Megaloblastic Anemia

MEGALOBLASTIC ANEMIA

Introduction

This results from a deficiency of vitamin B12 or folic acid, or from disturbances in folic acid metabolism. Folate is an important substrate of, and vitamin B12 a co-factor for, the generation of the essential amino acid methionine from homocysteine. This reaction produces tetrahydrofolate, which is converted to thymidine monophosphate for incorporation into DNA. Deficiency of either vitamin B12 or folate will therefore produce high plasma levels of homocysteine and impaired DNA synthesis. The end result is cells with arrested nuclear maturation but normal cytoplasmic development: so-called nucleocytoplasmic asynchrony. All proliferating cells will exhibit megaloblastosis; hence changes are evident in the buccal mucosa, tongue, small intestine, cervix, vagina and uterus. The high proliferation rate of bone marrow results in striking changes in the haematopoietic system in megaloblastic anaemia. Cells become arrested in development and die within the marrow; this ineffective erythropoiesis results in an expanded hypercellular marrow. The megaloblastic changes are most evident in the early nucleated red cell precursors, and haemolysis within the marrow results in a raised bilirubin and lactate dehydrogenase (LDH), but without the reticulocytosis characteristic of other forms of haemolysis. Iron stores are usually raised. The mature red cells are large and oval, and sometimes contain nuclear remnants. Nuclear changes are seen in the immature granulocyte precursors and a characteristic appearance is that of ‘giant’ metamyelocytes with a large ‘sausage-shaped’ nucleus. The mature neutrophils show hypersegmentation of their nuclei, with cells having six or more nuclear lobes. If severe, a pancytopenia may be present in the peripheral blood. Vitamin B12 deficiency, but not folate deficiency, is associated with neurological disease in up to 40% of cases, although advanced neurological disease due to B12 deficiency is now uncommon in the developed world. The main pathological finding is focal demyelination affecting the spinal cord, peripheral nerves, optic nerves and cerebrum. The most common manifestations are sensory, with peripheral paraesthesiae and ataxia of gait.

 Clinical features of megaloblastic anaemia

Symptoms

 • Malaise (90%)
 • Breathlessness (50%)
 • Paraesthesiae (80%)
 • Sore mouth (20%)
 • Weight loss
 • Impotence
 • Poor memory
 • Depression
 • Personality change
 • Hallucinations
 • Visual disturbance

Signs 

 • Smooth tongue
 • Angular cheilosis
 • Vitiligo
 • Skin pigmentation
 • Heart failure
 • Pyrexia

Neurological findings in B12 deficiency

Peripheral nerves

• Glove and stocking paraesthesiae
• Loss of ankle reflexes

Spinal cord

• Subacute combined degeneration of the cord
Posterior columns – diminished vibration sensation and
proprioception
Corticospinal tracts – upper motor neuron signs
Cerebrum
• Dementia
• Optic atrophy
Autonomic neuropathy

Investigations 

• Haemoglobin Often reduced, may be very low
• Mean cell volume Usually raised, commonly > 120 fL 
• Erythrocyte count Low for degree of anaemia Blood film Oval macrocytosis, poikilocytosis, red cell fragmentation, neutrophil hypersegmentation
• Reticulocyte count Low for degree of anaemia
• Leucocyte count Low or normal
• Platelet count Low or normal
• Bone marrow Increased cellularity, megaloblastic changes in erythroid series, giant metamyelocytes, dysplastic megakaryocytes, increased iron in stores, pathological non-ring sideroblasts  
• Serum ferritin Elevated
• Plasma lactate dehydrogenase Elevated, often markedly

