Targeted metabolomics and mathematical modeling demonstrate that vitamin B-6 restriction alters one-carbon metabolism in cultured HepG2 cells

Abstract

Low vitamin B-6 nutritional status is associated with increased risk for cardiovascular disease and certain cancers. Pyridoxal 5'-phosphate (PLP) serves as a coenzyme in many cellular processes, including several reactions in one-carbon (1C) metabolism and the transsulfuration pathway of homocysteine catabolism. To assess the effect of vitamin B-6 deficiency on these processes and associated pathways, we conducted quantitative analysis of 1C metabolites including tetrahydrofolate species in HepG2 cells cultured in various concentrations of pyridoxal. These results were compared with predictions of a mathematical model of 1C metabolism simulating effects of vitamin B-6 deficiency. In cells cultured in vitamin B-6-deficient medium (25 or 35 nmol/l pyridoxal), we observed >200% higher concentrations of betaine (P < 0.05) and creatinine (P < 0.05) and >60% lower concentrations of creatine (P < 0.05) and 5,10-methenyltetrahydrofolate (P < 0.05) compared with cells cultured in medium containing intermediate (65 nmol/l) or the supraphysiological 2,015 nmol/l pyridoxal. Cystathionine, cysteine, glutathione, and cysteinylglycine, which are components of the transsulfuration pathway and subsequent reactions, exhibited greater concentrations at the two lower vitamin B-6 concentrations. Partial least squares discriminant analysis showed differences in overall profiles between cells cultured in 25 and 35 nmol/l pyridoxal vs. those in 65 and 2,015 nmol/l pyridoxal. Mathematical model predictions aligned with analytically derived results. These data reveal pronounced effects of vitamin B-6 deficiency on 1C-related metabolites, including previously unexpected secondary effects on creatine. These results complement metabolomic studies in humans demonstrating extended metabolic effects of vitamin B-6 insufficiency.

Investigation of vitamin B6 inadequacy, induced by exposure to the anti-B6 factor 1-amino d-proline, on plasma lipophilic metabolites of rats: a metabolomics approach

“…However, targeted metabolomics of vitamin B 6 restriction in cultured HepG2 cells and healthy humans revealed fundamental changes in the metabolism of amino acids, carbohydrates, organic acid, neurotransmitters and metabolites of the one-carbon pathway [22,42]. This first metabolomics data on oral exposure of antivitamin B 6 in a rodent model identified novel plasma lipophilic biomarkers and their associated metabolic pathways of vitamin B 6 deficiency and antivitamin B 6 effect.…”

Pyridoxine supplementation does not alter in vivo kinetics of one-carbon metabolism but modifies patterns of one-carbon and tryptophan metabolites in vitamin B-6–insufficient oral contraceptive users

“…Increased glycine has been reported in liver (49,50), muscle, and plasma of vitamin B-6-deficient rats (50,51), in the plasma of vitamin B-6-deficient humans (16,18,20,23), and in vitamin B-6restricted cultured cells (52), concurrent with modeling predictions (24). Major processes for the metabolism of glycine include PLP-dependent reactions catalyzed by serine hydroxymethyltransferase isoforms and by glycine decarboxylase in the glycine cleavage system (53).…”

Metabolite Profile Analysis Reveals Association of Vitamin B-6 with Metabolites Related to One-Carbon Metabolism and Tryptophan Catabolism but Not with Biomarkers of Inflammation in Oral Contraceptive Users and Reveals the Effects of Oral Contraceptives on These Processes,

“…A discriminant analysis of the contribution of each variable to group differences according to vitamin B-6 status was conducted through use of the variable influence on projection (VIP) method (33,35). As stated previously (20,36), the VIP values represent weighted sums of squares of the PLS weights of the various metabolites and take into account the proportion of Y-variance in each dimension (34). In each plot, each variable is provided with a VIP value and a 95% CI derived from jack-knifing (34,35,37).…”