Anemia

A Systematic Approach to Anemia Syllabus by Allan Platt, PA-C, MMSc [email protected] www.EmoryPA.org NORMAL BLOOD Blood is the life sustaining substance of mankind. Without the circulation of blood in the body, organs and tissues quickly cease to function and die. Blood is 54% water based plasma, 1% white blood cells and platelets, and 45% red cells. The blood transports life sustaining elements such as oxygen, glucose, proteins, vitamins, enzymes, and electrolytes to every living cell in the body through arteries, veins, and capillaries. Red Blood Cells Red blood cells, or erythrocytes, are the predominant cellular component of blood. Red cells usually make up 45% of the blood volume and are primarily responsible for carrying oxygen from the lungs to the peripheral tissues and organs returning to the lungs with carbon dioxide as a waste product to be exhaled. A 170-pound person has about 30 trillion red cells about 300 million in each drop of blood. The red cell is perfectly designed as a pliable biconcave disc (7 - 8 micron size) to traverse the small capillary beds (3 - 4 micron size) of the micro vasculature. The red cell contains the oxygen carrying protein hemoglobin and enzymes for energy production all encased by a semipermeable and pliable lipid membrane. Hemoglobin Hemoglobin is the predominant protein in the red cell with a wonderful design for transporting oxygen molecules from the lungs for peripheral cellular metabolism and carbon dioxide as a waste product for exhalation. The normal red blood cell is composed of three types of hemoglobin, type A (97%), F (1%) or fetal, and A (2%). Fetal hemoglobin, predominant in utero, tightly binds oxygen 2

facilitating transfer from mother to fetus across the placenta. A manufacturing switch occurs at birth to decrease F and produce A. The most abundant hemoglobin present after one year of life is type A. This hemoglobin is composed of two beta globin chains and two alpha globin chains bonded to four iron containing heme groups. Hemoglobin production requires iron , the synthesis of the protoporphyrin ring and production of the globin chains. Reductions in any of these result in anemias. The globin chains are protein made up of a precise sequence of amino acids that are coded by genes located on chromosome 16 and chromosome 11 contained in the nucleus of the bone marrow stem cells. Any alteration of the amino acid sequence or "blue print" on the chromosomes that code the production of the beta globin chain results in one of the over 600 different types of hemoglobins identified through out the world. Common variants result from substituting valine for glutamic acid in the 6th position of the beta chain producing sickle hemoglobin or Hb S and substituting lysine for valine results in hemoglobin C or Hb C. Other abnormal hemoglobins are formed by other amino acid substitutions. An inherited deletion or dysfunction of one or more of the alpha or beta gene decreases alpha or beta chain production causing thalassemia. The degree of decreased production is related to the inability to manufacture functioning RNA. If alpha chains are reduced alpha thalassemia results and compensatory increase in hemoglobin H (beta4) and Barts (gamma4). If beta chains are reduced, Beta thalassemia results with compensatory increase in hemoglobin A2 and F. Cell Production In the adult, red cells, white cells and platelets are all manufactured in the axial skeleton bone marrow factory where stem cells divide and differentiate into the various cell types under regulation by growth factors. The proper building materials must be supplied to keep the factory producing such as amino acids, carbohydrates, lipids, vitamin B12, folate, vitamin C, and iron. Most of these elements are obtained from absorption in the digestive tract of one partaking of a balanced diet, from recycling red cell breakdown products, and from body stores. Iron is transported to the marrow by transferrin, then once in the maturing red cell, it is incorporated into the hemoglobin molecule. As the red cell matures and forms the hemoglobin necessary for it's life span, the nucleus is extruded and a reticulocyte is expelled into the blood circulation. This juvenile red cell still retains some residual RNA for 2 days that can be identified by staining with methyline blue. The normal reticulocyte count is between 0.8 and 2.4% of the red cell number. Increase or decrease reflects red cell production by the bone marrow factory. If intense marrow

