Pernicious Anaemia and B12 Deficiency

Pernicious anaemia accounts for 80% of cases of megaloblastic anaemia due to impaired absorption of vitamin B12.¹
Vitamin B12 is present in meat and animal protein foods. Absorption occurs in the terminal ileum and requires intrinsic factor, a secretion of gastric mucosal (parietal) cells, for transport across the intestinal mucosa. In pernicious anaemia, intrinsic factor production is deficient. It is believed to be an autoimmune disease.

Parietal-cell antibody and antibodies to intrinsic factor are found in nearly all cases. 90% of patients have antibodies to parietal cells and their components, including antibodies to intrinsic factor and the proton pump H , K -ATPase.² 50% of patients have thyroid antibodies.¹

Pernicious anaemia may be associated with simple gastric atrophy in 15% of people age 40-60 and 20-30% of the older population. Pathology shows gastritis with all layers of the body and fundus atrophied. The antrum is spared in > 80% of patients (type A gastritis).³

The incidence of the disease is 1:10,000 in northern Europe. The disease occurs in all races. The peak age is 60, and the condition is commoner in those with blue eyes, early greying, a positive family history and blood group A. The condition has a female:male ratio of 1.6:1.0.

• The onset is usually insidious as B12 stores in the liver are depleted, and starts with symptoms of anaemia - i.e. lethargy and breathlessness. The anaemia may be more severe than symptoms suggest due to physiological adaptation.
• Other symptoms may include anorexia, weight loss, diarrhoea, and dyspepsia. Glossitis may be an early symptom.
• Pernicious anaemia may first present as an incidental finding during the investigation of (reversible) diarrhoea.⁴
• Neurological involvement may be present even in the absence of anaemia. This is particularly common in patients over the age of 60.⁵ The peripheral nerves are most commonly involved, followed by the spinal cord (subacute degeneration of the cord).
• Peripheral loss of vibratory sense and position are early indications of CNS involvement, accompanied by reflex loss and mild to moderate weakness. Later stages may be characterised by spasticity, Babinski's responses and ataxia.
• Other uncommon neurological symptoms include impairment of pain, temperature and touch sensations may be impaired. The legs and feet are involved earlier and more consistently than the hands.
• Yellow-blue blindness may occur.
• Psychiatric symptoms (usually more prominent in advanced cases) may include depression, paranoia (megaloblastic madness), delirium, confusion and dementia.³
• Signs may include anaemia and jaundice.
• Severely anaemic patient may present with heart failure, often triggered by an infection. Hepatomegaly⁶ and splenomegaly⁷ may be present.

Causes of vitamin B12 deficiency

• Poor quality diet, vegetarian diet
• Gastric causes - gastrectomy, congenital intrinsic factor deficiency
• Intestinal causes - stagnant loop, congenital selective malabsorption, ileal resection, inflammatory bowel disease
• Infestation - fish-tapeworm
• Metabolic causes - transcobalamin II deficiency, nitrous oxide anaesthesia
• Drugs causing decreased B12 levels - oral calcium-chelating agents,³ aminosalicylic acid,³ biguanides

Causes of megaloblastic anaemia

• Folate deficiency - poor diet, goat's milk,⁸ gluten-induced enteropathy, tropical sprue, pregnancy, prematurity, chronic haemolytic anaemias (e.g. sickle cell anaemia), malignant disease, increased renal loss (congestive cardiac failure, dialysis), drugs (anticonvulsants, sulphasalazine)

Causes of macrocytosis¹

• Alcohol excess - commonest cause of macrocytosis in the UK,⁹ may co-exist with folate deficiency in spirit drinkers¹⁰ (not seen in beer drinkers due to high folic acid content in beer¹¹)
• Liver disease
• Severe hypothyroidism
• Reticulocytosis (e.g. post acute blood loss or haemolytic anaemia)¹²
• Other blood disorders - red-cell aplasia, aplastic anaemia, myeloid leukaemia, myelodysplastic disorders
• Changes in plasma proteins (e.g. increased paraprotein secondary to multiple myeloma) may cause a spurious rise in mean cell volume without the presence of macrocytes¹³
• Drugs that affect DNA synthesis - e.g. azathioprine, hydroxyurea

