- Email: [email protected]
Periconceptional multivitamin supplementation and multimalformed offspring
Periconceptional Multivitamin Supplementation and Multimalformed Offspring Andrew E. Czeizel, MD, DSc, Erika Medveczky, MD OBJECTIVE: To study the human teratogenic risk of a folic acid– containing multivitamin. METHODS: We evaluated the data set of two Hungarian intervention studies: a randomized double-blind, controlled trial and a two-cohort, controlled study of the same folic acid– containing multivitamin in participants of the Hungarian periconceptional service. RESULTS: Of 2471 supplemented and 2391 unsupplemented women, 18 and 21, respectively, had multiple congenital abnormalities in the randomized, controlled trial. Of 3056 supplemented and unsupplemented pairs in the two-cohort, controlled study, 33 and 32, respectively, were affected with multiple congenital abnormalities. After the combination of two data sets, the number of cases with multiple congenital abnormalities was 51 in the supplemented group and 53 in the unsupplemented group (odds ratio 0.89; 95% confidence interval 0.45, 1.68). In addition, there was no difference in the occurrence of specified multiple congenital abnormality entities or of unidentified multimalformed informative offspring. CONCLUSION: We found no evidence that periconceptional folic acid– containing multivitamin supplementation either prevents or induces multiple congenital abnormalities. (Obstet Gynecol 2003;102:1255– 61. © 2003 by The American College of Obstetricians and Gynecologists.)
When evaluating structural birth defects (congenital abnormalities), it is worth differentiating isolated and multiple congenital abnormalities.1 Isolated congenital abnormality is a morphologic defect that can be traced back to one localized error of morphogenesis.2,3 Multiple congenital abnormality is a developmental disturbance caused by two or more different localized errors in morphogenesis of the same person.1 Multiple congenital abnormalities include three types2,3: syndromes, associations, and random combinations of congenital abnormalities. The Hungarian randomized, double-blind, controlled trial (RCT) of periconceptional folic acid (0.8 mg)– From the Foundation for the Community Control of Hereditary Diseases, Budapest, Hungary.
containing multivitamin supplementation demonstrated a significant reduction in the first occurrence of isolated (ie, nonsyndromic) neural tube defects (odds ratio [OR] 0.13; 95% confidence interval [CI] 0.03, 0.65),4 congenital abnormalities of urinary tract (OR 0.22; 95% CI 0.05, 0.99), mainly obstructive defects, and cardiovascular malformations (OR 0.42; 95% CI 0.19, 0.98), mainly conotruncal defects, including ventricular septal defect (OR 0.29; 95% CI 0.09, 0.97).5,6 In addition, there was a trend in the reduction of isolated limb deficiencies (OR 0.19; 95% CI 0.03, 1.18) in the multivitamin group.6 Subsequent publications confirmed the possible protective effect of periconceptional folic acid– containing multivitamin supplementation for defects of urinary tract,7,8 cardiovascular malformations,9 –11 and limb deficiencies.9,8,12 The recent Hungarian two-cohort, controlled intervention study of periconceptional multivitamin supplementation13 also confirmed the findings of the previous Hungarian randomized trial. All of the above preventive effects concerned isolated congenital abnormalities. However, Shaw et al14 published the result of a case– control study from two California centers based on telephone interviews in 112 case and 195 control mothers. They were compared with women who used or did not use multivitamin supplements containing folic acid in the period 3 months before through 3 months after conception. Women who used multivitamins during this period were observed to have an elevated risk of delivering fetuses or infants with multiple congenital abnormalities (OR 2.6; 95% CI 1.1, 6.2. The adjusted OR was not substantially altered: 2.9; 95% CI 0.8, 10.3). However, as the authors stated, “The observed elevated risk associated with maternal vitamin use is considered to be preliminary and needs to be replicated in other populations.” Thus, the objective of the study reported here was to evaluate the occurrence of Financial Disclosure Roche Pharmaceutical supplied the multivitamins and placebo capsules used in the studies. Dr. Czeizel has received some speakers support from Roche Pharmaceutical.
