The effect of vitamin B12 deficiency on methylfolate metabolism and pteroylpolyglutamate synthesis in human cells.

Abstract

1. The uptake of 14C from [methyl-14C]methyItetrahydrofolate was significantly reduced in the phytohaemagglutinin (PHA)-stimulated lymphocytes from nine patients with untreated pernicious anaemia compared with the uptake in seven normal subjects. 2. The uptake of 14C from [14C]methyltetrahydrofolate by the lymphocytes from seven of the patients with pernicious anaemia was consistently increased by addition of vitamin B12in vitro. 3. The proportion of 14C taken up from [14C]methyltetrahydrofolate transferred to non-folate compounds was found to be significantly reduced in the PHA-stimulated lymphocytes from nine patients with untreated pernicious anaemia compared with the proportion transferred in the PHA-stimulated lymphocytes from seven normal subjects. Addition of vitamin B12in vitro consistently increased the transfer in vitamin B12-deficient cells but had no consistent effect in normal cells. 4. Normal and vitamin B12-deficient PHA-stimulated lymphocytes took up [3H]folic acid and after 72 h incubation converted this largely into pteroylpolyglutamate forms. 5. The proportion of labelled lymphocyte folate as pteroylpolyglutamate after incubation with [3H]folic acid was the same in vitamin B12-deficient as in normal lymphocytes and the proportion of pteroylpolyglutamates formed in vitamin B12-deficient lymphocytes was unaffected by addition of vitamin B12in vitro. 6. No radioactivity could be decteted in pteroylpolyglutamates after incubating normal PHA-stimulated lymphocytes with [14C]methyltetrahydrofolate for 72 h, suggesting that pteroylpolyglutamate forms of folate cannot be made directly from methyltetrahydrofolate. 7. These results are consistent with the ‘methyltetrahydrofolate trap’ hypothesis in vitamin B12 deficiency. It is suggested that reduced synthesis of pteroylpolyglutamates reported by others in vitamin B12-deficient cells may be secondary to the failure of removal of the methyl group from methyltetrahydrofolate rather than to a direct effect of vitamin B12 deficiency on the enzyme responsible for pteroylpolyglutamate synthesis. 8. Reduced entry of methyltetrahydrofolate into vitamin B12-deficient cells may be secondary to failure of conversion of this compound into tetrahydrofolate.