Megaloblastic Anemia: Vitamin B12 deficiency & Folate deficiency

Introduction 

Megaloblastic anemias are characterized by ineffective erythropoiesis, results from a defect in red cell DNA synthesis which alters and delays RBC maturation resulting in altered (“megaloblastic”) differentiation.

The RBCs resulting from megaloblastic differentiation are large (macroovalocytes) due to delayed maturation; corresponding changes in the granulocytic cell line lead to multisegmented polymorphonuclear leukocytes.

What's Causes of Megaloblastic anemias? 

The cause of megaloblastic anemia is vitamin B12 or folic acid deficiency. Both types of anemia result in the production of large, immature red blood cells (megaloblasts), but the specific deficiencies and their causes distinguish the two conditions.

• Folate is required for normal DNA synthesis; inadequate availability of folate causes megaloblastic differentiation in the red cell line.
• B12 deficiency results in megaloblastosis by reducing folate availability for RBC maturation 
• In pernicious anemia (PA) the absence of B12 causes a deficiency of usable folate at the cellular level resulting in megaloblastic differentiation. 

Combined deficiencies of cobalamin & folic acid is seen in tropical sprue.

Features of Megaloblastic anemia

• The earliest manifestation of megaloblastic anemia is presence of hypersegmented neutrophils (neutrophils having > 5 lobes). 
• Triad of megaloblastic anemia includes oval macrocytes, Howell Jolly bodies and hypersegmented neutrophils.
• Diagnosis is made if even a single neutrophil with ≥ 6 lobes is seen or > 5% neutrophils with 5 lobes are seen.
• Significant and marked macrocytosis (MCV > 100 fL) suggests presence of a megaloblastic anemia.
• Reticulocyte index is low, & leukocyte and platelet count may also be decreased, particularly in severely anemic patients. 
• MCHC is not elevated in mega­loblastic anemia be­cause hemoglobin increases proportionately to the increased volume of the RBCs.
• The serum homocysteine level is increased in both folate deficiency and vitamin B12 deficiency. However, serum methylmalonic acid levels are only seen in vitamin B12 deficiency.

Raised urine urobilinogen, reduced haptoglobins and positive urine hemosiderin, and a raised serum lactate dehydrogenase provides evidence of ineffective erythropoiesis.

Important to determine whether folate or vitamin B12 deficiency is the cause megaloblastic anemia prior to initiating treatment.  

Megaloblastic anemia caused due to folic acid deficiency is clinically indistinguishable from vitamin B12. However, folic acid deficiency is NOT associated with neurological abnormalities.

Vitamin B12 is necessary for normal myelin synthesis and deficient supply of this vitamin underlies the neurologic abnormalities that occur in PA.

Folate supplements can improve the anemia of vitamin B12 deficiency, but not the neurologic impairment. Therefore, if the vitamin B12 deficiency remains untreated, irreversible neurologic disease can result. Folic acid supplements should never be given to patients with megaloblastic anemia until PA has been ruled out.

Note that patients with vitamin B12 deficiency can have moderate-to-severe neurologic symptoms with little-tono anemia (i.e., blood counts may be normal). Delay in diagnosis and treatment may lead to irreversible neurologic disease.

Vitamin B12 Deficiency 

The normal range of serum cobalamin in serum is 160 - 1000 ng/L. 

• Values between 100 & 200 ng/L are regarded as borderline.
• Values <200 pg/mL indicate clinically significant cobalamin deficiency.

Most commonly occurs due to impaired absorption (either via lack of intrinsic factor or lack of absorptive ileal surface)

• Cobalamin absorption can be passive through buccal, duodenal & ileal mucosa.
• More efficient normal physiologic active mechanism is through ileum mediated by gastric intrinsic factor (IF).
• Vitamin B12 binds to IF to form a complex.
• Vitamin B12–IF complex is absorbed in the terminal ileum.
• Vitamin B12 binds to transcobalamin II and enters the plasma.
• Delivered to metabolically active cells or stored in the liver (6- to 9-year supply). Body stores of 2 - 3 mg are sufficient for 3 - 4 years 

Increased serum homocysteine and methylmalonic acid (95% of cases) 

• ↑Methylmalonic acid: most sensitive test for B12 deficiency

Causes of Vitamin B12 Deficiency

1. Pernicious anemia—lack of intrinsic factor due to autoimmune destruction of parietal cells, the most common cause of vitamin B12 deficiency. 
2. Gastrectomy—lack of intrinsic factor due to removal of parietal cells 
3. Inadequate dietary intake (e.g., strict vegetarianism, alcoholism)
4. Malabsorption: Crohn disease or Removal of 1.2 meters of terminal ileum causes malabsorption of cobalamin.
5. Diphyllobothrium latum (fish tapeworm), the worms absorb more than 80% of the vitamin B12 intake
6. Blind loop syndrome (bacterial overgrowth): bacteria use available vitamin B12
7. Chronic pancreatitis: enzyme deficiency leads to inability to cleave R-binder from the vitamin B12–R-binder complex

Clinical Features

The clinical features of cobalamin deficiency involve the blood, the gastrointestinal tract, and the nervous system.

