Vitamin B-6 restriction tends to reduce the red blood cell glutathione synthesis rate without affecting red blood cell or plasma glutathione concentrations in healthy men and women.

BACKGROUND: Glutathione plays various protective roles in the human body. Vitamin B-6 as pyridoxal-5'-phosphate (PLP) is required as the coenzyme in the formation of glutathione precursors. Despite this obligatory role of PLP, previous studies from this laboratory showed that vitamin B-6 deficiency caused elevated glutathione concentrations in rat liver and human plasma. OBJECTIVE: Our aim was to determine the effect of marginal vitamin B-6 deficiency (plasma PLP 20-30 nmol/L) on the rate of red blood cell (RBC) glutathione synthesis. DESIGN: We measured plasma and RBC glutathione concentrations and the fractional and absolute synthesis rates of RBC glutathione using the stable-isotope-labeled glutathione precursor [1,2-(13)C(2)]glycine in 13 healthy volunteers aged 21-39 y. RESULTS: Dietary vitamin B-6 restriction did not significantly affect the glutathione concentration in plasma (6.9 +/- 1.9 compared with 6.7 +/- 1.1 micromol/L) or RBCs (2068 +/- 50 compared with 2117 +/- 48 micromol/L). For RBC glutathione, the mean fractional synthesis rates were 54 +/- 5%/d and 43 +/- 4%/d (P = 0.10), and the absolute synthesis rates were 1116 +/- 100 and 916 +/- 92 micromol . L(-1) . d(-1) (P = 0.14) before and after vitamin B-6 restriction, respectively. CONCLUSIONS: Marginal vitamin B-6 deficiency tended to decrease mean RBC glutathione synthesis with no effect on RBC glutathione concentration, but the responses varied widely among individuals. Because the cysteine concentration in plasma and RBC did not change during vitamin B-6 restriction, we conclude that the effects of marginal vitamin B-6 deficiency on glutathione synthesis are not caused by altered precursor concentrations.

Production of 1-carbon units from glycine is extensive in healthy men and women.

Glycine undergoes decarboxylation in the glycine cleavage system (GCS) to yield CO(2), NH(3), and a 1-carbon unit. CO(2) also can be generated from the 2-carbon of glycine by 10-formyltetrahydrofolate-dehydrogenase and, after glycine-to-serine conversion by serine hydroxymethyltransferase, from the tricarboxylic acid cycle. To evaluate the relative fates of glycine carbons in CO(2) generation in healthy volunteers (3 male, 3 female, aged 21-26 y), primed, constant infusions were conducted using 9.26 micromol x h(-1) x kg(-1) of [1,2-(13)C]glycine and 1.87 micromol x h(-1) x kg(-1) of [5,5,5-(2)H(3)]leucine, followed by an infusion protocol using [1-(13)C]glycine as the glycine tracer. The time period between the infusion protocols was >6 mo. In vivo rates of whole-body glycine and leucine flux were nearly identical in protocols with [1,2-(13)C]glycine and [5,5,5-(2)H(3)]leucine and with [1-(13)C]glycine and [5,5,5-(2)H(3)]leucine tracers, which showed high reproducibility between the tracer protocols. Using the [1-(13)C]glycine tracer, breath CO(2) data showed a total rate of glycine decarboxylation of 96 +/- 8 micromol x h(-1) x kg(-1), which was 22 +/- 3% of whole-body glycine flux. In contrast, infusion of [1,2-(13)C]glycine yielded a glycine-to-CO(2) flux of 146 +/- 37 micromol x h(-1) x kg(-1) (P = 0.026). By difference, this implies a rate of CO(2) formation from the glycine 2-carbon of 51 +/- 40 micromol x h(-1) x kg(-1), which accounts for approximately 35% of the total CO(2) generated in glycine catabolism. These findings also indicate that approximately 65% of the CO(2) generation from glycine occurs by decarboxylation, primarily from the GCS. Further, these results suggest that the GCS is responsible for the entry of 5,10-methylenetetrahydrofolate into 1-carbon metabolism at a very high rate ( approximately 96 micromol x h(-1) x kg(-1)), which is approximately 20 times the demand for methyl groups for homocysteine remethylation.

Moderate dietary vitamin B-6 restriction raises plasma glycine and cystathionine concentrations while minimally affecting the rates of glycine turnover and glycine cleavage in healthy men and women.

