Acquired and inherited disorders of cobalamin and folate in children

Cobalamin deficiency in the newborn usually results from cobalamin deficiency in the mother. Megaloblastic anaemia, pancytopenia and failure to thrive can be present, accompanied by neurological deficits if the diagnosis is delayed. Most cases of spina bifida and other neural tube defects result from maternal folate and/or cobalamin insufficiency in the periconceptual period. Polymorphisms in a number of genes involved in folate and cobalamin metabolism exacerbate the risk. Inborn errors of cobalamin metabolism affect its absorption, (intrinsic factor deficiency, Imerslund‐Gräsbeck syndrome) and transport (transcobalamin deficiency) as well as its intracellular metabolism affecting adenosylcobalamin synthesis (cblA and cblB), methionine synthase function (cblE and cblG) or both (cblC, cblD and cblF). Inborn errors of folate metabolism include congenital folate malabsorption, severe methylenetetrahydrofolate reductase deficiency and formiminotransferase deficiency. The identification of disease‐causing mutations in specific genes has improved our ability to diagnose many of these conditions, both before and after birth.     

Genomic mutations associated with mild and severe deficiencies of transcobalamin I (haptocorrin) that cause mildly and severely low serum cobalamin levels

Transcobalamin (TC) I deficiency, like the function of TC I itself, is incompletely understood. It produces low serum cobalamin levels indistinguishable from those of true cobalamin deficiency. Diagnosis is especially elusive when TC I deficiency is mild. To provide new, more substantive definition, the TCN1 gene was examined in two well-characterised families that included members with both severe and mild TC I deficiencies. A severely deficient proposita with undetectable TC I levels displayed compound heterozygosity for two mutations, each causing a premature stop codon. Relatives in both families who had mildly low or low-normal plasma levels of TC I and cobalamin were heterozygous for one or the other of these mutations. An unrelated patient with mild TC I deficiency and unknown familial TC I and cobalamin status was then tested and found to be similarly heterozygous for one of the mutations. The two nonprivate mutations identify a genetic basis for TC I deficiency for the first time. They also add new approaches to studying mild and severe TC I deficiency and to reducing confusion of its low cobalamin levels with those of cobalamin deficiency and its often dramatically different prognosis and management.     

Serum cobalamin, folate, methylmalonic acid and total homocysteine as vitamin B12 and folate tissue deficiency markers amongst elderly Swedes--a population-based study

OBJECTIVES: The possibilities of detecting tissue cobalamin and folate deficiency are under debate. In this report the levels of serum cobalamin, folate, methylmalonic acid (MMA) and total homocysteine (tHcy) and their interrelations in a representative random population sample are presented. DESIGN: Cohort study. SETTING: A general mid-Swedish population. SUBJECTS: A 20% random sample of persons 70 years or older in a defined geographical area were invited to a survey. A total of 235 (85%) persons responded, out of whom 224 had no interfering diseases. MAIN OUTCOME MEASURES: Serum cobalamin, folate, MMA and tHcy. RESULTS: The serum levels of cobalamin, folate, MMA and tHcy were all correlated to cobalamin and folic acid treatment. They were also correlated to the intake of multivitamin preparations. In addition, serum cobalamin was higher in untreated women than in men but not correlated to age. Serum folate was correlated neither to sex nor age. Serum tHcy and MMA were both directly correlated to age but MMA not to sex. MMA was inversely correlated to serum cobalamin but not to serum folate, whereas serum tHcy was inversely correlated to serum cobalamin, folate and creatinine. Neither serum cobalamin, folate, MMA nor tHcy had any significant correlation to haemoglobin, erythrocyte volume fraction (EVF) or mean red cell volume. Half of the study population had abnormal MMA or tHcy levels, suggesting a latent or overt tissue deficiency of cobalamin or folate. CONCLUSIONS: A substantial proportion of the elderly general population had signs of low tissue levels of cobalamin or folate. Amongst those who took multivitamin preparations this proportion was much lower.     