1. Glossitis associated with a smooth, sore tongue and atrophy of the papillae 
2. Peripheral neuropathy with sensorimotor dysfunction
3. Subacute combined degeneration (demyelination) of spinal cord 
4. Dementia from involvement of the brain  

Treatment

Cyanocobalamin (vitamin B12) IM—parenteral therapy is preferred

1. Replenishment of body stores of cobalamin is complete with six 1000-µg IM injections of hydroxocobalamin given at 3- to 7-day intervals.
2. For maintenance therapy, 1000 µg hydroxocobalamin IM once every 3 months is satisfactory.

Reticulocytosis begins 4 to 5 days after intramuscular cyanocobalamin therapy is started & peaks at ~ day 7.

Pernicious anemia

Pernicious anemia is the most common cause of cobalamin deficiency. It is caused by absence of IF, due to atrophy of gastric mucosa or autoimmune destruction of parietal cells.

Most characteristic finding in pernicious anemia is gastric atrophy affecting the acid and pepsin-secreting portion of the stomach while sparing the antrum.

• 90% patients with PA have antiparietal cell antibody directed against H+,K+ -ATPase, while IF antibodies are detected in gastric juice in ~80%.

Helicobacter pylori does not cause parietal cell destruction in pernicious anemia. 

The anemia in PA develops chronically and may be very severe with Hgb as low as 3 or 4 g/dL. This necessitates slow and cautious transfusion to avoid pulmonary edema. 

Plasma levels of B12 levels are much less reliable than methylmalonic levels in the diagnosis of PA.

Three antibodies that are associated with the pathophysiology of PA and its clinical findings include: 

1) Type I antibodies: Antibodies that block the binding of vitamin B12 to intrinsic factor (in 75% patients; Most specific test for PA).
2) Type II antibodies: Antibodies that prevent the binding of vitamin B12 –intrinsic factor complexes to ileal receptors (30%–50% of cases)
3) Type III antibodies: Antibodies directed against the proton pump in parietal cells (seen in 90% patients but not specific as it also seen in idiopathic chronic gastritis).

Antibodies that attack parietal cells in the body/fundus (type II hypersensitivity reaction) produce a chronic atrophic gastritis that is associated with achlorhydria (lack of gastric acid) and a loss of IF. 

1. Loss of acid prevents proper digestion of food in the stomach, which interferes with the release of vitamin B12 from food. 
2. Loss of IF, decreases absorption of vitamin B12 in the terminal ileum.
3. Achlorhydria leads to a corresponding increase in serum gastrin (hypergastrinemia; negative feedback).
4. Intestinal metaplasia in the chronic atrophic gastritis, increases the risk for developing adenocarcinoma of the body/fundus.

Other antibodies associated with PA also contribute to producing vitamin B12 deficiency.

Clinical Features

1. Megaloblastic anemia
2. Pancytopenia (Leucopenia with hypersegmented neutrophils, thrombocytopenia) 
3. Jaundice due to ineffective hematopoiesis and peripheral hemolysis 
4. Neurological features due to posterolateral spinal tract involvement.
5. A peculiar lemon yellow appearance of the skin: The anemia (pallor) in concert with low-grade jaundice may produce a distinctive “lemon yellow” coloration of the skin.
6. Personality change and outright dementia may occur as well.

Pernicious anemia is associated with increased risk of gastric cancer and increased chances of atherosclerosis and throm­bosis (because of elevated homocysteine levels).

Subacute combined degeneration of the spinal cord.

• Initial pathology is demyelination, followed by axonal degeneration & eventual neuronal death.
• Sites of involvement include peripheral nerves; the spinal cord, where the posterior and lateral columns undergo demyelination; and the cerebrum itself.
• C/F: Numbness and paresthesia in the extremities (the earliest neurologic manifestations), Weakness, and ataxia. Also can lead to urinary and fecal incontinence, impotence

Folate Deficiency  

Folic acid deficiency is the other cause of megaloblastic anemia, is caused by inadequate nutrition or impaired folate absorption.

• Normal serum folic acid ranges from 6 to 20 ng/mL. 
• Values < 4 ng/mL are considered diagnostic of folate deficiency.

Inadequate dietary intake (e.g., “tea and toast” diet, alcoholism)—most common cause

• Folate deficiency is very common in alcoholics due to poor dietary intake and impaired absorption.
• Pregnancy produces folate deficiency due to increased requirements.
• Folate malabsorption is common in celiac sprue. 

Folic acid stores in the body are limited, and inadequate intake over a 3-month period can lead to deficiency. Neonatal folate level falls rapidly to the lowest values at about 6 weeks of age.

• Total-body folate in the adult is ~10 mg, liver containing the largest store.

Clinical features folate deficiency is Similar to those of vitamin B12 deficiency, but no neurologic symptoms 

Treatment: Daily oral folic acid replacement
Fruits and vegetables are the primary dietary source of folic acid.

Patients on chronic hemodialysis require folate supplementation to replace that lost in dialysate.