Inherited errors of cobalamin metabolism and their management.

Cobalamins are essential biological compounds structurally related to haemoglobin and the cytochromes. Although the basic cobalamin molecule is only synthesized by micro-organisms, all mammalian cells can convert this into the coenzymes adenosylcobalamin (AdoCbl) and methylcobalamin (MeCbl). AdoCbl is the major form in cellular tissues, where it is retained in the mitochondria. MeCbl predominates in blood plasma and certain other body fluids such as breast milk; in cells MeCbl is found in the cytosol. Inherited disorders of cobalamin metabolism are single gene defects, transmitted as recessive traits. They affect absorption, transport or intracellular metabolism of cobalamin. At least 12 different mutations are known, including defects or deficiencies of IF, IF-receptor and TCII, MM-CoA mutase and of the various reductases and synthases required for synthesis of AdoCbl and MeCbl. These have been designated cblA to cblG. Abnormalities are detectable by urine and plasma assays of methylmalonic acid and homocysteine, and plasma and erythrocyte analysis of cobalamin coenzymes, which can reveal deficiencies of MeCbl or AdoCbl. Fibroblast studies discriminate between closely similar defects. In man, AdoCbl is required in only two reactions: the catabolic isomerization of MM-CoA to succinyl-CoA and interconversion of alpha- and beta-leucine. MeCbl is required in the anabolic transmethylation of homocysteine to methionine. Intestinal absorption of cobalamin requires the glycoproteins TCI and IF from the stomach and IF-cobalamin receptors in the ileum. Cobalamin is transported to cells bound to a polypeptide, TCII, is captured by surface receptors and absorbed by endocytosis. The complex is then split in the lysosomes, cobalamin is released and the coenzymes are synthesized. In plasma, 80-90% of the cobalamin is bound to TCI, whose function is uncertain. Megaloblastic anaemia at birth or in the first few weeks of life is a rare but serious event. Myelopathy and developmental delay, with or without seizures may also occur without anaemia. If urine and light-protected blood samples are collected and sent to an appropriate metabolic unit, an inborn error of cobalamin metabolism, including TCII deficiency in which the serum B12 may be normal, can quickly be diagnosed. IF deficiency or Imerslund-Gräsbeck disease usually presents with signs of cobalamin deficiency within the first year of life and can be diagnosed by absorption studies. Current treatment involves dietary protein restriction and/or parenteral OHCbl and the prognosis is very variable. Since lack of MeCbl leads to depressed DNA synthesis affecting rapidly dividing cells in the brain and elsewhere, treatment with this coenzyme should be considered at the earliest stage in appropriate cases.(ABSTRACT TRUNCATED AT 400 WORDS)

Multiple sclerosis associated with vitamin B12 deficiency.

We describe 10 patients with a previously unreported, to our knowledge, association of multiple sclerosis and unusual vitamin B12 deficiency. The clinical features and the age at presentation were typical of multiple sclerosis, with eight cases occurring before age 40 years, which is a rare age for vitamin B12 deficiency. Nine patients had hematologic abnormalities, but only two were anemic. All six patients examined had low erythrocyte cobalamin levels. Only two patients had pernicious anemia; in the remaining patients the vitamin B12 deficiency was unexplained. A vitamin B12 binding and/or transport is suspected. The nature of the association of multiple sclerosis and vitamin B12 deficiency is unclear but is likely to be more than coincidental. Further studies of vitamin B12 metabolism, binding, and transport in multiple sclerosis are indicated, as these cases may offer a clue to the understanding of a still mysterious neurologic disorder.

The biochemical systematics of red and sika deer (genus Cervus) in Ireland.

Twenty enzyme loci were investigated using starch gel electrophoresis in managed populations of red deer (Cervus elaphus) and sika deer (C. nippon) from Ireland. The red deer, which originated from several European countries, were polymorphic (%P) at 25% of the loci examined and exhibited a mean heterozygosity (H) of 0.025. The sika, originally from Japan, were introduced to Ireland in very limited numbers. Here, %P was 5% and H was 0.006. Mean genetic distance (D) between these taxa was 0.160. Because Scottish red deer, the source of much of the Irish herd, had previously been investigated for 16 of the same enzyme loci, it was possible to merge the present data with results from that earlier study, which also included North American wapiti (C. elaphus canadensis). The resulting phenogram based on genetic distance demonstrates that sika are twice as distantly related to red deer and wapiti, than the latter are to each other and suggests that sika-like forms are ancestral. Also, the demonstration of absolute mobility differences in the products of two enzyme loci in red deer and sika has many practical applications.

