Risk of Neural Tube Defect-Affected Pregnancy Is Associated with a Block in Maternal One-Carbon Metabolism at the Level ofN-5-Methyltetrahydrofolate:Homocysteine Methyltransferase

The disposition of whole blood mono- to hexaglutamyl methylfolate and plasma homocysteine (HCY) was used to evaluate potential lesion sites in one-carbon metabolism which could be responsible for neural tube defect(NTD)-affected pregnancies. An isocratic high-performance liquid chromatographic system (HPLC) with photodiode array detection was used to quantify and speciate whole-blood methylfolate into mono-, di-, tri-, tetra-, penta-, and hexaglutamate forms. This technique was also used with off-line radioassay to identify nonmethyl whole-blood folates. Isocratic HPLC with fluorescence detection was used to quantify SBDF derivatized homocysteine in plasma. The study investigated blood from 11 women who had experienced a previous NTD- affected pregnancy and 11 controls of similar age and social class. No subjects were pregnant. HCY levels were significantly higher in NTD subjects (P= 0.0486, 95% CI −2.799,0.001 using the Mann–Whitney test), as was the ratio of known intracellular (tri- to hexaglutamyl) methylfolate compared to extracellular (mono- and diglutamyl) methylfolate (P= 0.0062 95% CI −0.543, 3.862 using the Mann–Whitney test). Vitamin B₁₂, red cell folate, circulating total methylfolate, and circulating mono- to hexaglutamyl methylfolates showed no difference between population groups. The disposition between individual and cumulative glutamate chain lengths of methylfolate showed significant trends which differed between population groups: (i) total blood methylfolate (Glu_1–6) appears to be utilized byN-5-methyltetrahydrofolate:homocysteine methyltransferase (MS) in control blood but not NTD blood, where it appears to accumulate following a 45-min incubation; (ii) whole-blood hexaglutamyl methylfolate (5CH₃-H₄PteGlu₆) becomes a larger proportion of the total blood methylfolate in NTD than in control populations; and (iii) the intermediate glutamate chains of methylfolate (Glu_1–5) remain relatively constant as 5CH₃-H₄PteGlu₆accumulates in NTD but appear to increase linearly with 5CH₃-H₄PteGlu₆in controls. The significant elevation of HCY in the NTD population is associated with the increasing proportion of 5CH₃-H₄PteGlu₆relative to the total methylfolate, since, when corrected for HCY level, the proportion of 5CH₃-H₄PteGlu₆to total methylfolate is similar in NTD and control populations. These trends are consistent with a defect at the level of vitamin B₁₂dependent MS which “traps” folate at the 5CH₃-H₄PteGlu₆level.