VOL. 102, NO. 6, DECEMBER 2003 © 2003 by The American College of Obstetricians and Gynecologists. Published by Elsevier.
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total and different multiple congenital abnormality categories after periconceptional folic acid (0.8 mg)– containing multivitamin supplementation in two Hungarian intervention studies.4,13 Practically, multiple congenital abnormalities were defined as two or more congenital abnormalities affecting more than one organ or the combination of one congenital abnormality with two or more minor anomalies both in the study of Shaw et al14 and in our analysis. MATERIALS AND METHODS Both the Hungarian RCT (from February 1, 1984 until April 30, 1991) and the two-cohort controlled study (from May 1, 1993 until April 30, 1996) were based on the participants of the Hungarian periconceptional service,15,16 and they were supplied with the same multivitamin (Elevit Pronatal, Roche, Basel, Switzerland) during the periconceptional period (at least 1 month before and 2 months after conception). Elevit Pronatal contains 12 vitamins (vitamin A 4000 IU, but 6000 IU until the end of 1989, B1 1.6 mg, B2 1.8 mg, nicotinamide 19.0 mg, B6 2.6 mg, calcium pantothenate 10 mg, biotin 0.2 mg, B12 4.0 g, C 100.0 mg, D 500.0 IU, E 15.0 mg, and folic acid 0.8 mg), four minerals (calcium 125 mg, phosphorous 125 mg, magnesium 100 mg, and iron 6.0 mg), and three trace elements (copper 1 mg, manganese 1 mg, and zinc 7.5 mg). The change in the vitamin A content of the supplement was connected with the possible teratogenic effect of vitamin A.17 However, the teratogenic effect of vitamin A can be excluded under the dose of 10,000 IU.18 The placebo-like “trace element” included copper 1 mg, manganese 1 mg, zinc 7.5 mg, and vitamin C 7.5 mg in the RCT. The participants were advised to take a single tablet each day. The method of the RCT, including the “blind” use of one of two kinds of tablets, has been described previously.4 In the two-cohort, controlled study, the recruitment of routine care subjects for an unsupplemented cohort took place at their first visit in the regional antenatal care clinics between the 8th and 12th weeks of gestation. There were two criteria for preliminary recruitment: 1) appropriate matching to a pregnant woman in the supplemented cohort with regard to age (plus or minus 1 birth year), socioeconomic status (number of school grades and/or employment status), and residence, and 2) no multivitamin and/or folic acid was used in the periconceptional period before the first visit. This and the diet were checked on the basis of data obtained by a questionnaire through personal interview. The final recruitment of unsupplemented cohort subjects occurred at the 14th week of gestation with existing pregnancy. For the reduction of matched control pair losses, each
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participant in the supplemented cohort had two unsupplemented subjects. Compliance with the regimen of supplementation was verified 1) verbally in discussion with the women, 2) by evaluating the check marks on the form for the basal body temperature measurement, and 3) by counting unused tablets when boxes were returned. A full course of the supplement was completed when women took the supplement on each day or only 1 day was missing during 1 month before and at least 2 months after conception. Women who received a partial course of the supplement were defined as those who failed for more than one day to take the supplement during the preand/or postconceptional periods (in general, these women failed to take the supplement for only a few days). Women who conceived before or during the first month of multivitamin administration were considered as unsupplemented in the RCT. The rate of congenital abnormalities was calculated with a denominator including the three groups of the so-called informative offspring: 1) antenatally diagnosed malformed fetuses in electively terminated pregnancies, 2) stillborn fetuses (ie, late fetal death after the 28th