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intense marrow stimulation occurs, larger stress reticulocytes or shift cells may be observed on the peripheral smear. These cells mature after 2 to 3 days in the circulation and may artificially inflate the reticulocyte count. The normal red cell has a life span of 120 days. It is metabolized in the reticuloendothelial system ( spleen , liver, and bone marrow) and the iron, amino acids, carbohydrate and lipids are reclaimed. (Haptoglobin binds free serum hemoglobin released in the blood from red cell lysis for transport to the reticuloendothelial system.) A breakdown product of hemoglobin is bilirubin which is conjugated in the liver and excreted in the biliary system into the gut. Isolated increase in indirect, or pre-liver, bilirubin is a very specific indicator of increased red cell breakdown. The kidney cells secrete erythropoeitin in response to low tissue oxygen levels. Erythropoeitin stimulates bone marrow red cell production. This hormonal feedback loop is less responsive to anemia in renal disease and can cause elevated hemoglobin levels with chronic hypoxia. The Workup of Anemia and Bleeding Anemia is clinically suspected by the presenting symptoms and signs of weakness, fatigue, palpitations, increased heart rate, dyspnea, positional dizziness, syncope, bleeding from any site, increased or new onset angina. It may be suspected when the physical findings of tachycardia, orthostasis, pallor, or jaundice are observed. Anemia may be found on the routine diagnostic screening CBC. The diagnostic approach should contain a thorough history, physical examination and laboratory evaluation . HISTORY - History of melena, abdominal pain, Aspirin or non-steroidal anti-inflammatory agents (NSAIDs) use, anticoagulant use, past peptic ulcer disease, then consider GI bleeding or platelet dyfunction - In females the menstrual history quantifying the amount of blood loss, or possible pregnancy should be obtained. - History of pica or abnormal craving for ice, clay, starch dysphagia then consider iron deficiency. - Poor diet, then consider iron or folate deficiency, and general malnutrition - History of gastric surgery, distal paresthesias, gait problems -consider B12 deficiency - History of alcohol abuse - consider folate deficiency or liver disease - If moonshine use or lead paint/pipe exposure, consider lead toxicity. - Family history of blood cell disorder: consider Sickle Cell disease, G6PD,and Thalassemia - History of jaundice, transfusion, new medication, infection - consider hemolytic process - History of weight loss, Cancer, HIV, rheumatoid arthritis, thyroid disease, renal disease -History of fever and chills, cough, dyspnea, then consider Infection. - History of prolonged bleeding, epistaxis, gum bleeding, easy bruising - consider low platelets or dysfunction. PHYSICAL EXAM GENERAL INSPECTION- Pallor in conjunctiva, nails, or palmar creases Nails - Spoon nails- consider Iron deficiency; Petechiae or purpura- Low platelets, clubbing in TB or lung cancer Skin- Hypothyroid, SLE, Bruises, lesions, Petechiae or purpura- Low platelets Weight - Loss in Cancer, HIV, Chronic disease, gain in hypothyroid VITAL SIGNS- Pulse: Tachycardia from increased cardiac output Respirations: Tachypnea from decreased oxygen transport BP: Orthostatic if volume depleted Temp: Fever in infections and drug or transfusion reactions, hypo in hypothyroid HEENT- Eye: Jaundice if hemolysis, pallor in palpebral conjunctiva Mouth: Glossitis and angular stomatitis in iron or B12 deficiency NECK- Thyroid enlargement or nodules, lymph nodes HEART- Increased output/murmur- consider high output failure LUNG- consider infection, lesion ABDOMINAL- Liver/spleen size, masses, tenderness, surgical scars RECTAL- Stool guaiac, prostate exam in men PELVIC/BREAST- Uterine abnormality, Pap smear, Breast nodule LYMPHNODES- consider lymphoma, leukemia, Infection, connective tissue disease NEUROLOGIC- Decreased vibratory and position sense in B12 deficiency