Vitamin B12 absorption

The average daily diet contains 5–30 μg of vitamin B12, mainly
in meat, fish, eggs and milk – well in excess of the 1 μg daily
requirement. In the stomach, gastric enzymes release vitamin B12
from food and at gastric pH it binds to a carrier protein termed
R protein. The gastric parietal cells produce intrinsic factor, a
vitamin B12-binding protein that optimally binds vitamin B12 at
pH 8. As gastric emptying occurs, pancreatic secretion raises
the pH and vitamin B12 released from the diet switches from
the R protein to intrinsic factor. Bile also contains vitamin B12
that is available for reabsorption in the intestine. The vitamin
B12–intrinsic factor complex binds to specific receptors in the
terminal ileum, and vitamin B12 is actively transported by the
enterocytes to plasma, where it binds to transcobalamin II, a
transport protein produced by the liver, which carries it to the
tissues for utilisation. The liver stores enough vitamin B12 for
3 years and this, together with the enterohepatic circulation,
means that vitamin B12 deficiency takes years to become manifest,
even if all dietary intake is stopped or severe B12 malabsorption
supervenes.
Blood levels of vitamin B12 (cobalamin) provide a reasonable
indication of tissue stores, are usually diagnostic of deficiency and
remain the first-line tests for most laboratories. Additional tests
have been evaluated, including measurement of methylmalonic
acid, holotranscobalamin and plasma homocysteine levels, but donot add much in most clinical situations. Levels of cobalamins fall
in normal pregnancy. Reference ranges vary between laboratories
but levels below 150 ng/L are common and, in the last trimester,
5–10% of women have levels below 100 ng/L. Spuriously low
B12 values occur in women using the oral contraceptive pill and
in patients with myeloma, in whom paraproteins can interfere
with vitamin B12 assays.
Causes of vitamin B12 deficiency
Dietary deficiency
This occurs only in strict vegans but the onset of clinical features
can occur at any age between 10 and 80 years. Less strict
vegetarians often have slightly low vitamin B12 levels but are not
tissue vitamin B12-deficient.
Gastric pathology
Release of vitamin B12 from food requires normal gastric acid
and enzyme secretion, and this is impaired by hypochlorhydria
in elderly patients or following gastric surgery. Total gastrectomy
invariably results in vitamin B12 deficiency within 5 years, often
combined with iron deficiency; these patients need life-long
3-monthly vitamin B12 injections. After partial gastrectomy, vitamin
B12 deficiency only develops in 10–20% of patients by 5 years;
an annual injection of vitamin B12 should prevent deficiency in
this group.

Pernicious anaemia
This is an organ-specific autoimmune disorder in which the gastric
mucosa is atrophic, with loss of parietal cells causing intrinsic
factor deficiency. In the absence of intrinsic factor, less than 1%
of dietary vitamin B12 is absorbed. Pernicious anaemia has an
incidence of 25/100 000 population over the age of 40 years in
developed countries, but an average age of onset of 60 years.
It is more common in individuals with other autoimmune disease
(Hashimoto’s thyroiditis, Graves’ disease, vitiligo or Addison’s
disease) or a family history of these or pernicious anaemia.
The finding of anti-intrinsic factor antibodies in the context of B12
deficiency is diagnostic of pernicious anaemia without further
investigation. Antiparietal cell antibodies are present in over 90%
of cases but are also present in 20% of normal females over the
age of 60 years; a negative result makes pernicious anaemia
less likely but a positive result is not diagnostic. The Schilling
test, involving measurement of absorption of radio-labelled B12
after oral administration before and after replacement of intrinsic
factor, has fallen out of favour with the availability of autoantibody
tests, greater caution in the use of radioactive tracers, and limited
availability of intrinsic factor.
Small bowel pathology
One-third of patients with pancreatic exocrine insufficiency fail
to transfer dietary vitamin B12 from R protein to intrinsic factor.
This usually results in slightly low vitamin B12 values but no tissue
evidence of vitamin B12 deficiency.
Motility disorders or hypogammaglobulinaemia can result in
bacterial overgrowth, and the ensuing competition for free vitamin
B12 can lead to deficiency. This is corrected to some extent by
appropriate antibiotics.
A small number of people heavily infected with the fish
tapeworm develop vitamin B12 deficiency.
Inflammatory disease of the terminal ileum, such as Crohn’s disease, may impair the absorption of vitamin B12–intrinsic factor
complex, as may surgery on that part of the bowel.

