MLS Review - Hematology - Hematopoiesis & Anemias

MEDICAL LABORATORY SCIENCE REVIEW PEDIATRIC AND ADULT HEMATOPOIESIS HEMATOLOGY: HEMATOPOIESIS & ANEMIAS BONE MARROW OUTLINE • • Hematopoiesis o Intrauterine Hematopoiesis o Pediatric and Adult Hematopoiesis o Erythrocyte Anemias o Introduction o Classification Of Anemia o Impaired Or Defective Production Anemias o Blood Loss Anemias o Hemolytic Anemias o Anemia And The Elderly • • • • • HEMATOPOEISIS • • Process of blood cell production Hematopoietic disorders may be attributed to issues with hematopoiesis Main site of hematopoiesis Newborn: 80 – 90% of bone marrow is active bone marrow Young adult (age 20): 60% of bone marrow is active (hematopoiesis: proximal ends of large flat bones, pelvis, sternum) Older adult (age 55): 40% of bone marrow is active and 60% is fat (yellow marrow) Cellularity: ratio of marrow cells to fat (red : yellow marrow) o Normocellular: marrow has 30-70% hematopoietic cells o Hypercellular/hyperplastic: marrow has >70% hematopoietic cells ▪ More blood components may be produced o Hypocellular/hypoplastic: marrow has <30% hematopoietic cells o Aplastic: marrow has few or no hematopoietic cells M:E (MYELOID:ERYTHROID) RATIO INTRAUTERINE HEMATOPOEISIS • MESOBLASTIC PHASE (YOLK SAC PHASE) • • • Begins at 19th day of gestation Primitive nucleated erythroblasts - first cell Embryonic hemoglobins: Gower I, Gower II, Portland • • Ratio of granulocytes and their precursors to nucleated erythroid precursors Normal: 3:1 or 4:1 Lymphocytes & monocytes are excluded from M:E ratio o Lymphocytes are part of lymphoid system o Monocytes mature into macrophages HEPATIC PHASE HEMATOPOIESIS • • • 1. 2. 3. • Begins at 6 weeks of gestation Occurs in the liver Production of red blood cells, granulocytes, monocytes, and megakaryocytes Alpha and gamma globin chain production (HbF/Fetal Hgb) MYELOID/MEDULLARY PHASE • • • • Begins around 5th month of gestation Bone marrow producing mainly granulocytes M:E ratio 3:1 6th months of age gamma-beta globin chain switch SUMMARY OF INTRAUTERINE HEMATOPOIESIS Aka Start End Site Mesoblastic Yolk-sac phase 19th day after fertilization 2nd to 4th month after fertilization Yolk-sac, intravascularly Primitive erythroblasts Detectable Gower I, Gower hemoglobin II, Portland Product Notes Hepatic Medullary Myeloid phase 4 to 5 weeks after 5th to 6th month fertilization of development Peak: 3rd month throughout lifetime Extravascular: Extravascular: Liver (primary); Bone marrow thymus, spleen, (primary); Liver kidneys (2°) and spleen (2°) RBC, and some Cells of various other blood cells lineages are seen HbA, HbA2, HbF HbA1, HbA2 Pluripotential stem cells Committed progenitor cells (Lymphoid or Myeloid) Mature Blood Cells o Uses PBS to identify blood cell maturity & morphology Committed Growth Factors/ Mature Cell Progenitor Cells Interleukins CFU-MEG Thrombopoietin, Thrombocytes GM-CSF CFU-GM CFU-M GM-CSF, M-CSF, IL-3 Monocytes CFU-GM CFU-G GM-CSF, G-CSF, IL-3 Neutrophils BFU-E CFU-E Erythropoietin, Erythrocytes GM-CSF, IL-3 CFU-Eo GM-CSF, IL-3, IL-5 Eosinophils CFU-Bs IL-3, IL-4 Basophils CFU, Colony forming unit; CSF, Colony stimulating factor; IL, interleukin; MEG, Megakaryocyte ERYTHROCYTE Thymus - T cell Kidney - B cell Spleen - RBC, lymphocytes 1 CLASSIFICATION OF ANEMIA MORPHOLOGIC CLASSIFICATION OF ANEMIA • ANEMIAS INTRODUCTION TO ANEMIAS • • • • Anemias o Defined as a decrease in RBCs and hemoglobin, resulting in decreased oxygen delivery to the tissues o Impoverished condition of the blood caused by reduction in RBC, Hgb, or both; considered to be present if the Hgb conc. of the Hct is below the lower limit of the 95% reference interval for the individual’s age, sex, and geographic location o Caused by: ▪ Decrease in RBC production (insufficient production at site of blood production) ▪ Increase RBC destruction (shortened lifespan) o Can be classified morphologically using RBC indices (MCV, MCH, MCHC) to determine etiology/cause Anemia is suspected when the hemoglobin level is <12 g/dL in men or <11 g/dL in women Total Erythropoiesis - total production of RBC and can be measured by M:E ratio, fecal urobilinogen (stool pigment), and plasma iron o Urobilinogen - byproduct of stercobilin from senescent RBC or RBC destruction Effective Erythropoiesis - production of RBC that reach the circulation of the peripheral blood measured by RBC turnover utilization of iron, reticulocyte count, RBC lifespan o Retic count - normal in small amounts in PBS CLINICAL SIGNS & SYMPTOMS • • General: o Low Hgb conc, & blood volume o Fatigue o Dyspnea on exertion (shortness of breath) o Faintness o Vertigo o Palpitation o Headache Common: o Pallor o Rapid bounding pulse o Low blood pressure (below 90/6) o Slight fever o Some dependent edema o Systolic murmurs • Macrocytic Normochromic Anemia - MCV greater than 96 fL.; MCHC is normal (MCHC more accurate than MCH) o Megaloblastic anemia - erythroblasts in bone marrow show abnormality and maturation is delayed ▪ Causes: • Vitamin B12 deficiency - pernicious anemia • Folic acid deficiency - nutritional megaloblastic anemia • Abnormalities of Vit. B12 or folate metabolism • Inherited disorders of DNA synthesis • Drug-induced disorders of DNA synthesis ▪ Laboratory Findings: • RBCs are macrocytic, MCV greater than 95 fL, and often high as 120-140 fL • Macrocytes are typically oval shape • Reticulocyte count is low in relation to the degree of anemia • Total WBC count and platelets counts may be moderately reduced, especially in severely anemic patients ▪ Peripheral smears: • Show normal RBC in morphology despite the drop of Hgb, Hct, and RBC count • Increased bone marrow activity (increased retic ct) w/o increased RBC or Hgb breakdown o Women after menstruation may have increased retic count o Non-megaloblastic anemia - macrocytosis (increased large RBCs); bone marrow shows normoblastic than megaloblastic cells ▪ Causes: • Accelerated erythropoiesis o Rapid blood loss due to accidents induce accelerated erythropoiesis • Increased membrane surface area • Obscure causes (hypoplastic & aplastic anemias) • Alcohol • Liver disease • Cytotoxic drugs (Metronidazole drug for amoebiasis) Microcytic Hypochromic Anemia - MCV less than 80 fL., MCHC is less than 32% o Causes: ▪ Iron deficiency ▪ Disorder or globin synthesis as in thalassemia ▪ Disorders or porphyrin & heme synthesis as in sideroblastic anemia ▪ Other disorders of iron metabolism o Laboratory findings: ▪ Small RBCs ▪ Low Hgb content o Peripheral smears: ▪ Aniso- and poikilocytosis ▪ RBCs are microcytic since many are smaller than the nucleus of the lymphocyte ▪ RBCs are hypochromic w/ increased central pallor ▪ Elliptocytic and pencil-shaped forms are present 2 • • Normocytic Normochromic Anemia - MCV is 80-96 fL. (normal), MCHC is normal o Causes: ▪ Recent blood loss (accident, wound) ▪ Overexpansion of plasma volume as in pregnancy ▪ Hemolytic diseases ▪ Hypoplastic bone marrow (aplastic anemia0 ▪ Infiltrated bone marrow (leukemia) ▪ Endocrine abnormality ▪ Chronic disorders ▪ Renal disorders ▪ Liver diseases o Normochromic, normocytic anemia with effective erythropoiesis ▪ Increased reticulocyte count ▪ Acute blood loss • Very acutely, with hypovolemia, may have normal blood counts, will become anemic with volume replenishment ▪ Hemolytic anemia • Increased reticulocyte production cannot keep pace with loss of RBCs peripherally ▪ Response to specific therapy in nutritional anemias o Laboratory findings: ▪ Plasma volume and RBC volume - reduced in proportionate amount ▪ Hct is normal ▪ Platelet count is reduced ▪ Plasma fibrinogen level is reduced ▪ Neutrophilic leukocytosis is present ▪ Normocytes and normochromic cells are present Differential Diagnosis of the Microcytic/Hypochromic Anemias using Iron-related Parameters ANEMIA DUE TO IMPAIRED RED CELL PRODUCTION: IMPAIRED OR DEFECTIVE PRODUCTION ANEMIAS