Clinical Genetics: Problems in Diagnosis and Counseling

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NUTRITIONAL SUPPLEMENTATION AND PREVENTION OF NEURAL TUBE DEFECTS

Mary J. Seller

Publisher Summary

This chapter discusses nutritional supplementation and prevention of neural tube defects (NTD). In 1976, Smithells, Sheppard, and Schorah published a prospective study of the blood vitamin status of an unselected group of more than 900 women in the first trimester of pregnancy. Their first finding was that there was a significant social class gradient in blood vitamin levels. Their second finding was that the six women who subsequently gave birth to infants with NTD had, in the first trimester, lower serum, red cell folate, white blood cell vitamin C, and riboflavin than women who had children without NTD. Smithells and his colleagues submitted that these findings were compatible with the hypothesis that nutritional deficiencies are significant in the causation of neural tube defects. As a consequence, they planned an intervention study, which was subsequently undertaken in a collaborative way by five centers in the United Kingdom and started in 1976.

In 1976, Smithells, Sheppard and Schorah¹ published a prospective study of the blood vitamin status of an unselected group of over 900 women in the first trimester of pregnancy. Their first finding was that there was a significant social class gradient in blood vitamin levels. Social classes I and II* had significantly higher levels of red cell folate, leucocyte vitamin C, riboflavin and serum vitamin A, and higher (but not significantly so) serum folate, than classes III, IV and V. Their second finding was that the six women who subsequently gave birth to infants with neural tube defects (NTD) had, in the first trimester, lower serum and red cell folate, white blood cell vitamin C and riboflavin than women who had children without NTD. This finding held even when the social classes were taken into account, and despite small numbers the differences were significant for red cell folate and white blood cell vitamin C. Smithells and his colleagues submitted that these findings were compatible with the hypothesis that nutritional deficiencies are significant in the causation of neural tube defects.

As a consequence, they planned an intervention study which was subsequently undertaken in a collaborative way by five centers in the United Kingdom and started in 1976. This work, to be described, has been done jointly by myself at Guy’s Hospital, London; by R.W. Smithells, S. Sheppard and C. Schorah at Leeds University; by N.C. Nevin, Queen’s University, Belfast; by R. Harris and A. Read of Manchester University; and by D. Fielding, Chester Hospital, and has been published.²,³ The incidence of NTD varies geographically within the United Kingdom, from around 2.9/1000 births in the southeast to about 8/1000 births in Northern Ireland. Our study incorporated areas of low, high and medium incidence.

Women who had already had one or more children with NTD, who were contemplating another pregnancy, but were not yet pregnant, were offered periconceptional multivitamin supplementation. Originally a double-blind placebo trial was planned, and indeed, the placebo tablets were already prepared. However, this was rejected independently by both the Leeds and Guy’s Hospitals’ Ethical Committees, the first two centers to be involved in the project, and so a less scientifically satisfactory experimental design was adopted.

The women who agreed to participate all received vitamin tablets, Pregnavite Forte F (Bencard), which in a daily dose of three tablets provides a total of 4000 IU vitamin A, 400 IU vitamin D, 1.5 mg thiamine, 1.5 mg riboflavin, 1 mg pyridoxine, 15 mg nicotinamide, 40 mg ascorbic acid, 480 mg calcium phosphate, 0.36 mg folic acid and ferrous sulphate equivalent to 75.6 mg Fe. They were asked to take them for at least a month before conceiving and for the first eight weeks of their pregnancy. Those who complied with this regime form a fully supplemented group. The women actually took the vitamin tablets for varying lengths of time from the minimum 28 days to many months, according to how long it took them to achieve pregnancy; the mean was 110 days. During the supplementation period, tablets were sent regularly to the women by post at one to three month intervals.

Some women who had agreed to take part conceived before they had taken a full month’s therapy; others were found to have begun supplementation a few days after conception. AU these women were placed in a separate group which was called partially supplemented. The women were assigned to these two groups when they reported to us that their pregnancies were confirmed at around seven to eight weeks’ gestation. At this time the period over which they had been receiving supplementation was calculated.

In the absence of being able to do a properly controlled trial, the eventual control women were those who were at the same risk of producing NTD as the supplemented women, living in the same general area, who conceived within the same month, and as near as possible in age to the supplemented women. All but one center matched control and supplemented individuals one for one; Belfast was not able to do this. The control women were selected prospectively when they applied to our genetic counseling clinics at 8–12 weeks gestation because they wanted to book an amniocentesis. They comprised at risk women who did not participate in the trial because they were not previously known to us, or in a few cases were women who had been approached but had refused to participate. It should be noted that since the controls were picked at this early stage in pregnancy, long before maternal serum alpha fetoprotein screening or amniocentesis took place, no bias could be introduced by selecting those women who were especially likely to have a recurrence of NTD. The only known difference between the supplemented and control women is that early first trimester (before eight weeks) spontaneous abortions are recorded for the former, but are necessarily absent in the latter because of the manner of selection of the controls after eight weeks from amniocentesis candidates.

The pregnancies of all women were followed. If amniocentesis was performed the result was recorded and also the eventual outcome of the pregnancy.