week of gestation and/or greater than 1000 g of fetus), and 3) live-born infants. Each congenital abnormality had welldefined diagnostic criteria, and isolated and multiple congenital abnormalities were differentiated. The evaluation of congenital abnormalities had three time windows in both the RCT and the two-cohort, controlled study: 1) antenatally diagnosed fetal defects in terminated pregnancies. All available medical records (ultrasound films, fetal pathologic description, etc) were collected to have an accurate diagnosis of fetal defects, 2) at birth in all newborn infants. In Hungary, all deliveries took place in hospitals, and neonates were examined by pediatricians. Autopsy was obligatory stillborn fetuses and infant deaths. Furthermore, antenatal (minimum four) and neonatal ultrasound screening became routine care during the study period. If the completed pregnancy outcome certificate indicated any congenital abnormality, all available medical records (detailed physician’s description, autopsy record, karyotype, etc) were obtained to evaluate as much clinical and pathologic information as possible, and 3) at approximately the 12th month of life, all infants were invited for an examination to the coordinating or countryside centers of the Hungarian periconceptional service, and infants were “blindly” checked by one pediatrician. If families did not take part in this examination of infants after the first invitation, we repeated the invitation. If there was still no response, we contacted the pediatrician of infants studied and obtained the case history, particularly data concerning congenital abnormalities diagnosed after birth, recent
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Table 1. Data Sets and Distribution of Informative Offspring in the Supplemented and Unsupplemented Pregnant Women of Two Intervention Studies: Randomised Controlled Trial and Two-Cohort, Controlled Study Intervention studies, informative offspring RCT Confirmed pregnancies Drop-outs Evaluated pregnancies Noninformative offspring* Informative offspring Malformed fetuses in electively terminated pregnancies Stillborn fetuses Live-born infants Two-cohort, controlled study Recruited pregnancies Drop-outs Evaluated pregnancies Unevaluated offspring† Informative offspring Malformed fetuses in electively terminated pregnancies Stillborn fetuses Liveborn infants
Supplemented
Unsupplemented
2819 (100.0) 26 (0.9) 2793 (100.0) 322 (11.5) 2471 (100.0) 3 (0.1) 11 (0.5) 2457 (99.4)
2683 (100.0) 23 (0.9) 2660 (100.0) 269 (10.1) 2391 (100.0) 13 (0.5) 9 (0.4) 2369 (99.1)
3123 (100.0) 54 (1.7) 3069 (100.0) 13 (0.4) 3056 (100.0) 20 (0.7) 7 (0.2) 3029 (99.1)
3123 (100.0) 47 (1.5) 3076 (100.0) 20 (0.7) 3056 (100.0) 9 (0.3) 8 (0.3) 3039 (99.4)
RCT ⫽ randomized, controlled trial. Data are presented as n (%). * Early fetal death: ectopic pregnancy, miscarriage, induced abortion. † Minus: late miscarriage, unmatched pairs; plus: twins and triplets.
death, serious and/or chronic disorders, etc. Thus, all live-born infants except cases of infant death had a double check (birth and follow-up data concerning congenital abnormality). For the fetal and infant deaths, however, autopsy reports were available. Student t and 2 tests were used for the comparison of demographic data. Odds ratios with their 95% CIs were used to estimate effects of multivitamin supplementation. On the other hand, the Manzel-Haenszel test seemed to be appropriate to check the possible heterogeneity of data sets in the RCT and two-cohort, controlled study.
RESULTS The data sets and the distribution of informative offspring in supplemented and unsupplemented women of the two intervention studies are shown in Table 1. The total numbers of supplemented and unsupplemented women were 5527 and 5447, respectively. Among confounders, all were similar between the two study groups in the RCT (Table 2). Mean birth order was higher, thus the proportion of primiparas was lower, and the mean school grades were lower in the unsupplemented group of the two-cohort, controlled study.