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LABORATORY- INITIAL SCREENING TESTS: URINANALYSIS- Hematuria/proteinuria in renal disease hemoglobinuria in hemolysis. CBC, red cell morphology and white blood cell differential, Reticulocyte count, Platelet count, Chemistry profile (LDH, Bilirubin- Direct and Indirect, BUN, Creatinine, GPT) Anemia Anemia is from the Greek word anaimia, meaning without blood. Anemia is defined as a reduction in the number of red blood cells, blood hemoglobin content, or hematocrit. There are three primary causes of anemia: (1) reduced production of red blood cells in the bone marrow factory, (2) excessive destruction of red blood cells, and (3) excessive blood loss. THE COMPLETE BLOOD COUNT AND OTHER RELATED TESTS PARAMETER HB - Hemoglobin HCT - Hematocrit RBC - Red Blood Cell Count WBC - White Blood Cell Count Platelet Count

NORMAL ADULT Male= 15.5 +/- 2 mg/dl Female = 13.5 +/- 2 Male= 46.0 +/- 6% Female= 41.0 +/- 6% Male = 4.3 - 5.9 Million/uL Female= 4.0 - 5.2 4.5 - 11.0 K/uL 150 - 400 K cell/uL

Retic - Reticulocyte Count

0.5 -1.5 % 25 - 85 K cell/ul

COMMENTS Low = Anemia High = Polycythemia " " " " Low = Leukopenia High = Leukocytosis Low = Thrombocytopenia High = Thrombocytosis Low in anemia = low High = RBC loss

RED CELL INDICIES MCV - Mean Corpuscular 80 - 90 fl Low = Microcytosis Volume High = Macrocytosis MCH - Mean Corpuscular 27 -32 pg Low = Hypochromic Hemoglobin High = Hyperchromic MCHC - Mean Corpuscular 30 - 36 gm/dl Low = R/O Fe Deficiency Hemoglobin Concentration High = R/O Sperocytosis RDW - Red Cell Distribution 11.5 - 14.5 Variation in RBC size Width Red Cell Morphology - Have human eyes look at the blood smear Burr Cells Uremia, Low K, artifact, Ca stomach, PUD Spur Cell Post-splenectomy, Stomatocyte Hereditary, Alcoholic liver disease, Spherocyte Hereditary, Immune hemolytic anemia, water dilution, post-transfusion Schistocyte - helmet TTP, DIC, vasculitis, glomerulonephritis, heart valve, burns Elliptocyte - Ovalocyte Hereditary, Thalassemia, Iron deficiency, Myelophthistic, megaloblastic anemias Tear Drop Iron deficiency, Myelophthistic, megaloblastic Sickle Cells Sickle cell disease Target Cells Thalassemias, hemoglobinopathies Parasites Malaria, babesiosis Basophilic Stippling Lead toxicity Bite Cells G6PD Deficiency Anisocytosis Red cells are of unequal size.(high RDW) iron def. Poikilo cytosis Red cells are of different shapes

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Reticulocyte Count The reticulocyte count must be corrected for the reduction in red cell count to accurately reflect marrow production of erythrocytes. The most accurate correction is to determine the absolute reticulocyte count, obtained by multiplying the red cell count by the recticulocyte percentage. The normal mean is 50,000 +/- 25,000/ml. Alternatively the raw reticulocyte percentage can be multiplied times the observed hematocrit or hemoglobin divided by the normal hematocrit or hemoglobin as shown: Corrected Retic Raw Retic Pt's Hematocrit =

Percentage Percentage X 45 (normal) The normal corrected reticulocyte percentage is 1 - 2%. Hypoproliferative Anemias: A corrected reticulocyte percentage greater than 2.0 indicates that the anemia is caused by red cell loss either from bleeding or hemolysis. A corrected reticulocyte percentage less than 2.0 indicates that the anemia is from decreased production of erythrocytes . Once this value is known to be under 2, the mean red cell volume or MCV is the next critical value needed to guide the diagnostic work-up. An MCV greater than 94 leads to a macrocytic work-up, an MCV of 80 - 94 leads to a normocytic path, and an MCV under 80 leads to the microcytic differential diagnosis. Figure 2 is a summary of the diagnostic pathway:

Reticulocyte Production Index <2 Decreased Production Red Cell Indicies MCV >94 Macro

80-94 Normo

>2 Increased Loss Hemolysis

<80 Micro

Extrinsic Coombs Positive

Drug

Bleeding

Warm Antibody

Intrinsic Coombs Negative

Membrane

Hb Enzyme

Cold Antibody

If the reticulocyte count is reduced, the bone marrow factory has slowed or stopped production because of a lack of erytropoeitin stimulation, a lack of raw materials such as iron, protein, lipids, folate, vitamin B or the marrow is infiltrated or insulted by a virus, toxin or cancer. In this setting, the mean 12,

corpuscular volume or MCV is the next parameter to guide your evaluation. If the peripheral blood tests do not indicate the diagnosis, referral to a hematologist for a bone marrow aspirate is appropriate. Specific therapy is based on the underlying cause, but in the symptomatic patient, a type specific blood transfusion may be needed to improve oxygen carrying capacity. MICROCYTIC ANEMIAS An MCV of less than 80 with pale, poorly hemoglobinized red cells on peripheral smear characterizes this group of anemias. All result from a disorder in cytoplasmic maturation caused by reduced hemoglobin production. Hemoglobin is produced using iron, protoporphyrin and globin chains. Microcytic anemia can result if iron delivery to the marrow is decreased as in iron deficiency and the anemia of chronic inflammation, heme synthesis is defective as in sideroblastic anemias, or globin chain synthesis is defective as in thalassemias. The differential diagnosis is remembered by the mnemonic

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"TICS": Thalassemias, Iron Deficiency, Chronic diseases and Sideroblastic. Diagnostic evaluation is initiated by excluding iron deficiency. Iron stores are assessed using the serum ferritin, iron, Total Iron Binding Capacity - TIBC (amount of transferrin to transport iron), and percent saturation ( the % of transferrin saturated with iron). The iron and TIBC will also allow diagnosis of the anemia of chronic inflammation. If iron deficiency and the anemia of chronic inflammation are excluded, hemoglobin electrophoresis with quantification of hemoglobin A2 and F will aid the diagnosis of thalassemia. Diagnosis of sideroblastic anemia ultimately requires a bone marrow aspirate stained for iron. The most common sideoblastic anemia is caused by lead toxicity. This treatable cause should always be suspected and worked-up with lead levels. Summary of Findings in Microcytic Anemias Iron Chronic Sideroblastic Deficiency Inflammation Anemia Thalassemia __________ __________ __________ ___________ Red Cells Microcytic Normocytic or Dimorphic Microcytic Hypochromic Microcytic Hypochromic Serum Iron

Decreased

Decreased

Increased

Normal or Increased

TIBC

Increased

Decreased

Normal

Normal

Saturation

< 16%

10-20%

50-100%

30-100%

Serum Ferritin

Decreased

Increased

Increased

Normal or Increased

Bone Marrow Iron Storage

Absent

Increased

Increased

Increased

Sideroblasts

Absent

Absent or Decreased

Increased & Ring Forms

Increased

DIFFERENTIAL DIAGNOSIS: "TICS" T: The thalassemia syndromes result from inherited genetic abnormalities that cause decreased synthesis of a-globin or ß-globin chains, and thus, decreased production of hemoglobin A. In ßthalassemia major, or Cooley’s anemia, patients present with severe anemia, jaundice, and hepatosplenomegaly in the first year of life. In ß-thalassemia minor, splenomegaly may be present, and the peripheral smear often shows target cells and basophilic stippling. Iron study results are normal. Hemoglobin electrophoresis shows increases in hemoglobin types A2 and F.10 Symptoms are usually absent, and no specific therapy is required. Therapeutic iron supplements are not recommended unless a low serum ferritin level confirms concurrent iron deficiency.11 a-Thalassemia has its highest incidence in peoples of Southeast Asian, Mediterranean, and African descent. Diagnosis is based on CBC evidence of more microcytosis than anemia and an elevated red cell count. Hemoglobin electrophoresis is normal. I: Iron deficiency anemia occurs when body iron stores are depleted by prolonged bleeding without adequate replacement.9,11 The daily iron requirement for a man or a nonmenstruating woman is 1.0 mg; for a woman who menstruates, 2.0 mg; and for a pregnant woman, 3.0 to 4.0 mg. The normal daily diet contains 10 mg of iron, of which 5% to 10% is absorbed. Bleeding can lead to a loss of 50 mg of iron per 100 cc of whole blood.