Efficacy of short‐term oral cobalamin therapy for the treatment of cobalamin deficiencies related to food‐cobalamin malabsorption: A study of 30 patients

Background: It has been suggested that oral cobalamin (vitamin B₁₂) therapy may be an effective therapy for treating cobalamin deficiencies related to food‐cobalamin malabsorption. However, the duration of this treatment was not determined. Patients and method: In an open‐label, nonplacebo study, we studied 30 patients with established cobalamin deficiency related to food‐cobalamin malabsorption, who received between 250 and 1000 μg of oral crystalline cyanocobalamin per day for at least 1 month. Endpoints: Blood counts, serum cobalamin and homocysteine levels were determined at baseline and during the first month of treatment. Results: During the first month of treatment, 87% of the patients normalized their serum cobalamin levels; 100% increased their serum cobalamin levels (mean increase, +167 pg/dl; P < 0.001 compared with baseline); 100% had evidence of medullary regeneration; 100% corrected their initial macrocytosis; and 54% corrected their anemia. All patients had increased hemoglobin levels (mean increase, +0.6 g/dl) and reticulocyte counts (mean increase, +35 × 10⁶/l) and decreased erythrocyte cell volume (mean decrease, 3 fl; all P < 0.05). Conclusion: Our findings suggest that crystalline cyanocobalamin, 250–1000 μg /day, given orally for 1 month, may be an effective treatment for cobalamin deficiencies not related to pernicious anemia.     

Whole blood folate values in pernicious anaemia: relation to treatment

27 patients with pernicious anaemia, followed for a long period, were consecutively treated with three different vitamin B12 preparations, while during intervening period no therapy was given until signs of B12 deficiency developed. After vitamin B12 treatment, a peak whole blood folate value some 70% higher than the starting value was noted after about 75 d in patients treated with B12 injections and after about 165 d in perorally treated patients. During the same time significantly increased whole blood folate/plasma folate ratio was noted. In most of the patients, whole blood folate declined from peak to plateau level in about 150 d and thereafter remained almost unchanged until signs of cobalamin deficiency developed. The mean time from peak to plateau values was significantly longer in "long-reactors" than in "short-reactors". Higher peak and plateau values were noted in patients with neuropathy and mucosal symptoms, as compared to patients with hypersegmentation.     

Mild hyperhomocysteinemia in adult patients with sickle cell disease: a common finding unrelated to folate and cobalamin status

Homocysteine has associations with both vitamin insufficiency and vascular complications, and its status is therefore of interest in sickle cell disease (SCD). However, information is limited, especially in adults. We studied plasma total homocysteine (tHcy) and three of its major modifiers, cobalamin, folate, and creatinine, in 90 adult patients with SCD and 76 control subjects. The patients had higher tHcy levels than did controls (P = 0.03) and had elevated tHcy more often (20% vs. 3%, P = 0.0005). None of the hyperhomocysteinemic patients had low cobalamin or folate levels; on the contrary, patients with SCD had high folate levels more often than control subjects (32% vs. 7%; P < 0.0001). Although serum creatinine values were lower in SCD patients than in control subjects (P = 0.03), high levels also tended to occur more often (8% vs. 1%; P = 0.054). Most importantly, creatinine levels correlated significantly with tHcy (P < 0.0001) and logistic regression analyses showed creatinine to be the only significant predictor of high tHcy levels in SCD (P = 0.01). Our results show that hyperhomocysteinemia affects 20% of adults with SCD despite routine folate supplementation and is independent of folate and cobalamin status. Creatinine was the major identifiable influence on tHcy, but renal insufficiency explained only 4 of the 18 elevated tHcy levels. Longitudinal studies will be needed to determine whether the frequent hyperhomocysteinemia of SCD influences the vascular complications in SCD. If reducing tHcy becomes advisable, then interventions other than folate therapy will be needed.     

Subtle and atypical cobalamin deficiency states

Evidence for cobalamin (vitamin B₁₂) deficiency usually involves some combination of low serum cobalamin levels, clinical abnormalities (classically, megaloblastic anemia and neurologic defects), metabolic abnormalities, and response to therapy. However, cobalamin deficiency may often display few of the expected clinical findings. Identification of the underlying cause is also important in the diagnosis of deficiency, and its value may be particularly great when the expression of deficiency is subtle. The cause of cobalamin deficiency is usually malabsorptive, but may sometimes be limited to malabsorption of food cobalamin while free cobalamin is absorbed normally. Nongastroenterologic entities may sometimes also be found. All of these considerations allow the proposal of four patterns of cobalamin deficiency. The first type is classical deficiency; typical megaloblastic anemia with or without neurologic dysfunction occurs because of classical cobalamin malabsorption such as lack of intrinsic factor (pernicious anemia). The second type consists of classical cobalamin malabsorption in which the cobalamin deficiency is expressed subtly rather than in classical fashion. There is no megaloblastic anemia and sometimes the only evidence of deficiency may be metabolic. In the third type, cobalamin deficiency is expressed classically but is attributable to a subtle or atypical cause, such as food-cobalamin malabsorption. In the fourth type, deficiency is both expressed subtly and arises from subtle or atypical causes. Such presentations require further investigation but are a challenging expansion of our understanding and recognition of cobalamin deficiency.     