Table 2. Demographic Data of Supplemented and Unsupplemented Pregnant Women in Two Intervention Studies Intervention studies, variables
Supplemented
Unsupplemented
RCT Maternal age (y) Birth order Proportion of primiparous Pregnancy order School grades Two-cohort, controlled study Maternal age (y) Birth order Proportion of primiparous Pregnancy order School grades
n ⫽ 2471 26.9 (3.4) 1.31 (0.80) 2182 (88.3) 1.52 (0.91) 14.4 (2.9) n ⫽ 3056 27.4 (3.9) 1.55 (0.75) 1742 (57.0) 1.91 (1.05) 13.5 (2.3)
n ⫽ 2391 26.9 (3.4) 1.29 (0.80) 2150 (89.9) 1.50 (0.89) 14.5 (2.7) n ⫽ 3056 27.4 (4.0) 1.75 (0.88) 1424 (46.6) 1.90 (1.04) 13.1 (2.2)
Difference t or 21 0.00 0.87 3.27 0.77 1.49
P 1.00 .39 .07 .44 .15
0.00 8.45 66.70 0.33 2.19
1.00 ⬍.01 ⬍.01 .74 .04
Abbreviation as in Table 1. Data are presented as mean (standard deviation) or n (%).
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Table 3. Number and Rate (per 1000) of Informative Offspring With Multiple Congenital Abnormalities Supplemented
Categories of multiple congenital abnormalities, name of congenital abnormalities
Unsupplemented
Two-cohort Two-cohort OR (95% CI) RCT study Total RCT study Total Mantel-Haenszel (n ⫽ 2471) (n ⫽ 3056) (n ⫽ 5527) (n ⫽ 2391) (n ⫽ 3056) (n ⫽ 5447) P
Monogenic syndromes Alagille (Au.D.) Collodion fetus (Au.R.) Exostoses, multiple (Au.D.) Situs inversus viscerum (Au.R.) Treacher-Collins (Au.D.) Subtotal No. Rate Chromosomal syndromes Down (⫹21) Edwards (⫹18) Deletions (18q⫺) Turner (45, X) Sex chromosomes (47XXY, 46XX/92XXXX) Subtotal No. Rate Teratogenic/maternal syndromes Hydantoin Diabetic Subtotal No. Rate Recognizable multiple congenital abnormalityentities GAM* Non-immune hydrops Postural* Subtotal No. Rate Unidentified multiple congenital abnormalities Components 2 3 4 6 7 8 Subtotal No. Rate Total No. Rate
1 1 0 0
0 0 0 1
1 1 0 1
0 0 0 0
0 0 1 1
0 0 1 1
0
1
1
0
0
0
2 0.81
2 0.65
4 0.72
0 0.00
2 0.65
2 0.37
2 0 0 0 0
8 1 2 1 2
10 1 2 1 2
5 0 0 1 0
8 1 0 0 0
13 1 0 1 0
0.76 (0.76, 1.73)
16 2.89
6 2.51
9 2.94
15 2.75
1.06 (0.50, 2.26) 0.87
2 0.81
14 4.58
0 0
1 0
1 0
0 0
0 1
0 1
0 0.00
1 0.33
1 0.18
0 0.00
1 0.33
1 0.18
3 1 4
1 1 2
4 2 6
4 0 6
3 0 2
7 0 8
8 3.24
4 1.31
12 2.17
3 3 0 0 0 0
9 0 0 1 1 1
12 3 0 1 1 1
2 2 1 0 0 0
10 4.18
5 1.64
1.98 (0.31, 1.55) 0.42
1.00 (0.10, 9.62) 1.00
15 2.75
0.78 (0.34, 1.72) 0.53
10 2 3 0 0 0
12 4 4 0 0 0
1.00 (0.53, 2.46)
6 2.43
12 3.93
18 3.26
5 2.09
15 4.91
20 3.67
0.89 (0.45, 1.68) 0.72
18 7.28
33 10.80
51 9.23
21 8.78
32 10.47
53 9.73
0.95 (0.63, 1.42) 0.79
OR ⫽ odds ratio; CI ⫽ confidence interval; Au.D. ⫽ autosomal dominant; Au.R. ⫽ autosomal recessive; GAM ⫽ genital anomalies of the male; other abbreviation as in Table 1. * See definition in text.
