Functional methionine synthase deficiency due to cblG disorder: A report of two patients and a review

Functional methionine synthase deficiency due to abnormal methylcobalamin metabolism causes megaloblastic anemia, moderate to severe developmental delay, lethargy, and anorexia in association with homocystinuria. Patients with this disorder of cobalamin metabolism can be classified into two separate groups, cblE or cblG, primarily on the basis of complementation analysis with cultured skin fibroblasts. We describe two unrelated boys, ages 3 and 5 years, with the cblG defect in methylcobalamin synthesis. Both children presented with severe developmental delay, lethargy, anorexia, and megaloblastic anemia. The diagnosis of homocystinuria was delayed in each case due to difficulties with detection of small amounts of homocystine in physiologic samples. The clinical course of cblG disease is favorably altered by treatment with intramuscular hydroxycobalamin. Megaloblastosis in the presence of adequate supplies of cobalamin and folate in the blood must alert the clinician to the possibility of functional methionine synthase deficiency and should prompt a careful search for associated biochemical hallmarks, including homocystinuria/emia. Am. J. Med. Genet. 71:384–390, 1997. © 1997 Wiley-Liss, Inc.     

Experimental evidence of oxidative stress in plasma of homocystinuric patients: a possible role for homocysteine

Homocystinuria is an inherited disorder biochemically characterized by high urinary excretion of homocystine and increased levels of homocysteine (Hcy) and methionine in biological fluids. Affected patients usually have a variety of clinical and pathologic manifestations. Previous experimental data have shown a relationship between Hcy and oxidative stress, although very little was reported on this process in patients with homocystinuria. Therefore, in the present study we evaluated parameters of oxidative stress, namely carbonyl formation, malondialdehyde (MDA) levels, sulfhydryl content and total antioxidant status (TAS) in patients with homocystinuria at diagnosis and under treatment with a protein restricted diet supplemented by pyridoxine, folate, betaine, and vitamin B(12). We also correlated plasma Hcy and methionine concentrations with the oxidative stress parameters examined. We found a significant increase of MDA levels and carbonyl formation, as well as a reduction of sulfhydryl groups and TAS in plasma of homocystinuric patients at diagnosis relatively to healthy individuals (controls). We also verified that Hcy levels were negatively correlated with sulfhydryl content and positively with MDA levels. Furthermore, patients under treatment presented a significant reduction of the content of MDA, Hcy and methionine concentrations relatively to patients at diagnosis. Taken together, the present data indicate that lipid and protein oxidative damages are increased and the antioxidant defenses diminished in plasma of homocystinuric patients, probably due to increased reactive species elicited by Hcy. It is therefore presumed that oxidative stress participates at least in part in the pathogenesis of homocystinuria.     

Homocystinuria and megaloblastic anemia responsive to vitamin B12 therapy. An inborn error of metabolism due to a defect in cobalamin metabolism

We describe an inborn error of vitamin B12 metabolism in an infant who had severe developmental delay, megaloblastic anemia, and homocystinuria. There was no evidence of methylmalonic aciduria or deficiency of folate or vitamin B12. Treatment with hydroxocobalamin, but not with cyanocobalamin and folic acid, resulted in rapid clinical and biochemical improvement. Cultured fibroblasts showed an absolute growth requirement for methionine, defective incorporation of radioactivity from [14C]5-methyltetrahydrofolate into protein, and normal incorporation of radioactivity from [14C]propionate, thus assigning the intracellular defect to methionine synthesis. The proportion of intracellular methylcobalamin in the fibroblasts was decreased, but that of 5'-deoxyadenosylcobalamin was normal. Methionine synthetase activity in cell extracts was normal, as was cobalamin incorporation into cultured cells. This defect differs from those described previously in being limited to methylcobalamin accumulation and defective use of 5-methyltetrahydrofolate by intact cells with normal activity of methylmalonyl CoA mutase.     

Is disordered folate metabolism the basis for the genetic predisposition to neural tube defects?