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Laboratory findings may include anemia with a low reticulocyte production index, low MCV, and low mean corpuscular hemoglobin concentration. Physical examination may reveal glossitis, koilonychia (spoon nails), gastritis, and angular stomatitis. The clinician must determine the deficiency’s underlying cause by excluding occult gastrointestinal bleeding, excessive menstrual loss, and inadequate dietary intake. Replacement therapy is Ferrous Sulfate 300mg TID on empty stomach. May give with vitamin C to enhance absorbtion. Follow – up in 3-4 weeks hemoglobin should be normalized and the Ferritn should be normal in 8 weeks. C: Anemia of chronic inflammation. Long-standing infections, neoplastic diseases, and chronic inflammatory processes (eg, rheumatoid arthritis, systemic lupus erythematosus) block iron transportation from the storage sites to the bone marrow factory.12,13 An elevated Westergren sedimentation rate or Creactive protein level can be a nonspecific indicator of inflammation. A normal or elevated ferritin level with a high percent saturation should prompt a close review of the electronic CBC (which may reveal an elevated red cell count, suggesting thalassemia), hemoglobin electrophoresis, the blood smear, and a lead level. S: Sideroblastic anemias are a diverse group characterized by the presence of “ringed” sideroblasts in the bone marrow; basophilic stippling may be present on a peripheral smear. Acquired sideroblastic anemias are associated with use of antituberculous medications (eg, isoniazid, pyrazinamide14), alcohol abuse, lead poisoning, chronic inflammation, and preleukemic states, especially after chemotherapy. Lead levels should be obtained in children and in patients exposed to lead-based paints, car batteries, or “moonshine,”15 and in those with crampy abdominal pain, peripheral neuropathy, or encephalopathy. NORMOCYTIC ANEMIAS. MCV between 80-94 with normochromic normocytic red cells present on peripheral blood smear. Reviewing the patient's Blood Urea Nitrogen (BUN), Creatinine, SGOT, Alkaline Phosphatase, Bilirubin, Erythrocyte Sedimentation Rate (ESR), Urinalysis, and Thyroid profile may help in the diagnosis. A aspirated bone marrow biopsy is often necessary to aid the diagnosis if these simple tests do not provide the diagnosis DIFFERENTIAL DIAGNOSIS: "NORMAL SIZE" N: Normal pregnancy. Anemia of pregnancy results in part from a 30% expansion in the plasma volume.2 O: Overhydration and expanded plasma volume. R: Chronic renal disease. Anemia resulting from relative decrease in EPO production responds to recombinant EPO therapy.2,7,23 M: Myelophthisic anemia results from replacement of the bone marrow by a tumor, granulomatous infectious disease, fibrosis, or leukemia.24 A: Acute blood loss. L: Leukemia and liver disease. Chronic hepatitis, alcoholism, and cirrhosis may increase plasma volume and alter red cell survival. SI: Systemic inflammation (anemia of chronic inflammation).12,13 Z: Zero production. Aplastic anemia usually presents with pancytopenia.25 E: Endocrine disorders (eg, hypothyroidism, hyperthyroidism, adrenal insufficiency, hypogonadism).2 MACROCYTIC ANEMIAS. Common causes include Vitamin B12 and folate deficiency. Macrocytosis is seen with high reticulocyte counts, liver disease, obstructive jaundice, hypothyroidism, post splenectomy, and