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Table 4. Phenotypes of 18 Infants Affected With Unidentified Multiple Congenital Abnormalities Whose Mothers Consumed Multivitamins in the Periconceptional Period Number of congenital abnormalities
RCT
Two-cohort, controlled study
Two
Urethral stenosis ⫹ undescended testis, bilateral Ventricular septal defect ⫹ congenital inguinal hernia, bilateral Congenital hydrocephalus ⫹ hydronephrosis, bilateral
Three in the RCT
Cleft lips, bilateral and cleft palate ⫹ talipes equinovarus, bilateral ⫹ minor: spina bifida occulta Oesophageal atresia ⫹ duodenal atresia with annular pancreas ⫹ anal atresia Malrotation of right kidney ⫹ congenital inguinal hernia, right ⫹ undescended testis, right
Six–eight in the two-cohort study
Pyelon duplex, left ⫹ congenital inguinal hernia Short limbs ⫹ hydronephrosis, bilateral Renal agenesis, right ⫹ undescended testis, left Situs inversus abdominal ⫹ complex cardiovascular malformations: common truncus and ventricular septal defect, stenosis of pulmonary and renal arteries Microcephaly ⫹ congenital dislocation of the hip Tracheoesophageal atresia ⫹ ventricular septal defect Poly/syndactyly of hands ⫹ hypospadias, penile Trigonocephaly (needed surgery) ⫹ undescended testis, bilateral Achondroplasia ⫹ renal agenesis, left
Microcephalus ⫹ complex cardiovascular malformation: ventricular septal defect and patent ductus arteriosus ⫹ kidney hypoplasia and ren arcuatus ⫹ diaphragmatic defect: relaxation ⫹ undescended testis, bilateral, ⫹ minors: lowseat ear, simian crease. Arhinencephalus ⫹ cleft lip, bilateral and cleft palate ⫹ microphthalmos, bilateral ⫹ complex cardiovascular malformations: atrial septal defect and ventricular septal defect ⫹ pyelon ureter duplex ⫹ doubling of uterus and vagina ⫹ polydactyly, hands, bilateral, postaxial Complex cardiovascular malformations: atrial septal defect, ventricular septal defect and aortic valve stenosis ⫹ choanal atresia ⫹ oesophageal atresia ⫹ doubling of uterus and vagina ⫹ polydactyly, hands, preaxial ⫹ congenital dislocation of hips, bilateral ⫹ dysmorphic face ⫹ minors: Meckel diverticulum, clinodactyly
Abbreviation as in Table 1.
The numbers and rates of informative offspring affected with multiple congenital abnormalities per 1000 are shown in Table 3. Five categories of multiple congenital abnormalities were differentiated. The first category included mendelian–monogenic syndromes. Each of them occurred only once. Chromosomal syndromes are summarized in the second category. Karyotype analysis was performed in most malformed fetuses and all surviving multiple congenital abnormality cases. Of 16 supplemented and 15 unsupplemented women, 10 (57.1%) and 13 (75.0%) had Down syndrome. Only one case was diagnosed as teratogenic syndrome, such as fetal hydan-
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toin syndrome, and as maternal syndrome, such as diabetic embryopathy. The fourth category included the so-called recognizable multiple congenital abnormalityentities without known etiology. The genital anomalies of the male include at least two congenital abnormalities of male genital organs from the following three: hypospadias, undescended testis, and congenital inguinal hernia with or without minor anomalies of genital organ as phimosis, hydrocele, etc.19 Postural associations represent the combinations of two or more postural deformities as congenital dislocation of the hip, different manifestations of clubfoot, torticollis, and other deformities
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without nonpostural abnormalities.20 There was no difference in the occurrence of recognizable multiple congenital abnormality-entities between supplemented and unsupplemented groups. Finally, the fifth category of multimalformed subjects comprised the unidentified multiple congenital abnormalities differentiating according to the number of component congenital abnormalities. The total number of multiple congenital abnormality cases, and each category within them, did not show difference between supplemented and unsupplemented groups. Mantel-Haenszel test did not indicate a significant heterogeneity of multiple congenital abnormality cases between the RCT and the two-cohort, controlled study; thus it is possible to evaluate them together. The list of component congenital abnormalities in 18 infants affected with unidentified multiple congenital abnormalities whose mothers consumed multivitamins in the periconceptional period is shown in Table 4. Some informative offspring had the combination of congenital