Cobalamin absorption and serum homocysteine and methylmalonic acid in elderly subjects with low serum cobalamin

Abstract: We prospectively studied 41 consecutive elderly patients with serum cobalamin (vitamin B₁₂) levels lower than 125 pmol/l. The protein-bound cobalamin absorption test (PBAT) was performed in 34 of them and in 27 selected elderly control patients. The lower decision limit was 0.18% and an abnormal test was detected in only 9 (26%) of the 34 patients with low serum cobalamin level. When the PBAT was compared to the Schilling (Dicopac method) test, a concordant result was found in 80 %. Serum methylmalonic acid and/or total homocysteine concentrations were elevated in 75% (26/35) of the patients with low serum cobalamin levels but also in 30% (5/17) of the control patients. Of the 12 and 9 cobalamin-deficient patients with elevated serum levels of methylmalonic acid and homocysteine, normalization after cobalamin therapy was obtained in 11 and 5 respectively. In conclusion, determination of serum metabolites and their response to cobalamin therapy are a sensitive index of significant cobalamin deficiency and a useful means of distinguishing between cobalamin and folate deficiency. The PBAT offers little advantage over the Schilling test in diagnosing cobalamin malabsorption in elderly patients.     

Reported symptoms and clinical findings in relation to serum cobalamin,folate, methylmalonic acid and total homocysteine among elderly Swedes: a population-based study

OBJECTIVES: The early stages of tissue B12 or folate deficiency often cause diagnostic problems. In this report, the levels of serum cobalamin, folate, methylmalonic acid (MMA) and total homocysteine (tHcy), and their relationships with clinical findings and reported symptoms in a representative random population sample are presented. DESIGN: Cohort study. SETTING: A general central Swedish population 70 years or older. SUBJECTS AND METHODS: A 20% random sample of persons 70 years or older in a defined geographical area were invited to a survey. A total of 235 (85%) persons responded, of whom 161 had no interfering diseases or medication. Blood specimens for serum cobalamin, folate, MMA and tHcy analyses were drawn. MAIN OUTCOME MEASURES: Presence of anaemic, gastrointestinal, neurological and psychiatric symptoms, obtained by questionnaire, and vibration sense measurement and findings at a physical and Mini Mental State Examination. RESULTS: Among a large number of symptoms and clinical findings that traditionally have been linked to vitamin B12 or folate tissue deficiency, only changes in the tongue mucosa and mouth angle stomatitis turned out to be significantly associated with abnormal serum tHcy and serum folate levels. There were no relationships to serum cobalamin and serum MMA. CONCLUSIONS: Changes in the oral mucosa were the only signs and symptoms found in this study, indicating that these may be the very early markers of metabolic defects. The traditional symptoms of vitamin deficiency may appear later in the course.     

Hyperhomocysteinemia and cobalamin deficiency in young Asian Indians in the United States

Hyperhomocysteinemia, a risk factor for vascular disease, may be a particular problem in Asian Indians, but information is limited, especially in the U.S., despite its growing Asian population. Moreover, suggestions have been made that folate deficiency is responsible for the hyperhomocysteinemia in Indians. Therefore, we studied homocysteine status in healthy Asian Indians in the U.S. prospectively, determined the frequency of cobalamin and folate deficiency as contributors to it, and examined whether food-cobalamin absorption contributed to cobalamin deficiency. Homocysteine levels were higher in Asian Indian men than in 4 other ethnic groups (P < 0.0001); 10/39 Indian men (25.6%) were hyperhomocysteinemic. Cobalamin levels were lower in Indian men (P = 0.000005) and women (P = 0.03) than in non-Indians; low levels were found more frequently in both Indian men (23/39; 59.0%) and women (5/21; 23.8%) than in others. Measuring methylmalonic acid in 10 selected subjects showed that the low cobalamin levels reflected cobalamin deficiency, and high methylmalonic acid levels were found in some subjects without hyperhomocysteinemia. Evidence of folate deficiency was not found in any subjects. Food-cobalamin absorption was normal in all 13 Indian subjects tested, including those with Helicobacter pylori infection. The results show that hyperhomocysteinemia is strikingly common in apparently healthy, young Asian Indian men. The cause appears to be cobalamin deficiency, which affected more than half of the Indian men, may be largely subclinical, is underestimated by homocysteine levels alone which were not always abnormal, and is probably largely dietary in origin. Folate deficiency is rare. This public health problem is amenable to prevention and treatment in this growing segment of the U.S. population. It was, parenthetically, noteworthy that many of the affected subjects were young physician trainees.     