Vitamin levels were measured in twenty women under 35 years of age with a history of two or more neural tube defect pregnancies. Each index case was compared with a female control matched for age, obstetric history and social class. The mean concentration of red cell folate in the subjects was 178 ng/ml, significantly lower than the mean of 268 ng/ml for the control group (P = 0.005). Red cell folate levels showed a linear relationship with the number of neural tube defect pregnancies, the levels being lowest in women who had had three or four affected offspring. There was no significant difference in serum folate; plasma or white cell vitamin C; plasma vitamin A; thiamine, riboflavine or pyridoxine status; serum vitamin B12; plasma vitamin E; total protein, albumin, transferrin, magnesium, copper or zinc. Diet was assessed by a questionnaire. The dietary intakes of total folate and other vitamins except vitamin A were lower in the subjects than the controls but none of the differences were statistically significant. Regression analysis showed a difference between subjects and controls in the relationship of red cell folate to dietary folate. This study demonstrates an association between susceptibility to offspring with neural tube defects and depressed red cell folate levels which cannot be entirely attributed to a lower dietary intake of folate. It is postulated that one factor predisposing to the occurrence of neural tube defects may be an inherited disorder of folate metabolism.     

Haemoglobin A2 levels in vitamin B12 and folate deficiency.

Haemoglobin A2 levels were measured in 50 patients with vitamin B12 deficiency, 50 patients with folate deficiency, and six patients with combined deficiencies of these vitamins. All were normal except for three patients with vitamin B12 deficiency, who had a slightly elevated Hb A2 level; this fell to normal after vitamin B12 therapy. It is concluded that haemoglobin A2 levels are usually normal in vitamin B12 or folate deficiency. However, raised levels of haemoglobin A2 may be found, but these are not as high as is found in beta thalassaemia trait and should not cause difficutly in diagnosis.     

Serum levels of folate and cobalamin in women with recurrent spontaneous abortion

We evaluated the folate and cobalamin status in 29 non-pregnant women with a history of recurrent spontaneous abortion (three or more consecutive) of unknown aetiology in comparison to 29 healthy nulligravidae of similar reproductive age (controls). Serum concentrations of folate and cobalamin showed no significant differences between the two groups. No correlation between age and vitamin concentrations was found. In the study group there was a significant negative correlation of the number of previous abortions and the concentration of serum folate. Patients with at least four previous miscarriages had significantly lower serum values of folic acid than women with three abortions, but not than controls. The underlying cause of this finding remains unclear. In conclusion, the serum concentrations of folic acid and vitamin B12 are not significantly altered in women with unexplained recurrent spontaneous abortions, and an association between a deficiency of these vitamins and an increased risk of pregnancy loss appears to be questionable in the majority of gestations.      

A Common Variant in Methionine Synthase Reductase Combined with Low Cobalamin (Vitamin B12) Increases Risk for Spina Bifida

Impairment of folate and cobalamin (vitamin B(12)) metabolism has been observed in families with neural tube defects (NTDs). Genetic variants of enzymes in the homocysteine remethylation pathway might act as predisposing factors contributing to NTD risk. The first polymorphism linked to increased NTD risk was the 677C-->T mutation in methylenetetrahydrofolate reductase (MTHFR). We now report a polymorphism in methionine synthase reductase (MTRR), the enzyme that activates cobalamin-dependent methionine synthase. This polymorphorism, 66A-->G (I22M), has an allele frequency of 0.51 and increases NTD risk when cobalamin status is low or when the MTHFR mutant genotype is present. Genotypes and cobalamin status were assessed in 56 patients with spina bifida, 58 mothers of patients, 97 control children, and 89 mothers of controls. Cases and case mothers were almost twice as likely to possess the homozygous mutant genotype when compared to controls, but this difference was not statistically significant. However, when combined with low levels of cobalamin, the risk for mothers increased nearly five times (odds ratio (OR) = 4.8, 95% CI 1.5-15.8); the OR for children with this combination was 2.5 (95% CI 0.63-9.7). In the presence of combined MTHFR and MTRR homozygous mutant genotypes, children and mothers had a fourfold and threefold increase in risk, respectively (OR = 4.1, 95% CI 1.0-16.4; and OR = 2.9, 95% CI 0.58-14.8). This study provides the first genetic link between vitamin B(12) deficiency and NTDs and supports the multifactorial origins of these common birth defects. Investigation of this polymorphism in other disorders associated with altered homocysteine metabolism, such as vascular disease, is clearly warranted.     

