18:Megaloblastic and Macrocytic Anemias

• Megaloblastic anemia stems from impaired DNA synthesis, hindering proper red blood cell maturation and division, causing abnormally large cells (megaloblasts) with distinctive morphology.
• Reverse transcriptase inhibitors, used for HIV treatment, can interfere with DNA production and induce megaloblastic changes.
• Screening for megaloblastic anemia involves five tests: complete blood count (CBC), reticulocyte count, white blood cell (WBC) manual differential, serum bilirubin, and lactate dehydrogenase.

Macrocytic Nonmegaloblastic Anemias

• Characterized by larger-than-normal red blood cells (RBCs) with unimpaired DNA synthesis.
• Do not involve defects in DNA synthesis, unlike megaloblastic anemias.
• MCV values typically range between 100 and 115 fL, less pronounced than in megaloblastic forms.
• RBCs appear larger than normal but maintain a round shape; reticulocytes may increase if anemia results from increased RBC production.
• Bone marrow shows increased erythropoiesis without megaloblastic changes.
• Common causes include liver disease (altering lipid metabolism and affecting RBC membrane), chronic alcoholism (affecting bone marrow and RBC production), and reticulocytosis (immature RBCs are larger).
• Diagnosis involves CBC, peripheral blood smear, bone marrow biopsy, liver function tests, and alcohol use assessment.

Relationships Among Macrocytic Anemia, Megaloblastic Anemia, and Pernicious Anemia

• Macrocytic anemia features abnormally large red blood cells (macrocytes) with a high mean corpuscular volume (MCV).
• Macrocytic anemia is classified into megaloblastic and non-megaloblastic anemias.
• Megaloblastic anemia, a subtype of macrocytic anemia, includes megaloblasts (large, immature red blood cells) in the bone marrow.
• Megaloblastic anemia results from impaired DNA synthesis due to deficiencies in vitamin B12 or folate.
• Pernicious anemia, a specific type of megaloblastic anemia, is caused by an autoimmune response leading to vitamin B12 deficiency due to the destruction of gastric parietal cells.

Classification of Anemias

• Macrocytic Anemia:
  • Megaloblastic Anemia: Caused by vitamin B12 or folate deficiency with the presence of megaloblasts.
    • Pernicious Anemia: Subtype caused by autoimmune destruction of parietal cells and lack of intrinsic factor.
  • Non-Megaloblastic Macrocytic Anemia: Not related to impaired DNA synthesis, examples include liver disease, hypothyroidism, and alcoholism.

Vitamin B12 (cobalamin)

• Essential nutrient with a cobalt-containing ring found in forms like hydroxycobalamin and cyanocobalamin.
• Functions as a coenzyme in DNA production and metabolic pathways.
• Crucial in the transfer of a methyl group from 5-methyltetrahydrofolate (5-methyl THF) to homocysteine, generating methionine and tetrahydrofolate (THF) catalyzed by methionine synthase.
• Requires methylcobalamin as a coenzyme.
• Impaired activity leads to elevated levels of serum methylmalonic acid (MMA).

Folate (Vitamin B9)

• Involved in nucleotide synthesis, particularly in the conversion of deoxyuridine monophosphate (dUMP) to deoxythymidine monophosphate (dTMP).
• Catalyzed by thymidylate synthase and requires 5,10-methylene THF.
• Participates in the transfer of methyl groups in metabolic processes and is essential for amino acid metabolism and methylation reactions.

Absorption and Distribution of Vitamin B12

• Vitamin B12 is released in the acidic environment of the stomach aided by pepsin and is bound by salivary protein, haptocorrin.
• Pancreatic proteases release vitamin B12 from haptocorrin in the small intestine.
• Vitamin B12 is then bound by intrinsic factor, produced by gastric parietal cells and the vitamin B12-intrinsic factor complex is absorbed by ileal enterocytes through receptors.
• Once inside the enterocyte, vitamin B12 is freed from intrinsic factor and bound to transcobalamin, and released into the circulation.
• Vitamin B12-transcobalamin complex is termed holotranscobalamin (holoTC) and is the metabolically active form of vitamin B12.