Human cobalamin deficiency: alterations in serum tumour necrosis factor-α and epidermal growth factor

OBJECTIVES: We have previously demonstrated that vitamin B12 (cobalamin)-deficient central neuropathy in the rat is associated with local overexpression of neurotoxic tumour necrosis factor (TNF)-alpha combined with locally decreased synthesis of neurotrophic epidermal growth factor (EGF). The aims of this study were to investigate whether a similar imbalance also occurs in the serum of adult patients with clinically confirmed cobalamin deficiency and whether it can be corrected by vitamin B12 replacement therapy. PATIENTS AND METHODS: We studied 34 adult patients with severe cobalamin deficiency, 12 patients with pure iron deficiency anaemia and 34 control subjects. Haematological markers of cobalamin deficiency and serum TNF-alpha and EGF levels were measured using commercial kits. Thirteen cobalamin-deficient patients were re-evaluated after 3 and 6 months of parenteral vitamin B12 treatment. RESULTS: TNF-alpha was significantly higher (p < 0.01) and EGF significantly lower (p < 0.01) in the patients with cobalamin deficiency, but both were unchanged in patients with pure iron deficiency anaemia. In cobalamin-deficient patients the serum TNF-alpha levels correlated significantly with plasma total homocysteine levels (r = 0.425; p < 0.02). In the treated patients TNF-alpha and EGF levels normalised concomitantly with clinical and haematological disease remission. CONCLUSIONS: In humans, as in rats, cobalamin concentration appears to be correlated with the synthesis and release of TNF-alpha and EGF in a reciprocal manner, because cobalamin deficiency is accompanied by overproduction of TNF-alpha and underproduction of EGF.     

Spouses of demented patients with low cobalamin levels: a new risk group for cobalamin deficiency

Abstract: Low serum cobalamin levels are common in conditions such as dementia and often represent mild deficiency. We surveyed serum cobalamin levels prospectively in spouses and blood relatives of demented patients to determine if any familial predisposition exists for the low levels. Cobalamin status in most of the relatives found to have low levels was assessed further by means of blood counts, metabolic tests, neurologic evaluation, absorption studies and response to cobalamin therapy. Serum cobalamin levels in 36 spouses correlated with those of the 36 demented patients related to them (r = 0.46, p = 0.004). A significant association was not seen in 34 blood relatives of 34 demented patients (r = 0.27). Most importantly, 67% of the spouses of demented patients with low serum cobalamin had low values themselves, compared with only 3% of the spouses of patients with normal levels (p = 0.001). Detailed study of 4 of the 5 spouses (and 3 blood relatives) with low cobalamin levels showed no anemia in any case. Nevertheless, 4 of the subjects had metabolic evidence of deficiency and one had electrophysiological abnormalities; all these defects improved with cobalamin therapy. These observations identify a hitherto unsuspected group of people at high risk for cobalamin deficiency and suggest that spouses of demented patients with low cobalamin levels should also have their cobalamin levels measured. The increased frequency of low serum cobalamin levels in spouses of demented patients with low levels represents in most cases a true, mild cobalamin deficiency that responds to treatment.     

Serum transferrin receptor in the megaloblastic anemia of cobalamin deficiency.

In order to further study the relation between transferrin receptor and erythropoiesis we examined serum receptor levels in megaloblastic anemia, which is the classic example of ineffective erythropoiesis. We studied 33 patients with unequivocal cobalamin deficiency, only 22 of whom were anemic. High serum transferrin receptor levels were found in 12 patients, all of whom were anemic and had high lactate dehydrogenase (LDH) levels; in contrast, only 10 of the 21 patients with normal receptor levels were anemic. Receptor correlated most strongly with LDH (r = 0.573, p < 0.001) and, inversely, with hemoglobin values (r = -0.560, p < 0.001); it also correlated with ferritin and total bilirubin levels, but not with cobalamin, MCV or erythropoietin. No association was found with the hemolytic component of megaloblastic anemia, represented indirectly by haptoglobin levels. Changes induced by cobalamin therapy were also examined in 13 patients. Transferrin receptors rose in all 6 patients who initially had high levels and in 2 of 3 patients who had borderline levels, but not in the 4 patients with initially normal levels. The receptor levels began to rise within 1-3 days, peaked at about 2 weeks and returned to normal at about the 5th wk. The findings indicate that serum transferrin receptor levels reflect the severity of the megaloblastic anemia. The elevated receptor levels rise further with cobalamin therapy, however, as effective erythropoiesis replaces ineffective erythropoiesis, and these persist until the increased erythropoiesis returns to normal.     