Folate absorption
Folates are produced by plants and bacteria; hence dietary leafy
vegetables (spinach, broccoli, lettuce), fruits (bananas, melons)
and animal protein (liver, kidney) are a rich source. An average
Western diet contains more than the minimum daily intake of
50 μg but excess cooking destroys folates. Most dietary folate is
present as polyglutamates; these are converted to monoglutamate
in the upper small bowel and actively transported into plasma.
Plasma folate is loosely bound to plasma proteins such as albumin
and there is an enterohepatic circulation. Total body stores of
folate are small and deficiency can occur in a matter of weeks.
Folate deficiency
The causes and diagnostic features of folate deficiency are given below. The edentulous elderly or psychiatric
patient is particularly susceptible to dietary deficiency and this
is exacerbated in the presence of gut disease or malignancy.
Pregnancy-induced folate deficiency is the most common cause
of megaloblastosis worldwide and is more likely in the context
of twin pregnancies, multiparity and hyperemesis gravidarum.
Serum folate measurement is very sensitive to dietary intake;
a single folate-rich meal can normalise it in a patient with true
folate deficiency, whereas anorexia, alcohol and anticonvulsant
therapy can reduce it in the absence of megaloblastosis. For
this reason, red cell folate levels are a more accurate indicator
of folate stores and tissue folate deficiency.

Causes of folate deficiency

Diet
• Poor intake of vegetables
Malabsorption
• e.g. Coeliac disease, small bowel surgery
Increased demand
• Cell proliferation, e.g. haemolysis
• Pregnancy
Drugs*
• Certain anticonvulsants (e.g. phenytoin)
• Contraceptive pill
• Certain cytotoxic drugs (e.g. methotrexate)

Investigation of folic acid deficiency

Diagnostic findings
• Serum folate levels may be low but are difficult to interpret
• Low red cell folate levels indicate prolonged folate deficiency and
are probably the most relevant measure
Corroborative findings
• Macrocytic dysplastic blood picture
• Megaloblastic marrow

Management of megaloblastic anaemia

If a patient with a severe megaloblastic anaemia is very ill and
treatment must be started before vitamin B12 and red cell folate results are available, that treatment should always include both
folic acid and vitamin B12. The use of folic acid alone in the
presence of vitamin B12 deficiency may result in worsening of
neurological features.
Rarely, if severe angina or heart failure is present, transfusion
can be used in megaloblastic anaemia. The cardiovascular system
is adapted to the chronic anaemia present in megaloblastosis,
and the volume load imposed by transfusion may result in
decompensation and severe cardiac failure. In such circumstances,
exchange transfusion or slow administration of 1 U of red cells
with diuretic cover may be given.

Vitamin B12 deficiency

Vitamin B12 deficiency is treated with hydroxycobalamin. In
cases of uncomplicated deficiency, 1000 μg IM for 6 doses 2
or 3 days apart, followed by maintenance therapy of 1000 μg
every 3 months for life, is recommended. In the presence of
neurological involvement, a dose of 1000 μg on alternate days
until there is no further improvement, followed by maintenance
as above, is recommended. The reticulocyte count will peak
by the 5th–10th day after starting replacement therapy. The
haemoglobin will rise by 10 g/L every week until normalised.
The response of the marrow is associated with a fall in plasma
potassium levels and rapid depletion of iron stores. If an initial
response is not maintained and the blood film is dimorphic (i.e.
shows a mixture of microcytic and macrocytic cells), the patient
may need additional iron therapy. A sensory neuropathy may
take 6–12 months to correct; long-standing neurological damage
may not improve.

Folate deficiency

Oral folic acid (5 mg daily for 3 weeks) will treat acute deficiency
and 5 mg once weekly is adequate maintenance therapy.
Prophylactic folic acid in pregnancy prevents megaloblastosis
in women at risk, and reduces the risk of fetal neural tube
defects. Prophylactic supplementation is also given in
chronic haematological disease associated with reduced red cell
lifespan (e.g. haemolytic anaemias). There is some evidence that
supraphysiological supplementation (400 μg/day) can reduce
the risk of coronary and cerebrovascular disease by lowering
plasma homocysteine levels. This has led the US Food and
Drug Administration to introduce fortification of bread, flour and
rice with folic acid.