Clinical Laboratory Strategies: April 30, 2009

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Finding the Link between Vitamin B12 and Neural Tube Defects
Lower vitamin B12 levels put newborns at risk for neural tube defects
By Stuart Zehner

Neural tube defects, birth defects of the brain and spinal column, have long been linked to maternal deficiencies in folate. Researchers have speculated that in addition to folate, vitamin B12 might play a role in the occurrence of these defects. A recently published study looked for a possible link between these two metabolically related vitamins. This issue of Strategies examines those findings.

In a new study conducted by researchers from Trinity College Dublin, the U.S. National Institutes of Health, and the Health Research Board of Ireland, maternal vitamin B12 status and the risk for neural tube defects (NTDs) was evaluated in a population of pregnant women who had not received folic acid fortification and who were therefore at risk of giving birth to babies with NTDs (Pediatrics 2009; 123:917-923). Folic acid, the artificial form of folate also known as vitamin B9, can prevent many, but not all, NTDs. Based on this information, the Food and Drug Administration began requiring that certain grains be fortified with folic acid in 1996, which later became mandatory in 1998. Previous studies indicated that low maternal levels of vitamin B12, which is metabolically linked to folate, could play a similar role in the occurrence of NTDs.

In the new study, researchers found that vitamin B12 < 250 ng/L is an independent maternal risk factor for having a NTD-affected pregnancy. The study, which is the first to examine the NTD risk as a function of vitamin B12 levels, suggests that women, especially if they are planning to become pregnant, should be monitored for vitamin B12 levels and folate levels.

But researchers remain unsure about how the mother’s vitamin B12 status leads to NTDs. "We know that folate prevents NTDs, but there’s a lot of speculation as to the mechanisms," said James L. Mills, MD, MS, a senior investigator at the National Institute of Child Health and Human Development and one of the co-authors of the study. "We know that it is related to methylation, and that would be most applicable to vitamin B12. Folate and vitamin B12 interact in generating methyl groups. They convert homocysteine to methionine, and methionine provides methyl groups for many reactions involving DNA and proteins. Essentially, problems with vitamin B12 and folate lead to insufficient methylation and regulation of DNA."

Evaluating the Risks

The retrospective study assessed maternal vitamin B12 status in blood samples taken at an average 15 weeks gestation from three separate groups of Irish women. The blood samples originally were drawn between 1983 and 1990, a time that predates widespread food fortification with folic acid and when medical advice recommended that pregnant women avoid additional vitamin supplements. Today, the rates of NTDs are especially high in Ireland, around 1 per 1000 births, while rates in the U.S. have fallen to about 1 per 2000 births since grain fortification began, according to Mills.

The researchers analyzed samples from three different groups. The first group consisted of blood samples from 95 women during a NTD-affected pregnancy and 265 control subjects. The second cohort consisted of blood samples from 107 women who had a previous NTD-affected pregnancy, but whose current pregnancy -had not been affected, and 414 control subjects. The third group also contained blood samples from 76 women during an NTD-affected pregnancy and 222 control subjects. However, subjects from the third group were enrolled in a different study than those in the first group. Known vitamin supplement users were excluded from the study. In all three arms of the study, the researchers measured the levels of folate and vitamin B12 in the blood samples and compared them to controls whose pregnancies were not affected by NTDs.

Statistical methods were used to evaluate the role of vitamin B12 separately from that of folate. In all three study arms, subjects with vitamin B12 concentrations of <250 ng/L had a 2.5 to 3-fold higher risk of having an NTD-affected baby, even after adjusting for folate levels. The study also showed very clear risk trends for those with the lowest levels of vitamin B12: subjects with levels <150 ng/L had an approximately 5-times higher risk of having an NTD-affected pregnancy compared to those with vitamin B12 levels >400 ng/L. The researchers speculate that there may be some risk reduction with vitamin B12 levels between 320 to 350 ng/L, but no statistically significant association was found in this study. The majority of risk, however, was confined to those with vitamin B12 levels <250 ng/L, though further risk reduction was achieved by having B12 status between 320 to 350 ng/L.

These results were not completely expected by the researchers. "We were a little surprised there wasn’t a more across-the-board risk," said Mills. "But I guess in retrospect it makes sense that the people who are at a level where you might see physiological impairment would be the ones at the most risk."

"The findings of this study are significant because it shows that vitamin B12 is important to neural tube defects. There is a strong link between folate and vitamin B12, and we need to watch deficiencies in vitamin B12, especially during pregnancy, because metabolically these two are connected," said Majid Moridani, PharmD, PhD, DABCC, FACB, and assistant professor of pharmaceutical sciences and clinical chemistry at Texas Tech University Health Sciences Center. Moridani was not involved in the study. "It is important in terms of evidence-based medicine that they were able to come up with a cutoff."

Most importantly, women need to know the risks associated with deficiencies in vitamin B12 and take the necessary steps to make sure their levels are adequate before they become pregnant. The study suggests that women aim to enter pregnancy with serum B12 levels of >300 ng/L, and that levels >400 ng/L might be desirable even though there is no statistically significant benefit. "Women of child-bearing age who are capable of becoming pregnant—not if they are planning to, but if they are capable of becoming pregnant—need to take folic acid just to be safe," stressed Mills. "The neural tube closes by the 28th day after conception, and that’s why it’s important to get women to do all of this before they become pregnant because there really isn’t time once they realize they are pregnant."

It is uncertain whether the incidence of NTDs can be further reduced in the U.S. by increasing the level of fortification of grains with folic acid. The addition of vitamin B12 to fortified grains maybe an acceptable and useful way to further reduce the incidence of NTDs, but further studies would be required to establish the safety of B12 fortification and the necessary dose that would be needed to reach an effective level of protection, according to the authors. "Many people think about B12 fortification in flour, similar to folate. Alternatives to consider are fruit juices, milk and breakfast cereals but I think it’s better to consider fortifying drinks such as milk with folate and vitamin B12 for those who can tolerate milk as part of their diet because milk is a good source for vitamin D and calcium," suggested Moridani. "In addition, one should note that vitamin B12 and folate have a very low potential for toxicity, therefore the excess of these two vitamins are not harmful. Co-fortification of folate and vitamin B12 makes sense because an excess of folate can mask vitamin B12 deficiency," added Moridani.

Ultimately, most NTDs are easily preventable as long as people understand the risks and eat a diet rich in folate and vitamin B12. "Women need to take folic acid if they are capable of becoming pregnant because that is the best and surest way to prevent NTDs," stressed Mills. "As for vitamin B12, there are some populations, like vegans or people with GI problems that might interfere with absorption, who need to be identified before they become pregnant and warned about the need for vitamin B12." 

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