abnormalities that were similar to previously delineated multiple congenital abnormality-entities. For example, a multimalformed boy who died in infancy was affected with eight component congenital abnormalities in the two-cohort, controlled study and seemed to be CHARGE (coloboma, heart defect, atresia choanae, retarded growth and mental development, genital anomalies, ear anomalies) association. However, the lack of coloboma and ear anomalies, in addition to the existence of preaxial polydactyly in hands, refused this “registrydiagnosis.” There was a cluster of urinary tract defects among component congenital abnormalities because these congenital abnormalities were recorded in eight multiple congenital abnormalities, whereas complex cardiovascular malformations and undescended testis were found in 4-4 multiple – congenital abnormality cases. Poly/syndactyly, esophageal atresia, and congenital inguinal hernia in 3-3 multiple congenital abnormality cases. However, a specific pattern of component congenital abnormalities within multiple congenital abnormalities was not recognized. DISCUSSION The objective of this analysis was to evaluate the occurrence of multiple congenital abnormalities after periconceptional folic acid– containing multivitamin supplementation in the pooled materials of two Hungarian intervention studies. Our findings did not indicate either a protective or a triggering effect of this treatment for multiple congenital abnormalities. The strength of study designs of these two intervention studies lies in their prospective approach, whereby the same multivitamin was used in a racially homoge-
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neous European–Caucasian population. The periconceptional use of the multivitamin was achieved effectively in both intervention studies, and there was a low proportion of drop-outs. Moreover, the diet of women recruited was checked, and it corresponded with that of the Hungarian population.21 Finally, the diagnosis of well-defined congenital abnormalities had a double check: medically recorded and personally examined, or based on autopsy record. However, some methodologic limitations were found at the evaluation of the studies. Most participants in the RCT were primiparous, with low rates of comorbidity. Many women with previous unsuccessful pregnancy outcomes and maternal disorders took part in the Hungarian periconceptional service, because they expected a higher level of medical care. For ethical reasons, it was not possible to make any discrimination among voluntary participants in the Hungarian periconceptional service. These factors might explain the higher number of chromosomal syndromes and unidentified multiple congenital abnormalities in the two-cohort, controlled study compared with the RCT. Although the sample sizes of our studies appear adequate, the rates of multiple congenital abnormality groups are based on small numbers. Hence, larger sample sizes are necessary for a final conclusion. There was no difference in the rate of total multiple congenital abnormalities between the supplemented and unsupplemented groups; thus our data do not confirm the finding of Shaw et al14 concerning a higher occurrence of multiple congenital abnormalities after periconceptional intake of multivitamin supplements. The definition of multiple congenital abnormality was same in the Hungarian and US studies, but the study design was different. The Hungarian RCT and two-cohort, controlled study were intervention studies, the dropout was very low, all participants used the same multivitamin, and the diagnosis of congenital abnormalities was controlled personally. The US observational study had a prospective case– control approach, approximately 75% of eligible mothers were evaluated, different multivitamins were used, and the congenital abnormality diagnosis was based on the report to the California Birth Defects Monitoring Program. In addition, the Hungarian and Northern Californian populations might differ substantially on the basis of racial origin, diet (eg, American populations currently have a tendency to indulge in herbal supplements), and lifestyle. Nevertheless, the major conclusion was similar in both countries: We did not find multimalformed informative offspring with recognizable patterns of component congenital abnormalities. In general, environmental factors induce specific multiple congenital abnormality patterns, because nearly all