Major Determinants of Serum Homocysteine Concentrations in a Korean Population

The objective of this study was to identify the factors that determine serum homocysteine concentrations in Korean population. In a community-based study, 871 participants completed detailed questionnaires and physical examination. We found that increased age, male sex, family history of stroke, deficiencies of serum folate and vitamin B12, and elevated serum creatinine significantly increased the risk of hyperhomocysteinemia. However, hormonal and behavioral factors (smoking, alcohol drinking, coffee consumption, and sedentary time) were not associated with the risk of hyperhomocysteinemia. The risk of hyperhomocysteinemia was steeply increased in subjects with two or more risk factors among four selected risk factors (deficiencies of serum folate and vitamin B12, elevated creatinine, and family history of stroke) compared to subjects who did not have any risk factors, especially subjects over the age of 65 yr (odds ratio [OR], 33.5; 95% confidence interval [CI], 3.71-302.0 in men; OR, 39.2; 95% CI, 7.95-193.2 in women). In conclusion, increased age, male sex, family history of stroke, deficiencies of serum folate and vitamin B12, and elevated serum creatinine are important determinants of serum homocysteine concentrations with interaction effects between these factors.     

Vitamin replacement therapy in renal failure patients

Renal failure patients require vitamin replacement therapy that addresses the specialized needs of renal failure. Four factors including restricted diet, uremic toxins, drug-nutrient interactions, and in ESRD, the dialysis process, affect the normal absorption, retention and activity of necessary micronutrients which support all aspects of carbohydrate, protein, lipid and nucleic acid metabolism. Studies have shown that the typical renal failure diet is low in B vitamins, that uremic factors affect folate and pyridoxine activities and that many B vitamins are lost on dialysis at a rate greater than are lost with normal urinary excretion. In addition, retention of vitamin A or inappropriately high supplementation of vitamin C may cause toxicities which exacerbate existing pathologies. Further, emerging research suggests some vitamins such as folic acid and pyridoxine, if provided in higher than normal amounts, may have an impact on reducing the risk of some aspects of renal cardiovascular disease. It is therefore important to supplement some vitamins, and use restraint in the supplementation of others. It is clear that renal failure patients, including predialysis, ESRD and transplant patients need specialized supplementation that meets the requirements of disease and its management.     

A congenital anomaly of vitamin B12 metabolism: a study of three cases.

The clinical and morphologic findings of three patients with metabolic acidosis, methylmalonic aciduria, and homocystinuria are presented. The clinical evolution of the patients was similar and was characterized in the first weeks of life by failure to thrive, hypotonia, and lethargy associated with pancytopenia and hepatic dysfunction, eventually progressing to severe respiratory insufficiency and renal failure consistent with a hemolytic-uremic syndrome. The patients died at 40, 45, and 75 days of age. Biochemical analyses and complementation studies revealed a congenital anomaly of vitamin B12 metabolism (cobalamin C disease). Postmortem morphologic findings in all three cases were dominated by a thrombotic microangiopathy of the kidneys and lungs, diffuse hepatic steatosis, and megaloblastic changes in the bone marrow. A severe gastritis with striking cystic dysplastic mucosal changes and total absence of parietal and chief cells was a consistent finding in all three cases, the rest of the gastrointestinal tract appearing essentially normal. Cobalamin C disease is an intracellular defect of cobalamin metabolism with possible recessive inheritance that can result in multiorgan failure early in life, with a thrombotic microangiopathy and unusual changes in the gastric mucosa.     

Alzheimer's disease: a 'cobalaminergic' hypothesis.