Biochemical Basis for Development of Anemia with Vitamin B12 Deficiency

• Reduced gastric acidity prevents vitamin B12 release from food proteins for binding to intrinsic factor.
• Lack of gastric acidity or trypsin prevents vitamin B12 from separating from haptocorrin.
• Lack of intrinsic factor leads to insufficient vitamin B12 absorption, autoimmune diseases destroy gastric parietal cells, reducing intrinsic factor secretion with antibodies to intrinsic factor blocking vitamin B12 binding.
• Both Vitamin B12 and folate deficiencies impair DNA synthesis resulting in megaloblastic anemia.
• Impaired DNA synthesis results in the formation of large, immature red blood cells (megaloblasts) in the bone marrow.

Individuals at Risk for Megaloblastic Anemia

• Elderly at risk due to reduced gastric acidity and impaired vitamin B12 release from food.
• Strict vegetarians (vegans) at risk due to the absence of vitamin B12 in plant-based diets.
• Pregnancy increases demand for folate, renal dialysis patients lose folate, and certain drug regimens decrease absorption of folic acid.
• Pathological conditions include food-cobalamin malabsorption, chronic pancreatic disease, autoimmune diseases, Helicobacter pylori infection, gastrectomy, and hereditary intrinsic factor deficiency.

Screening Tests

• Five tests used to screen for megaloblastic anemia are:
  • Complete Blood Count (CBC)
  • Reticulocyte Count
  • White Blood Cell (WBC) Manual Differential
  • Serum Bilirubin
  • Lactate Dehydrogenase (LDH)

Complete Blood Count and Reticulocyte Count

• Macrocytosis is often the earliest sign.
• Decreased hemoglobin and hematocrit values, pancytopenia, and reticulocytopenia are expected, hemoglobin values can be less than 7 or 8 g/dL, and hematocrit can be less than 20%.
• MCV is usually 100 to 150 fL, but commonly greater than 120 fL, MCH is elevated, but MCHC is usually within the reference interval, and RDW is elevated.
• Oval macrocytes and hypersegmented neutrophils are characteristic morphologic findings.

White Blood Cell Manual Differential Count

• Hypersegmentation of neutrophils is essentially pathognomonic; mean lobe count should be greater than 3.4.

Bilirubin and Lactate Dehydrogenase Levels

• Elevation in levels of total and indirect bilirubin and lactate dehydrogenase (predominantly RBC derived) are indicative findings.

Bone Marrow Examination

• Nuclear-cytoplasmic asynchrony, bone marrow hypercellularity with reduced myeloid-to-erythroid ratio of about 1:1, and giant metamyelocytes and bands may be observed.

Sequence of Development of Megaloblastic Anemias

• Decrease in vitamin levels, hypersegmentation of neutrophils, oval macrocytes in peripheral blood, megaloblastosis in bone marrow, and anemia.

Step-by-Step Interpretation

• Low Vitamin B12 indicates a potential deficiency.
• Elevated MMA suggests a deficiency in vitamin B12; normal MMA is unlikely.
• Low Serum Folate indicates folate deficiency; normal serum folate needs RBC folate values.
• Elevated Homocysteine can indicate deficiencies in either folate or vitamin B12.
• Positive for Intrinsic Factor Antibodies is highly specific for pernicious anemia.
• If serum B12 is low, MMA is elevated, and homocysteine levels are high, indicated Vitamin B12 Deficiency.
• If positive intrinsic factor antibodies are present along with low serum B12, elevated MMA, and homocysteine, pernicious anemia is indicated.
• False increases and decreases in assay results can occur due to various factors, elevated serum gastrin and achlorhydria may support the diagnosis of pernicious anemia, and Metabolically active form of vitamin B12 can be measured for more precise diagnosis.

Results of Bilirubin and Lactate Dehydrogenase (LDH) Tests in Megaloblastic Anemia

• Elevated levels are indicative of hemolysis, Elevated LDH levels are due to high turnover of erythroid progenitors.
• Hemolysis leads to increased bilirubin production, and a constellation of findings justifies further testing.

Homocysteine Levels

• These are affected by deficiencies in either folate or vitamin B12.