Vitamin B12 homoeostasis after haemorrhage in the rat: the importance of skeletal muscle.

1. The effects of haemorrhage on vitamin B12 (cobalamin) homoeostasis have been studied in rats with and without prior administration of 57Co-labelled cyanocobalamin. 2. Arteriovenous differences in radioactivity and endogenous B12 were measured across the thigh extensor muscles. Radioactivity and endogenous B12 were also measured in skeletal muscle, liver and kidney. 3. Haemorrhage induced a marked rise in circulating B12 with a concomitant loss of the vitamin from skeletal muscle and liver, but in kidney there was a net gain. Restoration of the blood volume with autologous plasma did not prevent release of B12 from tissues. 4. The results suggest that vitamin B12 reserves may be seriously depleted by the wasting of skeletal muscle.

Tissue distribution of methylcobalamin in rats fed amino acid-defined, methyl-deficient diets.

Total cobalamin and methylcobalamin levels were determined in tissues of male F344 rats fed a complete, amino acid-defined diet or a diet deficient in methionine, choline and/or cyanocobalamin. Total cobalamin levels in rats fed the complete diet were (picograms/milligram tissue +/- SEM): liver, 67 +/- 13; kidneys, 738 +/- 133; spleen, 23 +/- 2; and adrenals, 268 +/- 36. Corresponding methylcobalamin levels were: liver, 1.6 +/- 0.5; kidneys, 107.6 +/- 22.2; spleen, 0.3 +/- 0.1; and adrenals, 26.9 +/- 5.3; these values represent 2.4, 14.5, 1.4 and 9.7%, respectively, of the total cobalamin levels. Total cobalamin levels of all tissues studied were altered by cobalamin deprivation alone or in conjunction with methionine and/or choline deprivation. Methylcobalamin levels were more resistant to dietary alteration. Regardless of the presence or absence of methionine and cobalamin in the diet, choline deprivation always decreased the proportion of methylcobalamin in the liver. Kidney levels of methylcobalamin, like those of total cobalamin, were decreased by removal of cobalamin from the complete or the methyl-deficient diets. The results demonstrate that cobalamin, methionine and choline exert quite different effects on tissue levels of the cobalamins in rats.

A sensitive micromethod for the routine estimation of methylmalonic acid in body fluids and tissues using thin layer chromatography.

A routine method for the rapid estimation of methylmalonic acid (MMA) has been developed using thin-layer chromatography (TLC) on cellulose, locating the separated MMA zones by coupling with a diazo reagent and quantitating the results by densitometric scanning of a photocopy of the chromatogram on transparency film. The method can detect as little as 25 pg MMA in small volumes of urine, plasma and other body fluids or tissue homogenates, has potential for the simultaneous estimation of MMA, other organic acids, methylmalonyl coenzyme A and creatinine and should be of particular value in the investigation of occult cobalamin deficiency or suspected errors of vitamin B12 or intermediary metabolism.

Problems with the serum vitamin B12 assay.

Quality control trials have shown that, in routine practice, serum vitamin B12 estimations vary so much from laboratory to laboratory that serious confusion can result irrespective of whether microbiological or radioassay methods are used. Even experienced centres find the assay too insensitive and non-specific for a low level to be used as the sole criterion of vitamin B12 deficiency; the haemopoietic and biochemical sequelae of vitamin B12 deficiency also correlate poorly with the serum level. These basic difficulties with the assay seem to stem from the pattern of vitamin B12 (or cobalamin) binding in serum. Human beings are unique in having virtually all of their cobalamin attached to an apparently functionless binder, transcobalamin I. It is therefore not surprising that the serum cobalamin is such a poor predictor of cobalamin deficiency. The metabolically important serum binder is transcobalamin II and deficiency of this protein causes a potentially lethal megaloblastic anaemia even though the serum cobalamin level is normal. Tissue cobalamin depletion with normal serum levels also occurs after nitrous oxide inhalation and in certain inborn errors of metabolism.