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teratogens cause well-delineated syndromes (as rubella, alcohol, hydantoin, etc). On the other hand, periconceptional folic acid– containing multivitamin supplementation indicated a protective effect for the occurrence of isolated neural tube defects, obstructive defects of urinary tract, and cardiovascular malformations, particularly ventricular septal defects in both Hungarian intervention studies.4 – 6,13 However, a protective effect was not found for these congenital abnormalities as component congenital abnormalities (particularly urinary tract defects) of multiple congenital abnormalities in these studies. It is understandable, because isolated and multiple congenital abnormalities have different etiologies.1 The rate of offspring with Down syndrome was also similar in the supplemented and unsupplemented groups, though recent publications have shown a possible association between polymorphism in genes involved with folate metabolism and maternal risk for Down syndrome.22,23 In conclusion, we found no evidence that periconceptional folic acid– containing multivitamin supplementation either prevents or induces multiple congenital abnormalities. REFERENCES 1. Czeizel AE, Telegdi L, Tusnady G. Multiple congenital abnormalities. Budapest, Hungary: Akade´miai Kiado´, 1988. 2. Spranger J, Benirschke K, Pinsky L, Schwarzacher HG, Smith DW. Errors of morphogenesis: Conceptional terms. J Pediatr 1982;100:160–5. 3. Opitz JM, Czeizel AE, Evans JA, Hall JG, Lubinsky MS, Spranger JW. Nosologic grouping of birth defects. In: Vogel F, Sperling K, eds. Human genetics. Berlin: Springer Verlag, 1987:382–5. 4. Czeizel AE, Duda´s I. Prevention of the first occurrence of neural-tube defects by periconceptional vitamin supplementation. N Engl J Med 1992;327:1832–5. 5. Czeizel AE. Prevention of congenital abnormalities by periconceptional multivitamin supplementation. Br Med J 1993;306:1645–8. 6. Czeizel AE. Reduction of urinary tract and cardiovascular defects by periconceptional multivitamin supplementation. Am J Med Genet 1986;62:179–83. 7. Li DK, Daling JR, Mueller B, Hickok DE, Fantel AG, Weiss NS. Periconceptional multivitamin use in relation to the risk of congenital urinary tract anomalies. Epidemiology 1995;6:212–8. 8. Werler MW, Hayes C, Louik C. Multivitamin use and risk of birth defects. Am J Epidemiol 1999;150:675–82. 9. Shaw GM, O’Malley CD, Wasserman CR, Tolarova HM, Lammer EJ. Maternal periconceptional use of multivita-
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mins and reduced risk for conotruncal heart defects and limb deficiencies among offspring. Am J Med Genet 1995; 59:536–45. Botto LD, Khoury MJ, Mulinare J, Erickson JD. Periconceptional multivitamin use and the occurrence of conotruncal heart defects. Results from a population-based case-control study. Am J Med Genet 1995;59:536–45. Botto LD, Mulinare J, Erickson JD. Occurrence of congenital heart defects in relation to maternal multivitamin use. Am J Epidemiol 2000;151:878–84. Yang Q, Khoury MJ, Olney RS, Mulinare J. Does periconceptional multivitamin use reduce the risk for limb deficiency in offspring? Epidemiology 1997;8:157–61. Czeizel AE. Folic acid containing multivitamin and primary prevention of birth defects. In: Bendich A, Deckelbaum RJ, eds. Preventive Nutrition, 3rd ed. Totowa: Humana Press (in press). Shaw GM, Croen LA, Todoroff K, Tolarova MM. Periconceptional intake of vitamin supplements and risk of multiple congenital anomalies. Am J Med Genet 2000;93: 188–93. Czeizel AE, Dobo´ M, Duda´s I, Gasztonyi Z, Lantos I. The Hungarian periconceptional service as a model for community genetics. Community Genet 1998;1:252–9. Czeizel AE. Ten years of experience in periconceptional care. Eur J Obstet Gynecol Reprod Biol 1999;84:43–9. Rothman KJ, Moore LL, Singer MR, Nguyen US, Mannino S, Milunsky A. Teratogenicity of high vitamin A intake. N Engl J Med 1995;346:393–6. Czeizel AE, Rockenbauer M. Prevention of congenital abnormalities by vitamin A. Int J Vit Nutr Res 1998;68: 219–31. Czeizel AE. Genital anomialies of males: GAM-complex. Eur J Pediatr 1987;146:181–3. Pazonyi I, Kun A, Czeizel AE. Congenital postural deformity association. Acta Paediatr Acad Sci Hung 1982;23: 431–45. Czeizel AE, Susza´nszky E. Diet intake and vitamin supplement use of Hungarian women during the periconceptional period. Int J Vitam Nutr Res 1994;64:300–5. James SJ. Abnormal folate metabolism and mutation in the methylenetetrahydrofolate reductase gene may be maternal risk factors for Down syndrome. Am J Clin Nutr 1999;70:495–501. Hobbs CA. Polymomorphisms in genes involved in folate metabolism as maternal risk factors for Down syndrome. Am J Hum Genet 2000;67:623–30.