IRON DEFICIENCY ANEMIA (IDA) • • • • Most common form of anemia Microcytic/hypochromic anemia Serum iron, ferritin, hemoglobin/hematocrit, RBC indices, reticulocyte count – LOW RBC Distribution Width (RDW) and Total Iron Binding Capacity (TIBC) - HIGH ANEMIA OF CHRONIC DISEASE (ACD) • • • • • • Inability to use available normal RBCs Impaired release of storage iron associated with increased Hepcidin levels Normocytic/normochromic anemia, or slightly microcytic/hypochromic anemia Associated with persistent infections, chronic inflammatory disorders ESR, ferritin – INCREASED Serum iron and TIBC – DECREASED SIDEROBLASTIC ANEMIA • • • • • • Caused by blocks on the protoporphyrin pathway resulting in defective hemoglobin synthesis and iron overload Excess iron accumulates in the mitochondrial region of immature erythrocytes (ringed sideroblasts) Two RBC populations (dimorphic) are seen Microcytic/hypochromic anemia Ferritin and serum iron – INCREASED TIBC – DECREASED THALASSEMIA • Inherited blood disorder that causes your body to have less hemoglobin than normal LEAD POISONING • • Multiple blocks in the protoporphyrin pathway Normocytic/normochromic anemia with characteristic course basophilic stippling PORPHYRIAS • • • • Group of inherited disorders characterized by a block in the protoporphyrin pathway Heme precursors before the block accumulate in the tissues, and large amounts are excreted in urine/feces Photosensitivity, abdominal pain, CNS disorders Hematologic findings are insignificant MEGALOBLASTIC ANEMIAS • • • CLASSIFICATION OF ANEMIA ACCORDING TO CAUSE • • • • • Anemia due to impaired RBC production Anemia due to blood loss - post-hemorrhagic anemia Anemia due to accelerated RBC destruction - hemolytic anemia Defective DNA synthesis causes abnormal nuclear maturation RNA synthesis is normal, so the cytoplasm is not affected Nucleus matures slower than the cytoplasm (asynchronism) Caused by either Vitamin B12 deficiency or folic acid deficiency o Increase of Folic acid in diet is an intervention Macrocytic/normochromic anemia 3 APLASTIC ANEMIA INTRINSIC DEFECTS • • • • • Hereditary spherocytosis/Congenital Hemolytic Jaundice Anemia o Inherited as a non-sex-linked dominant trait o Most common in North Europeans o Spherocytes die prematurely o Splenomegaly o Laboratory Findings of Hereditary Spherocytosis: ▪ Osmotic Fragility Test is increased. ▪ Reticulocytes are usually 5-20% ▪ Direct Coomb’s (antiglobulin test is negative). ▪ MCV is normal; MCHC is often increased. ▪ WBC, platelet counts are normal except during periods of hemolysis. ▪ Total bilirubin is elevated. o Peripheral Smears: ▪ Blood film shows microspherocytes. • Paroxysmal Nocturnal Hemoglobinuria (PNH)/ Machiafava-Micheli Syndrome o Chronic intravascular hemolysis o Nocturnal hemoglobinuria occurs during sleep or after awakening o pH of plasma is low due to red cells o Acidosis ▪ Depressed respiration during sleep ▪ Retention of CO2 resulting to acidosis Absence of blood components Bone marrow failure Decrease in hemoglobin/hematocrit and reticulocytes Normocytic/Normochromic anemia MYELOPHTHISIC (BONE MARROW REPLACEMENT) ANEMIA • Hypoproliferative anemia caused by replacement of bone marrow hematopoietic cells by malignant cells or fibrotic tissue Normocytic/normochromic anemia • ANEMIA DUE TO BLOOD LOSS POST-HEMORRHAGIC ANEMIA: BLOOD LOSS ANEMIAS ACUTE & CHRONIC BLOOD LOSS • • Acute Blood Loss Sudden loss of blood • resulting from trauma or • other severe forms of • injury Normocytic/ normochromic anemia Chronic Blood Loss Gradual, small loss of blood Normocytic/normochromic anemia (initially) Microcytic/hypochromic (caused by gradual loss of iron) BLOOD LOSS ANEMIAS • • Acute Posthemorrhagic Anemia o If blood is lost over a short period of time in amount sufficient to cause anemia o Laboratory Findings ▪ Decrease of Hgb, Hct, and RBC count ▪ Increase of bone marrow activity, without increase RBC or Hgb breakdown