Granulocyte dysfunction in transcobalamin II deficiency responding to leucovorin or hydroxocobalamin-plasma transfusion.

Granulocytes from a 6-year-old boy with congenital transcobalamin II (TC II) deficiency were found to have abnormally low antibacterial activity against Staphylococcus aureus and very low intracellular levels of the cobalamin coenzymes. Transfusion of hydroxocobalamin (OH-Cbl) bound to normal plasma temporarily restored granulocyte bactericidal activity and increased cellular levels of the cobalamin coenzymes. Granulocyte function was also temporarily restored by oral Leucovorin. The defect appeared to be causally related to the patient's TC II deficiency and indirectly to a deficiency of cobalamin and folate coenzymes.

Defective adenosylcobalamin synthesis in a case of transcobalamin II deficiency.

Cobalamin metabolism has been investigated in a new case of transcobalamin II (TC II) deficiency. Using the chromatobioautographic technique, an abnormal distribution of cobalamins was detected in the child's erythrocytes and reduced synthesis of adenosylcobalamin but not of methylcobalamin in cultured fibroblasts. These results suggest that there may be a close link between TC II-mediated cobalamin transport and intracellular synthesis of adenosylcobalamin (Ado-Cbl).

Interference by methylcobalamin analogues with synthesis of cobalamin coenzymes in human lymphocytes in vitro.

1. 72 h uptake of cyano[57Co]cobalamin and formation of 57Co-labelled methylcobalamin, adenosylcobalamin and hydroxocobalamin has been estimated with and without the addition of methylcobalamin analogues in phytohaemagglutinin-stimulated lymphocytes from healthy human subjects. 2. Difluorochloromethylcobalamin reduced cell uptake of cyanocobalamin and caused a disproportionate reduction in synthesis of adenosylcobalamin. 3. Methylcobalamin-palladium trichloride reduced cell uptake of cyanobalamin more effectively than did difluorochloromethylcobalamin and reduced the formation of methylcobalamin, adenosylcobalamin and hydroxocobalamin in proportion. 4. The results suggest that in addition to inhibiting uptake of cyanocobalamin, one or both compounds may have interfered directly with the mechanism of synthesis of the cobalamin coenzymes.

[Granulocyte dysfunction in transcobalamin II deficiency]. / Granulozytendysfunktion bei Transcobalamin-II-Mangel.

Granulocytes from a boy with congenital transcobalamin II (TC II) deficiency were found to have abnormally low antibacterial activity against Staphylococcus aureus. Transfusion of normal plasma supplemented with hydroxocobalamin temporarily restored granulocyte bactericidal activity to normal. Granulocyte function was also temporarily restored by oral leucovorin. The defect appears to be causally related to the patient's TC II deficiency and indirectly to an intracellular deficiency of cobalamin and folate coenzymes [1].

Effects of low cobalamin diet and chronic cyanide toxicity on cobalamin distribution in baboons.

This paper reports the bodily distribution of total cobalamin and individual cobalamins at the termination of an experiment on the effects of a low cobalamin diet and chronic cyanide or thiocyanate administration in baboons. The results show that the distribution of cobalamins in the tissues of the baboon can be altered by a low cobalamin diet and also by chronic intoxication with cyanide, whether or not the animals are on a low cobalamin diet. All animals on the low cobalamin diet showed a reduction in total and individual cobalamins. In blood plasma and erythrocytes, kidney, spleen, testis and brain, the proportion of methylcobalamin tended to be disproportionately reduced in cobalamin-depleted animals. This reduction was lessened or prevented by the administration of cyanide. Neither cyanide not thiocyanate produced a significant increase in the proportion of cyanocobalamin in plasma, though thiocyanate produced a large increase in cyanocobalamin in erythrocytes. In liver, cyanocobalamin was more than doubled by the administration of cyanide to cobalamin-depleted animals.