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megaloblastic anemias. Megaloblastic anemias result from impaired nuclear maturation secondary to defective DNA synthesis.. Less common causes are related to inherited defects in DNA synthesis, vitamin metabolism, chemotherapy or marrow malignancy. Abnormal nuclear maturation also occurs in white cells, megakaryocytes, and other rapidly dividing cells. The peripheral blood changes include: Anemia with decreased reticulocyte count, increased MCV, and macro ovalocytes; neutropenia with hypersegmented neutrophils; and thrombocytopenia with large platelets. In the bone marrow, inhibition of DNA synthesis is reflected by a delay in nuclear maturation relative to cytoplasmic maturation and by ineffective erythropoiesis. Erythroid precursors are large with immature nuclear chromatin despite cytoplasmic maturity reflected by normal concentrations of hemoglobin. In myeloid cells this nuclear cytoplasmic asynergy is reflected by giant bands, giant metamyelocytes, and hypersegmented polys ( over 6 lobed nucleus). Megakaryocytes are large with increased polyploidy. Despite reticulocytopenia, neutropenia, and thrombocytopenia, the marrow is usually hypercellular reflecting ineffective erythropoiesis. Measurement of serum and red cell vitamin levels represents the most important first step in evaluating the megaloblastic anemias. Because of complex interactions, serum B12, serum folate, and red cell folate should be determined to make an accurate diagnosis. Patterns in folate and B12 deficiency are outlined in the Table below Vitamin B12 and Folate Levels in Deficiencies Vitamin Status _____________ Normal

Serum B 12 __________ Normal

Serum Folate RBC Folate __________ _________ Normal Normal

B12 Deficiency

Low

Normal or Low

Normal or High

Folate Deficiency

Normal or Low

Low

Low

Folate Deficient (early) Folate Deficient (refed)

Normal

Low

Normal

Normal or Low

Normal

Low

Because B12 is required for cellular uptake of folate, RBC folate is low in 1/3 of patients with B12 deficiency. In 1/2 of cases of severe folate deficiency, the serum B12 is low for unexplained reasons and will normalize with folic acid administration. Serum folate will fall within three to four weeks of initiating a folate poor diet, however, RBC folate will remain normal for three to five months reflecting tissue levels. Additional technical problems with the radioimmunoassay for B12 cause low serum B12 without other evidence of true deficiency. Increased urinary excretion of methylmalonic acid is specific for B12 deficiency and can be of diagnostic utility in difficult cases. Carefully supervised therapeutic trials of the vitamins may also be helpful Common causes of macrocytic (or megaloblastic) anemias include chemotherapy and vitamin B12 and folate deficiency; a less common cause is marrow malignancy. Rarely, inherited defects in DNA synthesis or vitamin metabolism may be the cause. The peripheral blood changes include macro-ovalocytes, neutropenia with hypersegmented neutrophils, and thrombocytopenia with large platelets.16 Levels of serum vitamin B12, serum folate, and red cell folate should be measured. The differential diagnosis is represented by the mnemonic “BIG FAT RED CELLS.” B: Vitamin B12 (cobalamin) deficiency.16,17 Patients may present with weakness, fatigue, dyspnea, paresthesias, and mental clouding. Physical signs include edema, pallor, jaundice, smooth tongue, decreased vibratory and position sensation, peripheral neuropathy, and long tract findings.18 In the US, inadequate B12 intake usually occurs only in strict vegans. However, about 70% of cases of vitamin B12 deficiency are caused by pernicious anemia—in which inadequate intrinsic factor or other conditions result in B12 malabsorption. This diagnosis is confirmed by the presence of anti–intrinsic factor antibodies or by positive results on the Schilling test.19 I: Inherited disorders (eg, orotic aciduria, Lesch-Nyhan syndrome, Di Guglielmo’s syndrome). These are rare.18