Prospective evaluation of protein bound vitamin B12 (cobalamin) malabsorption in the elderly using trout flesh labelled in vivo with 57Co-cobalamin

Background—The frequency of dietary proteinbound vitamin B₁₂ malabsorption in elderly patients remains controversial.
Aims—To evaluate this malabsorption inelderly hospitalised patients using a modified Schilling test.
Patients—Fourteen elderly patients with lowB₁₂ blood levels were prospectively selected from 394 hospitalised patients.
Methods—The modified Schilling test was performedwith trout labelled in vivo.
Results—The test was normal in five healthyelderly subjects, in 7/8 patients with pancreatic insufficiency, and innine non-elderly patients with antral gastritis. The low decision limitwas established at 3.3% (median 4.8%). From the 14 elderly patientswith low B₁₂ prospectively selected from 394 hospitalisedpatients, seven had a real deficiency with anaemia and an increasedhomocysteine and/or methylmalonate serum level. The modified Schillingtest showed malabsorption in five of these patients, including two inwhich the standard Schilling test was normal, and three in which the standard Schilling test was partially corrected by an intrinsic factor.
Conclusions—Protein bound vitamin B₁₂malabsorption was detected in at least 0.5% of elderly hospitalisedpatients, using the labelled trout flesh absorption test.

Keywords:cobalamin deficiency; malabsorption; gastritis; Schilling test

     

Severe megaloblastic anaemia presenting as pancytopenia with red cell hypoplasia and elevated serum cobalamin and cobalamin binding proteins

In severe megaloblastic anaemia due to folate or cobalamin deficiency the bone marrow is usually hypercellular with megaloblastic change in erythropoiesis and giant metamyelocytes in the myeloid series. We report an unusual case of megaloblastic anaemia with marked erythroid hypoplasia and myeloid hyperplasia; the serum cobalamin and unsaturated cobalamin binding capacity were both grossly elevated.     

High serum folates and the simplification of red cell folate analysis

Recent substantial increases in clinical blood folate concentrations are noted. Since red cell folates (RCF) are calculated from whole blood folates (WBF) by subtraction of the endogenous serum folate (SF) component, the reporting of clinical RCF results may be delayed because an ever increasing proportion (15%) of diagnostic SF levels are high (> 20 ng/ml) and need a repeat analysis. We evaluated ‘plasma replacement’ as a simple preanalytical procedure in which endogenous blood plasma is removed from red cells by washing and substituted with ‘low‐folate’ plasma (serum) as an alternative conjugase (gamma‐glutamyl carboxypeptidase) source for folate polyglutamate hydrolysis. Washed and conventional RCF assays compared well after both manual (n=115, r=0.98, y=1x + 1.26) and automated washing of red cells (n=170, r=0.96, y=0.96x – 0.73 ng/ml) and were not significantly different. The interassay reproducibility of folate results from washed blood samples was good (CV=< 6%). This novel ‘plasma replacement’ step halves the cost of a valid RCF assay by eliminating the need for endogenous SF analysis, and it expedites the reporting of clinical results.     

A patient with the inability to maintain in vivo levels of bound cobalamin (Cbl) and manifestations of tissue deficiency of Cbl

A 39-year-old woman presented with mild anemia, glossitis, an increased MCV, a low serum cobalamin (Cbl) (vitamin B12), mild tissue deficiency of Cbl, but with neither malabsorption of Cbl, impaired intake, nor deficiency of or inactivity of transcobalamin II (TC II). Because of a persistently low holo-TC II (TC II carrying Cbl as the circulating complex of TC II-Cbl), much of the evaluation was focused on the patient's TC II. Her TC II promoted the uptake of Cbl, reacted with anti-TC II, and bound Cbl in vitro. A test dose of 200 micrograms of cyanocobalamin (CN-Cbl) i.m. increased her holo TC II to levels higher than those in healthy persons, but with a much more abrupt fall to a subnormal level. Two milligrams of CN-Cbl i.m. followed by 100 micrograms i.m. monthly failed to maintain normal amounts of circulating TC II-Cbl or to overcome the tissue deficiency of Cbl. One milligram i.m. weekly or daily p.o. corrected both. The low holo TC II was considered to be responsible for the clinical expression and may have been primary to the reduced amounts of total and holo R binder of Cbl in the circulation. This study of a newly recognized defect points out the need for circulating holo TC II, a rational use of pharmacologic amounts of Cbl, and a possible interrelationship between TC II and the R binder of Cbl.