An association between Alzheimer's Disease (AD) and low CSF and serum vitamin B12 (B12) has recently been described (1, 2, 3). This is apparently independent of nutritional intake (4). It has been suggested that such patients may exhibit an atypical form of cobalamin deficiency (3, 4). It is therefore proposed that these deficiencies may be aetiologically important, at least in sub-groups of AD, and a mechanism is described whereby B12 deficiency may result in the characteristic neurotransmitter changes of the disease. The hypothesis generates predictions regarding biochemical evaluation of such patients and suggests associations between the neurochemical disturbances and structural abnormalities of AD.     

Aging, chronic administration of ethanol, and acute exposure to nitrous oxide: effects on vitamin B12 and folate status in rats.

Elderly patients with alcoholism often require surgery and receive nitrous oxide (N2O) as a component of their anesthetic. Since aging, ethanol, and N2O may all perturb folate and/or vitamin B12 metabolism, we examined the combined influence of these parameters on vitamin B12/folate status in a rodent model. Aged male Fischer 344 rats (24 months old) were given a liquid ethanol diet (35% of calories as ethanol) and control rats were pair-fed a liquid diet with carbohydrate substituting for the caloric content of ethanol. After receiving liquid diets for 7 weeks, rats were exposed to 60% N2O/40% 0(2) for 6 h. Urinary excretion of formic acid, formiminoglutamic acid (FIGLU), and methylmalonic acid (MMA) were used as indirect markers of folate/vitamin B12 status. In both the aged ethanol-fed and control groups, excretion of formic acid and FIGLU markedly increased the first day after N2O exposure and returned towards background values by the second day. No changes occurred in MMA excretion. Exposure to N2O decreased methionine synthase activities in liver, kidney and brain, and recovery of methionine synthase activities occurred over a period of 4 days in both the aged ethanol-fed and control groups. Ethanol treatment for 7 weeks combined with acute exposure to N2O did not deplete the aged rats of folate or vitamin B12 in blood, liver, kidney or brain. Thus, in this animal model, aging, chronic ethanol administration, and acute N2O exposure did not act synergistically to produce prolonged and severe disturbances in folate and vitamin B12 metabolism.     

Diagnostic value of the serum folate assay

The diagnostic value of the serum folate assay has been assessed in 90 patients, each of whom had a macrocytic anaemia and a low serum vitamin B(12) level. Twenty-nine (32%) patients were found to have anaemia due primarily to folate deficiency. The cause of the low serum vitamin B(12) levels is uncertain in the 22 (25%) patients with normal or borderline vitamin B(12) absorption. The effect of folic acid therapy was studied in four of these patients, and in each case the serum vitamin B(12) rose slowly to a normal level.The serum folate was low in only four (7.5%) of the 54 patients with pernicious anaemia, and the levels rose to normal on treatment with vitamin B(12) alone. A high serum folate occurred in eight (15%) pernicious anaemia patients. A normal serum folate indicated the diagnosis of pernicious anaemia or megaloblastic anaemia following partial gastrectomy. However, a normal serum folate and a very low vitamin B(12) level was found in two patients with idiopathic steatorrhoea.It is concluded that the serum folate assay is a valuable routine test in patients who have a macrocytic anaemia and low serum vitamin B(12). A low folate level makes the diagnosis of pernicious anaemia unlikely and is a strong indication for full investigation of small intestinal function.     

Mitochondrial vitamin B12-binding proteins in patients with inborn errors of cobalamin metabolism

