What really causes neural tube defects?

Dr Clare Craig

Mandatory fortification with folic acid depends on two beliefs. The first is that these defects arise because women do not get enough folate from their food. The second is that raising folate intake across the whole population will therefore prevent them. Both the claim about cause and the claim about remedy must be true to justify the intervention.

Folic acid at pharmaceutical doses does reduce recurrence of neural tube defects in women who have already had an affected pregnancy. The trials were clear about that. However, that is not enough to support the idea that folate deficiency is the cause. Trials of lower doses in women at ordinary risk also reported fewer affected births, however, researchers in that study pointed out that whole effect could have been because of the increased risk of pregnancy loss. What none of these trials establishes is that the defects arose in the first place because the mothers were short of folate.

Nevertheless, the international resolutions and national policies on folic acid fortification are framed as the correction of micronutrient deficiency. At a minimum that is an oversimplification. For a start, the World Health Organization’s own guidance accepts that low folate cannot account for every case.

What would falsify it

If these defects are principally a disease of dietary folate shortage, these things ought to be true.

1. High risk folate deficient group. There should be a group of women who are short of folate. The affected pregnancies should be concentrated in that group.
2. Low risk folate deficient group. Women with the highest stores of folate should be protected. Affected pregnancies should not occur at the top of the distribution. While there is a correlation between folate stores and risk there is no level that provides protection.
3. Rescue. An intervention, giving extra folic acid, should impact women with low folate levels rather than all women.
4. Other exposures. The effect of folic acid should be consistent not conditional on other exposures.
5. Toxicity. High folic acid exposure should never be associated with a higher risk.

Let’s check each against evidence that already exists in the medical literature.

	Would be true	Is true
High risk folate deficient group	Affected pregnancies cluster in women short of folate.	No such group has ever been found.
Low risk folate deficient group	No affected pregnancies.	Even at the highest levels there is still a risk.
Rescue	Folic acid helps only women who are short of folate.	It acts like a drug, rescuing mice that were not short.
Other exposures	No other causes would be identifiable unless they acted through the folate pathway.	Evidence of multiple other causative factors.
Toxicity	More folic acid is never worse.	High levels caused neural tube defects.

High risk folate deficient group

For decades researchers looked for the women with a folate deficiency who ought to have proved the theory. They could not find them. Most mothers of affected pregnancies had plentiful folate.  High risk countries had good diets and every attempt to identify a deficient cohort failed, as a companion article sets out.

Low risk folate deficient group

What emerged instead, eventually was an analysis of an Irish cohort of more than 56,000 pregnancies. Daly et al showed a correlation. Risk fell steadily as red cell folate stores rose. Affected pregnancies occurred at the very top of the distribution. Three decades on there is still no agreement that the curve has a meaningful at risk cut off. The World Health Organization nonetheless adopted a red cell folate concentration as a population target, while stating that it cannot predict any individual woman’s risk.

This shows that folate deficiency is neither necessary nor sufficient for these defects. There is no discrete deficient subgroup to account for them. It could be evidence that folate modifies risk. It could be evidence that folate stores correlate with a third factor that is causative. It is not evidence of a deficiency disease.

Public Health authorities argue that increasing folate must reduce the risk for every group of women and is therefore justified. Where is the evidence that that is the case?

Rescue

If giving folic acid prevented neural tube defects solely by replacing a dietary deficiency then it should not be possible to produce the defect with a known toxin and still prevent it with folic acid. It is possible, and the experiment has been done. This indicates that folic acid is behaving as a drug not a food.

Moulds on maize produce a type of toxin, a mycotoxin called fumonisins. They inhibit an enzyme, ceramide synthase, which the cell needs to build specialised fats in its outer membrane. Folate receptor alpha sits in a patch of those fats, which holds it in shape and allows it to work. Fumonisin degrades the patch and the receptor stops taking folate into the cell. Folate receptor alpha is critical for getting folate into developing brain cells and preventing neural tube defects. When fumonisin B1 was given to laboratory mice across the two days of neural tube closure 79 percent of exposed fetuses developed a neural tube defect. When the mothers were given folic acid the rate was reduced but not restored to normal. When they were instead given GM1, one of the fats that makes up the patch, folate levels inside the cells recovered and the defects were almost entirely prevented.

Here the initiating cause is known and it is a toxin. Folic acid partly rescues it and an agent treating the actual lesion rescues it better. The mothers were not short of folate in their diet. The toxin created the shortage inside the cells by disabling the receptor, and folic acid was acting here as a drug forcing more folate through a damaged system.