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G: Gastrointestinal disease or surgery. Small bowel disease, nontropical sprue, regional enteritis, ileal resection, celiac disease, fish tapeworm, or bacterial overgrowth all may block absorption of vitamin B12. F: Folic acid deficiency.16 Patient history may reveal anemia symptoms, alcoholism, or a diet lacking in fresh, uncooked fruits and vegetables (folates are rapidly destroyed by heat). Physical examination may show stigmata of alcoholic liver disease. The minimum daily requirement of folic acid is 50 µg; the average diet contains 700 µg, of which 10% is absorbed. Pregnant women, patients with hemolytic anemia, and those undergoing dialysis have increased folic acid requirements. A: Alcoholism (chronic).17,19 T: Thiamine-responsive anemia. This rare, unexplained condition is correctable with thiamine. R: Reticulocytes in large numbers may inflate the MCV, because they are larger than mature red cells.19 E: Endocrine disturbances (hypothyroidism).19 D: Dietary deficiencies, particularly folate, vitamin B12, protein, or lipids.16 C: Chemotherapeutic drugs, including methotrexate and 5-fluorouracil.20 E: Erythroleukemia. Circulating abnormal immature erythrocytes are larger than normal mature cells, inflating the MCV. L: Liver disease.17 Chronic hepatitis, alcoholism, and cirrhosis may increase MCV by altering the red cell membrane lipid composition. L: Lesch-Nyhan syndrome.21 This rare hereditary deficiency of hypoxanthine-guanine phosphoribosyltransferase activity is manifested by hyperuricemia, self-mutilation, mental retardation, spasticity, and choreoathetosis. S: Splenectomy. Loss of splenic filtration function results in larger erythrocytes with increased membrane, nucleated red blood cells (nRBCs), and nuclear fragments (Howell-Jolly bodies).22 Hemolytic Anemias Hemolytic anemias, which result from increased rate in destruction of red cells, will be discussed in some depth. Medical history, The CBC, Reticulocyte count, Chemistry profile with indirect and direct bilirubin, and LDH, urinalysis, and haptoglobin levels are most useful in diagnosing hemolysis. The past medical and family history, direct Coombs' test, and blood smear are most useful in establishing the cause once hemolysis is proven. The hallmark of hemolysis is the presence of an elevated reticulocyte count, with a stable or falling hemoglobin. Other findings include: -Elevated indirect bilirubin -Elevated serum lactate dehydrogenase (LDH)- Decreased Haptoglobin levels -Hemoglobinemia and hemoglobinuria -Erythroid hyperplasia in bone marrow The diagnostic tests that are very helpful in establishing the etiology of hemolysis are the clinical history, the direct antiglobulin (Coombs') test, Specific tests, such as hemoglobin electrophoresis, Heinz body stain, osmotic fragility will prove the diagnosis suggested by the history, blood smear and negative direct Coomb's test.

DIFFERENTIAL DIAGNOSIS : "HEMATOLOGIST".