o Peripheral Blood Smear ▪ Shows normal red cells ▪ Elevated retic count Chronic Posthemorrhagic Anemia o If blood is lost in small amount over an extended period of time, both the clinical and hematologic features that characterize acute post hemorrhagic anemia are lacking o Laboratory Findings ▪ WBC count is normal or slightly decreased ▪ Platelet count is increased o Peripheral Blood Smear ▪ Retic count may be normal or slightly increased ▪ RBCs at first normochromic and normocytic and gradually the newly formed RBCs become microcytic and hypochromic ANEMIA DUE TO ACCELERATED RBC DESTRUCTION HEMOLYTIC ANEMIAS • Hemolytic anemia may be due to: o Intrinsic hemolytic anemias - defect of red cell itself, usually hereditary & grouped as membrane, metabolic or hemoglobin defects. o Extrinsic hemolytic anemias - a factor outside the red cell & acting upon it. Almost always acquired o o Laboratory Findings of PNH ▪ Positive sucrose hemolysis test or Ham’s acidified serum test or sugar water test. ▪ WBC count and platelet count are often low. ▪ Hemosiderinuria is a feature. Peripheral Smears: ▪ Reticulocyte count is elevated. ▪ Normocytic –normochromic anemia is present. 4 • • • Hereditary Elliptocytosis/Ovalocytosis o Inherited as dominant trait o Associated w/ severe hemolytic anemia in infants o Defect involves the impaired association of spectrin dimers resulting in free, unconnected dimers o Laboratory Findings: ▪ Osmotic fragility test is increased. ▪ Autohemolysis of red cells is present. o Peripheral Smears: ▪ Non-hypochromic elliptoytes are abundant on blood films. ▪ Reticulocytes and NRBC are normal in shape. • Hereditary Acanthocytosis (Abetalipoproteinemia) o Caused by absence of beta-lipoprotein o Associated w/ plasma lipid abnormalities o Low total lipid, cholesterol & phospholipids o Autohemolysis occurs o Reticulocyte count ranges from normal to increased o Presence of mild anemia • Rhnull Disease o Inherited due to gene suppression or presence of silent Rh gene (Xo) o Membrane abnormalities due to absence of all Rh-Hr antigens on the red cells. o Laboratory findings: ▪ High Reticulocyte count ▪ Autohemolysis & Osmotic Fragility are increased. o Peripheral Smears: ▪ Mild, chronic normocytic normochromic hemolytic anemia ▪ Smear shows stomatocytes & spherocytes • High Phosphatidyl Hemolytic Anemia o Inherited o Represents an imbalance in the membrane phospholipids in the red cells o Anemia may increase due to infection or under condition of stress. o Peripheral Smears: ▪ Mild anemia w/ morphologically normal cells Hereditary Pyropoikilocytosis o Rare, moderately severe congenital hemolytic anemia o Inherited as recessive autosomal traits o Occurs in blacks o Peripheral Smears: ▪ Microcytosis, striking micropoikilocytosis & fragmentation Hereditary stomatocytosis (Hydrocytosis) o Rare congenital anemia o Inherited as recessive autosomal trait caused by inc. Na & dec. K due to increased permeability of membrane o 0-30% red cells appear as mouth like o Laboratory findings of Hereditary Stomatocytosis: ▪ Osmotic Fragility Test is increased. ▪ Reticulocyte may be normal or elevated. o Peripheral Smears: ▪ 10-30% red cells appear as mouth like linear pallor instead of the normal central round pale area. ▪ 10-50% of red cells appear as a stomatocytes 5 HEMOLYTIC ANEMIA DUE TO METABOLIC DISORDERS • • Glucose-6-Phosphate Dehydrogenase (G6PD) deficiency o Inherited sex-linked o Complex heterogeneous disorder w/c is ubiquitous o Seen in enzyme deficient hemolytic anemia o Type A, Type B & Favism o Laboratory Tests: ▪ Methyl violet or crystal violet stains ▪ Dye Reduction test ▪ Ascorbate Cyanide test ▪ Fluorescent Spot test ▪ Quantitative Assay of G6PD Pyruvate kinase deficiency o The most common red cell enzyme deficiency involving the Embden-Meyerhof glycolytic pathway o Inherited disorder o PK converted to phosphophenolpyruvate to pyruvate in the EMP w/ the production of ATP o RBC loses