This is not just true in mice. Along the Texas border with Mexico the prevalence of these defects doubled in 1990 and 1991, in the same crop year as fumonisin outbreaks in livestock, with risk tracking tortilla consumption. That is a suggestive parallel to the animal work. Together this evidence indicates not only that folic acid has a drug effect but also that toxins can be the causative factor.

Other exposures

Valproate taken in early pregnancy is associated with a ten to twentyfold increase in risk. No doctor claims that is because of a dietary deficiency. The standard teratogen guidance states that attempts to mitigate with higher-dose folic acid have not helped. Pre-existing diabetes, obesity and early-pregnancy fever are all maternal factors that increase the risk with strong supporting evidence. None of them can be reduced to a shortage of folate in the diet.

Then there are the genetic causes, which are a major component and the least often mentioned. Closure is a mechanical process. A sheet of cells must lengthen, bend and fuse on time and a great many genes play a part in that choreography. Disrupt any of them and the tube fails to close in an animal with entirely normal folate. In humans this appears as the syndromic and chromosomal cases, and as a recurrence risk after one affected pregnancy running many times the population rate. That inheritance is not dietary. Of the 621 affected pregnancies recorded in England in 2022, 57 were recorded with an associated genomic condition.

Some of these defects can be prevented with other nutrients. The curly tail mouse is the classic model of a folate-resistant defect. Folic acid does not prevent it and inositol does, acting through an entirely separate signalling system that controls how fast the cells of the developing tail bud divide. In a folate-responsive animal model inositol is not protective. Mice lacking the gene Grhl-3 develop neural tube defects that cannot be prevented with either. In mice deficient in the iron transporter ferroportin, folate levels were measurably low but folic acid did not prevent the defects and high iron did.

In the American national study of pregnancies conceived after mandatory fortification, taking folic acid supplements was not associated with any further reduction in risk, and neither was dietary folate. The authors interpreted this as fortification having already treated any folate-responsive cases. This does not fit with a model where the problem lies entirely in inadequate dietary folate.

In Bangladesh an estimated seventy million people drink arsenic-contaminated groundwater. A study of 294 cases of spina bifida divided these women into low and high arsenic exposure groups measured by the arsenic in their toenails, which records exposure over the preceding months rather than on a single day. In the low arsenic group folic acid was associated with half the odds of the malformation, an estimate whose confidence interval reached exactly 1.00. In the high arsenic group there was no association at all. The two groups were not formally compared, so the difference between them is suggestive rather than demonstrated, and only about a fifth of these women took folic acid, so the numbers in each group are small. An earlier study by the same group showed the protective association fading as arsenic in drinking water rose. Arsenic and folate interact on the same pathway. Folate supplies methyl groups, the small chemical tags the body uses to switch genes on and off, and the body disposes of arsenic by attaching methyl groups to it. Chick embryos exposed to arsenic develop the defects with demonstrated depletion of that methyl supply.

Toxicity

Five mouse strains carrying mutations that cause neural tube defects were given high folic acid doses across generations. Three of the five were negatively affected. Two showed a higher rate of neural tube defects and the third showed increased loss of the affected embryos before birth. In one, the defect rate rose from 18 percent to 58 percent. In another, which had no defects at all on the ordinary diet, 16 percent were affected on the high dose. How can something which can prevent the defect produce it instead, depending on genotype? A shortage being corrected does not behave like that. Correcting a shortage helps those who are short and does nothing to the rest. It does not reverse direction and cause the disease it was given to prevent.

The same mouse studies showed embryonic loss before closure as an adverse response. Long-term supplementation substantially reduced the survival of the affected embryos. If an embryo is lost there is the possibility that the defect did not appear because the embryo died. When the same possibility was raised in 1997 about a human trial in which nine pregnancy losses accompanied every defect apparently prevented, it was treated as speculation and never properly investigated. The mouse work shows the mechanism is real in animals.

It is quite possible that certain human genotypes are more sensitive to folic acid toxicity than others but this has not been investigated.

What can we conclude?

These defects are clearly heterogeneous not due to one causal mechanism. Some are caused by genetic lesions, drugs and maternal metabolic disease. Importantly, no deficient subgroup has ever been identified and one large study in an already fortified population found no further reduction from supplements or dietary folate.

Prevention by folic acid does not identify the initiating cause which can include toxins. Some experimental defects are folate-resistant and preventable through other pathways. Responsiveness to folic acid depends on genotype and it can prove toxic.

Given this we should be asking:

●       How much disease can folic acid actually prevent?

●       What causes the remainder?

●   What is the consequence of exposing everyone?