Inborn errors of vitamin B12 (cobalamin, Cbl) metabolism are autosomal recessive disorders and have been classified into nine distinct complementation classes (cblA-cblH and mut). Disorders affecting methylcobalamin metabolism cause megaloblastic anemia, which may be accompanied by leukopenia and thrombocytopenia, and a variety of neurological problems. Disorders affecting adenosylcobalamin cause methylmalonic acidemia and metabolic acidosis. Previous studies have shown that cobalamin binds to two enzymes in humans: methylmalonyl-CoA mutase in mitochondria and methionine synthase in the cytosol. In this study, cobalamin binding patterns were analyzed in crude mitochondrial fractions obtained from both control and patient fibroblasts that had been incubated with [57Co]cyanocobalamin. Crude mitochondrial fractions from control fibroblasts confirmed that the majority of [57Co]Cbl eluted with methylmalonyl-CoA mutase. However, in six of the nine disorders, at least one previously unidentified mitochondrial cobalamin binding protein was observed to bind [57Co]Cbl. The proportion of [57Co]Cbl that binds, is increased compared to controls when a deficiency in either adenosylcobalamin synthesis or utilization prevents binding to methylmalonyl-CoA mutase. Furthermore, unique cobalamin binding profiles emerged demonstrating how known mutations in these patients affect cobalamin binding to as yet unidentified proteins.     

Clinical spectrum of cobalamin deficiency in Tunisia

PURPOSE: the aim of this study was to determine the prevalence of cobalamin (vitamin B12) deficiency in different populations of patients with clinical manifestations associated or secondary to cobalamin or folates deficiency and to analyse the demographic, clinical, paraclinical investigations in cobalamin deficient patients in Tunisia. METHODS: it was a prospective (1999-2001) multicenter study of 604 patients divided into four groups. The first group is composed of 478 consecutive patients with anaemia and/or macrocytosis with megaloblastic haemopoiesis on bone marrow examination without myelodyslasic or malignancy signs. The second group is made up of 34 patients with unexplained neurological symptoms without the presence of anaemia. The third group was composed of 82 invidious with isolated psychiatric disorders and the 10 patients with Hashimoto thyro?ditis constituted the last group. Results: serum cobalamin level was low in 98 %, 23%, 14% of cases, respectively, in the first three groups. Only one case of patients with Hashimoto thyroiditis has serum cobalamin deficiency. Pernicious anaemia (Biermer's disease) was established by dual isotope schilling examination in 103 patients among a sample of 120 serum cobalamin deficient patients (86%). The median age at presentation was 45.5 years. Severe chronic atrophic gastritis was diagnosed in 97.5% of patients with Biermer's disease. Serum antibodies against intrinsic factor and gastric parietal cells were detected in (42.5%) and (60.6%) patients, respectively; (25.5%) patients had the both types of antibodies. 23.4% patients were positive for antithyroid antibodies. Anti-nuclear antibodies were detected in 3% patients. CONCLUSION: an interesting finding of our study was the high frequency of cobalamin deficiency in Tunisia, particularly in relative young patients. Our patients had classic features of florid cobalamin deficiency (severe haematological manifestations and neuro-psychiatric disorders). The main underlying causes of such deficiencies were Biermer's disease. Subtle clinical manifestations should be recognized and investigated even in young patients at risk.     

Dietary folate and vitamins B12, B6, and B2 intake and the risk of postpartum depression in Japan: the Osaka Maternal and Child Health Study

BACKGROUND: Previous studies showed an inverse association between folate intake and depression. However, epidemiological evidence for folate intake and postpartum depression is unavailable. This prospective study examined the relationship of dietary consumption of folate and B vitamins during pregnancy with the risk of postpartum depression. METHODS: Study subjects were 865 Japanese women. Dietary data were obtained during pregnancy from a validated self-administered diet history questionnaire. Postpartum depression was defined as present when subjects had an Edinburgh Postnatal Depression Scale score of 9 or higher between 2 and 9 months postpartum. Adjustment was made for age, gestation, parity, cigarette smoking, alcohol intake, family structure, family income, education, changes in diet in the previous 1 month, season when data at baseline were collected, body mass index, time of delivery before the second survey, medical problems in pregnancy, baby's sex, and baby's birth weight. RESULTS: Postpartum depression developed in 121 subjects (14.0%) 2 to 9 months postpartum. There was no measurable association between intake of folate, cobalamin, or pyridoxine and the risk of postpartum depression. Compared with riboflavin intake in the first quartile, only riboflavin consumption in the third quartile was independently related to a decreased risk of postpartum depression (multivariate odds ratio: 0.53, 95% CI: 0.29-0.95, P for trend=0.55). LIMITATIONS: Personal and family psychiatric history, sociocultural factors, and personal and family relations were not controlled for. The possibility of misclassification of dietary information during pregnancy should be considered. CONCLUSIONS: Our results suggest that moderate consumption of riboflavin may be protective against postpartum depression.     