Tests commonly performed in primary care

• Full blood count
  • This may show low haemoglobin and increased mean cell volume (MCV) - although macrocytosis can precede the development of anaemia. Severe cases may show a pancytopaenia.
  • The reticulocyte count may be low for the degree of anaemia (1-3% only).
  • The MCV may be normal if there is associated iron deficiency.
• The blood film This may show macrocytic red cells, neutrophils with hypersegmented nuclei and Howell-Jolly bodies (residual fragments of the nucleus causing spherical blue-black inclusions on red blood cells seen on Wright-stained smears.¹⁴ Associated iron deficiency may result in the MCV being normal, in which case two types or red blood cells may be seen (a dimorphic blood film.¹⁵ The ferritin level should be checked if such a picture is seen.
• Biochemistry
  • There may be an increase in plasma unconjugated bilirubin due to increased destruction of red-cell precursors in the marrow. Liver and thyroid function tests, and protein electrophoresis may help in the differential diagnosis of macrocytosis.
  • Serum vitamin B12 is the most commonly used method of establishing B12 deficiency. In general, levels < 150 pg/ml reliably indicate deficiency. Neurological deficiency or anaemia is usually evident in patients with levels < 120 pg/ml. False positives (low levels in the absence of deficiency) can occur with pregnancy, folate deficiency,¹⁶ myeloma,¹⁷ and excessive vitamin C intake.
  • False negatives (normal levels in the presence of deficiency) may occur in true deficiency,¹⁸ liver disease, lymphoma, autoimmune disease, and myeloproliferative disorders. In borderline cases or where B12 deficiency is clinically suspected, other tests must be carried out. Tissue deficiency of B12 results in raised levels of serum methylmalonic acid, and this is a useful test where false positive of negative values are suspected. Other tests include trancobalamin II-B12 content, and plasma total homocysteine.¹⁹
  • Folic acid levels should be measured to exclude deficiency, which may co-exist with B12 deficiency. Red-cell folate is a better guide to deficiency than serum folate.¹ B12 deficiency may result in increased serum folate levels but reduced red-cell folate levels, because of the effect on intracellular folate metabolism.²⁰ Combined deficiency usually results in both reduced serum folate and vitamin B12 levels.¹³
• Autoantibody screen Intrinsic factor (IF) antibodies, if present, are virtually diagnostic of pernicious anaemia. However, they are absent in 50% of patients with pernicious anaemia. Gastric parietal-cell antibodies are present in 85% of people with pernicious anaemia, but are also found in 3-10% of people who do not have pernicious anaemia.

Tests which may be performed in secondary care

• The Schilling test The purpose of this test is to differentiate patients whose B12 deficiency is due to pernicious anaemia from those who have an intestinal lesion causing malabsorption. It measures the absorption of B12 with and without intrinsic factor.
  • The patient must not take B12 for five days prior to the test.
  • Radioactive B12 is given orally followed in one to six hours by a parental B12 'flushing' dose (1000 mcg) to avoid liver storage of radioactive B12.
  • The percentage or radiolabelled material in a 24 hour urine collection is then measured (normally > 9% of the dose given).
  • Reduced urinary excretion (< 5%) in the presence of normal kidney function supports the diagnosis of decreased absorption of vitamin B12.
  • Repeating the first test (Schilling I) using radiolabelled cobalt attached to intrinsic factor from a hog (Schilling II) will confirm if absorption is increased, thus supporting the diagnosis of pernicious anaemia.³
  The Schilling test has its limitations:
  • Radiolabelled vitamin B12 is difficult to obtain, it is complicated to perform, and test results can be difficult to interpret in (often elderly) patients with renal insufficiency.²¹
  • Because the Schilling test does not measure absorption of food-bound B12, the test will not detect defective liberation of food-bound B12 in the elderly patient.²² Furthermore, the test result often does not contribute much to the ultimate management of the patient.
  If a Schilling test is felt inappropriate, in elderly patients with a low vitamin B12 level and negative intrinsic factor antibodies, response to vitamin B12 may be adequate to confirm a diagnosis of pernicious anaemia if:
  • The person feels better in 1-2 days
  • The reticulocyte count increases in 2-3 days, and peaks in 5-8 days
  • The red blood cell count increases within 1 week, and normalizes in 4-8 weeks
  • The MCV increases for 3-4 days (due to the increased reticulocyte count), then decreases, reaching the normal range in 25-78 days
  • Haemoglobin level increases by 2-3 g/dl every 2 weeks
  • White blood cell and platelet counts normalise in 7-10 days
• Bone-marrow aspiration may be necessary to narrow the differential diagnosis, especially if myelodysplasia, aplastic anaemia, myeloma, or other marrow disorders are suspected. In B12 and folate deficiency, megaloblasts and giant metamyelocytes (early granulocyte precursors) are seen.²³
• Gastroscopy is appropriate on diagnosis to confirm gastric atrophy and exclude gastric cancer and polyps.²⁴ Gastric cancer is two to three times commoner in patients with pernicious anaemia than in matched controls.¹³

• Vitiligo
• Myxoedema
• Hashimoto's disease
• Addison's disease
• Giant cell myocarditis
• Hypoparathyroidism
• Diabetes mellitus

• For patients with no neurological involvement, treatment is with six injections of hydroxocobalamin, 1 mg at intervals of between 2-4 days.
• Subsequently, 1 mg should be given every 3 months for life.
• For patients with neurological involvement, referral to a haematologist is recommended. Initial treatment is with hydroxocobalamin 1 mg on alternate days until there is no further improvement, after which 1 mg should be given every 2 months for life.¹
• Care should be taken not to give folic acid (instead of B12) to any patient who is B12 deprived as this may result in fulminant neurologic deficit.³
• Oral iron therapy should be given before B12 if iron deficiency is diagnosed by an absence of stainable Fe in the bone marrow or other parameters (eg, serum ferritin < 200 ng/ml).³

When to refer¹

Referral to a gastroenterologist should be considered for any patient with pernicious anaemia who has gastric symptoms and/or co-existent iron deficiency. Patients with pernicious anaemia have a 2-3 times increased incidence of gastric carcinoma and gastric polyps compared to matched controls.

• Heart failure - this may be secondary to anaemia, or rarely, myocarditis.
• Angina
• Neuropathy - subacute combined degeneration of the cord, optic atrophy, neurosis, depression, and dementia
• Gastric carcinoma
• Infertility (rare).²⁵

Pernicious anaemia responds rapidly to replacement therapy. However, if the deficiency has been severe and prolonged, any neurological complications may be irreversible.¹