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H: Hemoglobinopathies, with sickle cell disease among the most common (Type SS, SC, S-Beta Thalassemia, SS increased fetal) Most common feature is pain crisis from hypoxia, dehydration, infection. Children should be on Penicillin from birth to age 6. Consider hydroxyurea to prevent crisis Paroxysmal Nocturnal Hemoglobinuria.2 Manifestations include episodic intravascular hemolysis, thrombotic episodes, and increased infections.10 Diagnosis is based on flow cytometry for loss of antibodies CD55 and CD59.26 E: Enzyme deficiency (eg, inherited pyruvate kinase deficiency, pyrimidine-5'-nucleotidase deficiency).27,28 M: Medications, including sulfonamides, high-dose IV penicillin, quinine, quinidine, and chlorpromazine.29,30 A: Antibodies. In immune (Coombs’-positive) acquired hemolytic anemias, hemolysis is caused by an antigen that triggers antibody- or complement-mediated red cell destruction. Immune hemolytic anemia may be alloimmune (related to transfusion), autoimmune, or drug induced.10,31 Immunoglobulin G (IgG): Warm reacting antibody anemia32 occurs secondary to lymphoproliferative syndrome (in 30% of cases), collagen vascular diseases (20%), and other tumors (20%), or it can be idiopathic (20%). Laboratory findings are those of anemia with an increased reticulocyte count and detection of microspherocytes on the blood smear. Direct Coombs’ testing is positive for IgG or IgG and C3.33 IgM: Cold reacting antibody anemia32 occurs secondary to viral and mycoplasmal infections, with lymphoproliferative disease, and as idiopathic disorders in the elderly. T: Trauma to the red cells, as from an abnormal endothelium.10 O: Ovalocytosis is an autosomal dominant disorder seen in Southeast Asians.34 L: Liver disease, when severe, may lead to abnormal lipid loading in erythrocyte membranes, inducing formation of spur cells with shortened red cell survival.35 O: Osmotic fragility occurs with hereditary spherocytosis and hereditary elliptocytosis.36 The former is caused by autosomal dominant or recessive membrane protein abnormalities that result in sphere formation, membrane budding, and increased permeability to sodium. Hereditary elliptocytosis is caused by a defective red cell membrane, resulting from abnormal structural proteins. A finding of numerous elliptocytes on the blood smear is diagnostic. G: Glucose-6-phosphate dehydrogenase (G6PD) deficiency. Inherited G6PD deficiency is linked to the X chromosome, fully affecting homozygous females and males who inherit the deficient gene. Drugs or other substances cause increased oxidant stress within the red cell, triggering precipitation of hemoglobin into Heinz bodies and thereby causing hemolysis (most effectively identified by Heinz bodies on methyl violet stain). G6PD deficiency can be confirmed by molecular techniques or enzyme assay after two to three months. Patients should be advised to avoid antimalarials, high-dose aspirin, sulfa drugs, and fava beans.27,37 I: Infection. Intra-erythrocytic parasites (eg, malaria, babesiosis) lyse emerging red cells.10 They may be identified on examination of a thick smear slide for parasites. Clostridium infections and ß-hemolytic streptococcal septicemia may cause massive hemolysis. S: Splenic destruction in hypersplenism can occur with splenomegaly, portal hypertension secondary to infections, infiltration with leukemia or lymphoma, and collagen vascular diseases.22 T: Transfusion. Hemolysis may occur in a patient with antibodies directed against an antigen on the transfused red cells.31 The patient may experience an immediate acute transfusion reaction, or more commonly, a reaction that is delayed for five to 14 days. Additionally, thalassemia must be considered in a patient with hemolytic anemia and a low MCV.

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What causes polycythemia or too many red cells 1. Primary or Polycythemia vera. 2. Secondary Polycythemia a. High altitude. People ascending to altitudes above 2000 meters (6500 feet) b. Chronic lung disease with arterial hypoxemia. c. Cardiovascular right-to-left shunt. d. High-oxygen-affinity hemoglobinopathies. Erythrocytosis results from impaired release of oxygen to the tissues. e. Carboxyhemoglobinemia ("smokers erythrocytosis"). Erythrocytosis results from a functional anemia and left shift of the oxyhemoglobin dissociation curve caused by binding of carbon monoxide to hemoglobin f. Congenital decreased erythrocyte 2,3-DPG. g. Tumors producing erythropoietin or other erythropoietic substances neoplasm. h. Renal disease. These disorders lead to regional renal hypoxia, presumably affecting the tissue oxygen sensor and resulting in increased erythropoietin production. i. Adrenal cortical hypersecretion. Increased red cell mass may result from the production of adrenal androgens.

General References Platt A. Differential Diagnosis Mnemonics.and the Medical History 3nd Edition. Palm book at http://www.EmoryPA.org Eckman J, Platt A. Problem Oriented Management of Sickle Syndromes. National Maternal and Child Health Clearing House. 1991 and 2000. Available online http://www.SCInfo.org. Platt A, Eckman J, Diagnosing Anemia, Clinician Reviews Vol 16 No 12 December 2006 p 43 – 50. Web http://www.clinicianreviews.com/print.asp?page=courses/105417/lesson.htm Wintrobe's Clinical Hematology by Author(s): John P Greer MD; John Foerster MD, FRCP(C); John N Lukens MD; George M Rodgers MD, PhD; Frixos Paraskevas MD; Bertil E Glader MD, PhD 11th edition (2003) Lippincott, Williams & Wilkins

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