Down syndrome and epilepsy: a nutritional connection?

Non-Asian individuals with Down syndrome are much more likely to develop epileptic seizure disorders than individuals without Down syndrome. Examination of nutrient and metabolite levels in patients with these two seemingly disparate disorders reveals numerous similarities. Compared to individuals without these disorders, individuals with Down syndrome and individuals with seizures may have lower levels of vitamin A, vitamin B1, folate, vitamin B12, vitamin C, magnesium, manganese, selenium, zinc, carnitine, carnosine, choline, and possibly serine. Excesses of copper, cysteine, phenylalanine, and superoxide dismutase are also sometimes encountered in both disorders. In addition to common nutritional lower levels and excesses, disorders of metabolism involving vitamin B6, vitamin D, calcium, and tryptophan may play a common role. This paper hypothesizes that nutritional factors may account for the high joint occurrence of these conditions. Further examination of these data may provide insights into nutritional, metabolic and pharmacological treatments for both conditions.     

Neuropsychiatric disorders caused by cobalamin deficiency in the absence of anemia or macrocytosis

Among 141 consecutive patients with neuro-psychiatric abnormalities due to cobalamin deficiency, we found that 40 (28 percent) had no anemia or macrocytosis. The hematocrit was normal in 34, the mean cell volume was normal in 25, and both tests were normal in 19. Characteristic features in such patients included paresthesia, sensory loss, ataxia, dementia, and psychiatric disorders; longstanding neurologic symptoms without anemia; normal white-cell and platelet counts and serum bilirubin and lactate dehydrogenase levels; and markedly elevated serum concentrations of methylmalonic acid and total homocysteine. Serum cobalamin levels were above 150 pmol per liter (200 pg per milliliter) in 2 patients, between 75 and 150 pmol per liter (100 and 200 pg per milliliter) in 16, and below 75 pmol per liter (100 pg per milliliter) in only 22. Except for one patient who died during the first week of treatment, every patient in this group benefited from cobalamin therapy. Responses included improvement in neuropsychiatric abnormalities (39 of 39), improvement (often within the normal range) in one or more hematologic findings (36 of 39), and a decrease of more than 50 percent in levels of serum methylmalonic acid, total homocysteine, or both (31 of 31). We conclude that neuropsychiatric disorders due to cobalamin deficiency occur commonly in the absence of anemia or an elevated mean cell volume and that measurements of serum methylmalonic acid and total homocysteine both before and after treatment are useful in the diagnosis of these patients.     

Psychiatric manifestations of homocystinuria due to cystathionine beta-synthase deficiency: prevalence, natural history, and relationship to neurologic impairment and vitamin B6-responsiveness

Homocystinuria commonly affects the central nervous system (CNS), primarily as mental retardation, seizures, and stroke. Case reports have long suggested a predisposition to schizophrenia, but no careful study of predisposition to psychiatric illness has been performed. Accordingly, we evaluated 63 persons with homocystinuria due to cystathionine beta-synthase deficiency for psychiatric disturbance, intelligence, evidence of other CNS problems, and responsiveness to vitamin B6. The overall rate of clinically significant psychiatric disorders was 51%, predominated by four diagnostic categories: episodic depression (10%), chronic disorders of behavior (17%), chronic obsessive-compulsive disorder (5%), and personality disorders (19%). The average IQ was 80 +/- 27 (1 SD); and an IQ of less than or equal to 79 was two-thirds more common among vitamin B6-nonresponsive patients compared to vitamin B6-responsive patients. Aggressive behavior and other disorders of conduct were particularly common among patients with mental retardation and among vitamin B6-nonresponsive patients.