its flexibility due to decrease ATP o Mild to moderately hemolytic anemia w/ splenomegaly o Peripheral smears: ▪ No notable red cell abnormalities until after splenomegaly ▪ Ecchinocytes ▪ Irregularly contracted red cells ▪ Crenated red cells may be prominent o Laboratory findings and tests: ▪ Reticulocyte count is elevated ▪ Fluorescent test – positive ▪ Quantitative assay of PK • Glucose Phosphate Isomerase Deficiency o This causes an abnormality in anaerobic glycolysis causes a moderately severe anemia o Laboratory findings: ▪ Reticulocyte count may be significantly increased o Peripheral smear: ▪ Red cells show anisocytosis and poikilocytosis • Triosephosphate Isomerase, Hexokinase and Diphosphoglycerate Mutase Deficiency o Other enzyme deficiencies that involve anaerobic glycolysis Glutathione synthase, glutathione peroxidase and glutathione reductase deficiencies o Also show hemolytic anemia o These enzymes are required in HMP like G-6-PD, hemolysis increase due to oxidant drug exposure or infection. • EXTRINSIC HEMOLYTIC ANEMIA • Causes: o Chemical agents – drugs and chemicals o Physical agents – heat, trauma o Vegetable and animal poisons o Infectious agents – malarial parasite, bacteria o Presence of autoantibodies, isoantibodies or drugrelated antibodies causes ANEMIA AND THE ELDERLY • • • Pyrimidine-5-Nucleotidase (PN) Deficiency o Inherited o Caused by an abnormality in nucleotide metabolism o Acquired occurs in lead poisoning & responsible for the basophilic stippling o Laboratory findings: ▪ Reticulocytosis is observed ▪ Positive in the demonstration of decreased Nucleosidase activities o Peripheral Smears: ▪ There are marked basophilic stipplings in red cells • • Hemoglobin less than 13g/dl in males and less than 12g/dl in females Factors contributing: o Decreased bone marrow function o Decline in physical activity o Nutritional deficiencies o Cardiovascular disease o Chronic inflammatory disorders Most common among elderly o Anemia of chronic inflammation o Iron deficiency anemia o Unexplained anemia Ineffective erythropoiesis o Vitamin b12 deficiency o Sideroblastic anemia o Thalassemia 6 • • • • • • • Hypoproliferation o Secondary to iron deficiency o Vitamin b12 or folate deficiency o Renal failure o Hypothyroidism o Chronic inflammation o Endocrine disease Lab evaluation: CBC, retic count, peripheral blood film review, chemistry panel, iron studies, vit b12 and folate levels, free erythrocyte porphyrin o Hypoproliferative anemia o Most common form of anemia in the hospitalized geriatric population ▪ Impaired erythropoietin-dependent erythrocytosis – involved in the pathogenesis of this disease Iron Deficiency Anemia o Affects both erythrocytes and metabolic pathways of iron-dependent tissue enzymes o Results from conditions leading to chronic gastrointestinal blood loss, including long term use of nonsteroidal inflammatory medications, gastritis, peptic ulcer disease, gastropharyngeal reflux disease and angiodysplasia Ineffective Erythropoiesis o Sideroblastic anemia – impaired heme synthesis and abnormal globin synthesis o Megaloblastic anemia – defective DNA synthesis; results in ineffective erythropoiesis (deficiencyofvitaminB12 and folate) Vitamin B12 Deficiency o Megaloblasticanemiain5-10%ofelderly o Attributed to inadequate intestinal absorption of found bound vitaminB12 and pernicious anemia o Loss of gastric acid – bacterial growth: Helicobacter pylori Folate Deficiency o Develops from inadequate dietary intake o Alcoholic elderly are more prone because alcohol interferes with folate absorption Hemolytic anemia o Shortened RBC survival time o Drug induced haemolytic anemia – high doses of antibiotics, non-steroidal inflammatory drugs, quinidine, phenacetin, etc. o Also results from collagen vascular disease, infections, chronic lymphocytic leukemia o Major types: ▪ Caused by immunologic mechanisms ▪ Due to intrinsic effects ▪ Resulting from extrinsic factors 7