The results are shown in Table 1. They take into account twin pregnancies, and also the fact that a number of pregnancies ended in spontaneous abortion. Although every effort was made to examine the products, this was not possible in every case and in some which were examined there was no recognizable fetus.

TABLE 1

Results of Periconceptional Multivitamin Supplementation with Recurrences of NTD in Parenthesis

There were 300 unsupplemented control women who had 305 babies or fetuses of which 295 were examined. There were 13 NTD, a recurrence of 4.4%. There were 200 fully supplemented women who had 204 babies or fetuses of which 195 were examined and there was one recurrence (0.5%). Among the 50 partially supplemented women (52 babies or fetuses, 51 examined) there were no recurrences. The difference between the fully supplemented and the control women is significant (p<0.01, one tailed, Fisher’s exact method).

The most likely explanation for the results is that vitamin supplementation has somehow prevented NTD. However, it is possible that the results might have been unwittingly biased in some way, so the results are now examined for that possibility.

First, if the results are broken down into the different individual centers (Table 2), it can be seen that there is an excess of controls from Belfast, the high incidence area of the United Kingdom. This might have advantageously biased the number of recurrences in the controls. But this is not so because the recurrences were 2 in 35 in Leeds (5.7%), 5 in 95 in London (5.3%) and 6 in 124 in Belfast (4.8%).

TABLE 2

Number of Women Participating in Periconceptional Multivitamin Supplementation According to Center with Recurrence of NTD in Parenthesis

From this breakdown by center it can also be seen that the one recurrence in the supplemented group was in the southeast of England, the low incidence area of the United Kingdom. While this might be chance, it is possibly significant. It may be that the therapeutic effect of an environmental agent is less in areas where the incidence of NTD is lower.

Second, the recurrence of NTD is known to be higher after two previous affected children than after only one, and a bias might have arisen if there were more of these higher risk women in the controls. When women are separated according to their risk category (Table 3), there is no excess of the former in the control women. Nine percent of fully supplemented women, and 10% of control women had two previous affected offspring.

TABLE 3

Results of Periconceptional Multivitamin Supplementation According to Number of Previous Affected Children with Recurrence of NTD in Parenthesis

It has recently been shown⁴ that not only is there a higher incidence of NTD, but also a higher recurrence among the lower socioeconomic classes. Analysis of the social class of our women (Table 4) shows that the proportion of women in the lower classes (conventionally regarded as IIIM, IV and V) was higher in the control group (64% of all the control women) than in the fully supplemented women (55% of the total). However, the large difference overall in the recurrence of NTD between the supplemented and control women cannot be accounted for by this relatively small excess of women of the lower social classes among the controls. Notwithstanding, vitamin therapy seems beneficial to this group. If the effects of vitamin therapy on the lower social class group of women alone are examined, the recurrence is 11 in 191 (5.7%) and 1 in 110 supplemented women (0.9%), a statistically significant difference (p=0.02>0.01). For women of classes I, II and IHN combined, the respective figures are 2 in 87 controls (2.2%) and 0 in 86 supplemented (0%).

TABLE 4

Results of Periconceptional Multivitamin Supplementation According to Number of Previous Affected Children with Recurrence of NTD in Parenthesis

US=unsupplemented, FS=fully supplemented, PS=partially supplemented

Finally, it is possible that the lack of recurrence in the supplemented group might be accounted for by the fact that fetuses with NTD are being spontaneously aborted. However, the incidence of spontaneous abortion in the supplemented and control groups is similar (Table 5), being 10% and 9.6% respectively. Further, of a total of 13 abortuses examined from supplemented women, none had NTD, while one of 19 from control women had an NTD.

TABLE 5

Numbers of Spontaneous Abortions with Recurrence of NTD in Parenthesis

We have also considered differences in maternal age and previous reproductive history which likewise seem negative. Thus, there is nothing we can think of to explain our results other than that vitamin supplementation has, in some way, prevented the recurrence of NTD, but we acknowledge that we have not yet proven this.

We are currently undertaking a second, identical trial, and the nearly completed results show that it is going exactly the same way.

The lack of a controlled double-blind placebo trial has meant that we cannot be sure that a special group of low-risk women has not selected itself for vitamin therapy. Further, because of the cocktail nature of the vitamin preparation we used, even if the therapy really is effective, it is not known which vitamin or vitamins are beneficial. Consequently, further studies in the field are to be encouraged.

In this connection, a related clinical trial was published earlier this year which has evoked much interest. It was a periconceptional folate supplementation study by Laurence, James, Miller, Tennant and Campbell⁵ in South Wales. I have been asked to examine critically this study. It is vital this is done because this paper entitled Double-blind randomized controlled trial of folate treatment before conception to prevent recurrence of neural tube defects states in its summary, There were no recurrences among those who received supplementation and six among those who did not, this difference is significant (p=0.04). This statement is of enormous potential importance. The results as stated appear compelling and the study seems to be the good scientific trial that our own failed to be. Also, it specifically used a single substance where we used a compound of many, thereby isolating which component could be the true therapeutic