Produces black colour . Alizarin red S - produces red colour . Purpurin , naphthochrome green B and nuclear fast red.

88. Clinical correlates Myocardial infarction(MI): Dystrophic calcification in atherosclerotic coronary arteries contributes to narrowing of those vessels leading to MI. Mitral and aortic valves stenosis : leads to impeded blood flow because it produces inflexible valve leaflets and narrowed valve orifices. Cerebral infarct. Lithopaedion .

89. Clinical correlates ct’d Mammography is based principally on the detection of microcalcifications in breast cancers. Congenital toxoplasmosis , an infection involving the central nervous system, is suggested by the visualization of calcification in the infant brain.

90. THANK YOU FOR LISTENING

91. Post lecture MCQ 1. Dystrophic calcification: a. Almost always associated with hypercalcaemia . b. Not associated with atheromas of advanced atherosclerosis. c. Seen in some types of papillary carcinomas. d. Cannot occur in normocalcaemia . e. Associated with renal failure.

92. Post lecture MCQ 2. Metastatic calcification is associated with: a. Necrosis b. Hypocalcaemia c. Hypercalcaemia d. Monckeberg medial calcific sclerosis e. Irreversibility

93. Post lecture MCQ 3 A 60-year-old woman with breast cancer and widespread bony metastases is found to have calcification of multiple organs. The calcifications are best described as dystrophic with decreased serum calcium. (B) dystrophic with increased serumcalcium . (C) metastatic with decreased serum calcium. (D) metastatic with increased serumcalcium .

94. Further reading Kuma V, Abbas AK, Aster JC. ROBBINS& COTRAN PATHOLOGIC BASIS OF DISEASE.10 th ed. Elsevier Inc.; 2021. Pgs 65 – 66. Sunil RL, Susan AD, Caroline JF. Basic Pathology. An introduction to the mechanisms of disease. 4 th ed. London: Hodder Education; 2009. Pgs 17 – 18. Rubin R, Strayer DS. Rubin's Pathology : Clinicopathologic Foundations of Medicine, 5th ed. 2008. Pgs 8 – 9.

95. Further reading Senba M, Kawai K, Mori N. Pathogenesis of Metastatic Calcification and Acute Pancreatitis in Adult T-Cell Leukemia under Hypercalcemic State. Leukemia Research and Treatment.2012.