Glycine is a precursor of purines, protein, glutathione, and 1-carbon units as 5,10-methylenetetrahydrofolate. Glycine decarboxylation through the glycine cleavage system (GCS) and glycine-serine transformation by serine hydroxymethyltransferase (SHMT) require pyridoxal 5'-phosphate (PLP; active form of vitamin B-6) as a coenzyme. The intake of vitamin B-6 is frequently low in humans. Therefore, we determined the effects of vitamin B-6 restriction on whole-body glycine flux, the rate of glycine decarboxylation, glycine-to-serine conversion, use of glycine carbons in nucleoside synthesis, and other aspects of 1-carbon metabolism. We used a primed, constant infusion of [1,2-(13)C(2)]glycine and [5,5,5-(2)H(3)]leucine to quantify in vivo kinetics in healthy adults (7 males, 6 females; 20-39 y) of normal vitamin B-6 status or marginal vitamin B-6 deficiency. Vitamin B-6 restriction lowered the plasma PLP concentration from 55 +/- 4 nmol/L (mean +/- SEM) to 23 +/- 1 nmol/L (P < 0.0001), which is consistent with marginal deficiency, whereas the plasma glycine concentration increased (P < 0.01). SHMT-mediated conversion of glycine to serine increased from 182 +/- 7 to 205 +/- 9 micromol x kg(-1) x h(-1) (P < 0.05), but serine production using a GCS-derived 1-carbon unit (93 +/- 9 vs. 91 +/- 6 micromol x kg(-1) x h(-1)) and glycine cleavage (163 +/- 11 vs. 151 +/- 8 micromol x kg(-1) x h(-1)) were not changed by vitamin B-6 restriction. The GCS produced 1-carbon units at a rate (approximately 140-170 micromol x kg(-1) x h(-1)) that greatly exceeds the demand for remethylation and transmethylation processes (approximately 4-7 micromol x kg(-1) x h(-1)). We conclude that the in vivo GCS and SHMT reactions are quite resilient to the effects of marginal vitamin B-6 deficiency, presumably through a compensatory effect of increasing substrate concentration.

Evaluation of long-term treatment of children with congenital lactic acidosis with dichloroacetate.

OBJECTIVE: The purpose of this research was to report results on long-term administration of dichloroacetate in 36 children with congenital lactic acidosis who participated previously in a controlled trial of this drug. PATIENTS AND METHODS: We conducted a randomized control trial, followed by an open-label study. Data were analyzed for each patient from the time they began treatment through May 2005. RESULTS: Subject exposure to dichloroacetate totaled 110.42 years. Median height and weight increased over time, but the standardized values declined slightly and remained below the first percentile. There were no significant changes in biochemical metabolic indices, except for a 2% rise in total protein and a 22% increase in 24-hour urinary oxalate. Both the basal and carbohydrate meal-induced rises in lactate were blunted by dichloroacetate. The median cerebrospinal fluid lactate also decreased over time. Conduction velocity decreased and distal latency increased in peroneal nerves. Mean 3-year survival for all of the subjects was 79%. CONCLUSIONS: Oral dichloroacetate is generally well tolerated in young children with congenital lactic acidosis. Although continued dichloroacetate exposure is associated with evidence of peripheral neuropathy, it cannot be determined whether this is attributable mainly to the drug or to progression of underlying disease.

Glycine turnover and decarboxylation rate quantified in healthy men and women using primed, constant infusions of [1,2-(13)C2]glycine and [(2)H3]leucine.

Glycine plays several roles in human metabolism, e.g. as a 1-carbon donor, in purine synthesis, and as a component of glutathione. Glycine is decarboxylated via the glycine cleavage system (GCS) that yields concurrent generation of a 1-carbon unit as 5,10-methylenetetrahydrofolate (methyleneTHF). Serine hydroxymethyltransferase (SHMT) catalyzes the interconversion of glycine and serine, another 1-carbon donor. The quantitative role of glycine in human 1-carbon metabolism has received little attention. The aim of this protocol was to quantify whole body glycine flux, glycine to serine flux, and rate of glycine cleavage in humans. A primed, constant infusion with 9.26 micromol x kg(-1) x h(-1) [1,2-(13)C2]glycine and 1.87 micromol x kg(-1) x h(-1) [(2)H3]leucine was used to quantify the kinetic behavior of glycine in young, healthy volunteers (n = 5) in a fed state. The isotopic enrichment of infused tracers and metabolic products in plasma, as well as breath (13)CO2 enrichment, were determined for use in kinetic analysis. Serine synthesis by direct conversion from glycine via SHMT occurred at 193 +/- 28 micromol x kg(-1) x h(-1) (mean +/- SEM), which comprised 41% of the 463 +/- 55 micromol x kg(-1) x h(-1) total glycine flux. Nearly one-half (46%) of the glycine-to-serine conversion occurred using GCS-derived methyleneTHF 1-carbon units. Based on breath (13)CO2 measurement, glycine decarboxylation (190 +/- 41 micromol x kg(-1) x h(-1)) accounted for 39 +/- 6% of whole body glycine flux. This study is the first to our knowledge to quantify human glycine cleavage and glycine-to-serine SHMT kinetics. GCS is responsible for a substantial proportion of whole body glycine flux and constitutes a major route for the generation of 1-carbon units.