Address reprint requests to: Andrew E. Czeizel, MD, DSc, To¨ro¨kve´sz lejto¨ 32, 1026 Budapest, Hungary; E-mail: [email protected]. Received October 3, 2002. Received in revised form May 14, 2003. Accepted June 5, 2003.
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When evaluating structural birth defects (congenital abnormalities), it is worth differentiating isolated and multiple congenital abnormalities.1 Isolated congenital abnormality is a morphologic defect that can be traced back to one localized error of morphogenesis.2,3 Multiple congenital abnormality is a developmental disturbance caused by two or more different localized errors in morphogenesis of the same person.1 Multiple congenital abnormalities include three types2,3: syndromes, associations, and random combinations of congenital abnormalities. The Hungarian randomized, double-blind, controlled trial (RCT) of periconceptional folic acid (0.8 mg)– From the Foundation for the Community Control of Hereditary Diseases, Budapest, Hungary.
containing multivitamin supplementation demonstrated a significant reduction in the first occurrence of isolated (ie, nonsyndromic) neural tube defects (odds ratio [OR] 0.13; 95% confidence interval [CI] 0.03, 0.65),4 congenital abnormalities of urinary tract (OR 0.22; 95% CI 0.05, 0.99), mainly obstructive defects, and cardiovascular malformations (OR 0.42; 95% CI 0.19, 0.98), mainly conotruncal defects, including ventricular septal defect (OR 0.29; 95% CI 0.09, 0.97).5,6 In addition, there was a trend in the reduction of isolated limb deficiencies (OR 0.19; 95% CI 0.03, 1.18) in the multivitamin group.6 Subsequent publications confirmed the possible protective effect of periconceptional folic acid– containing multivitamin supplementation for defects of urinary tract,7,8 cardiovascular malformations,9 –11 and limb deficiencies.9,8,12 The recent Hungarian two-cohort, controlled intervention study of periconceptional multivitamin supplementation13 also confirmed the findings of the previous Hungarian randomized trial. All of the above preventive effects concerned isolated congenital abnormalities. However, Shaw et al14 published the result of a case– control study from two California centers based on telephone interviews in 112 case and 195 control mothers. They were compared with women who used or did not use multivitamin supplements containing folic acid in the period 3 months before through 3 months after conception. Women who used multivitamins during this period were observed to have an elevated risk of delivering fetuses or infants with multiple congenital abnormalities (OR 2.6; 95% CI 1.1, 6.2. The adjusted OR was not substantially altered: 2.9; 95% CI 0.8, 10.3). However, as the authors stated, “The observed elevated risk associated with maternal vitamin use is considered to be preliminary and needs to be replicated in other populations.” Thus, the objective of the study reported here was to evaluate the occurrence of Financial Disclosure Roche Pharmaceutical supplied the multivitamins and placebo capsules used in the studies. Dr. Czeizel has received some speakers support from Roche Pharmaceutical.
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total and different multiple congenital abnormality categories after periconceptional folic acid (0.8 mg)– containing multivitamin supplementation in two Hungarian intervention studies.4,13 Practically, multiple congenital abnormalities were defined as two or more congenital abnormalities affecting more than one organ or the combination of one congenital abnormality with two or more minor anomalies both in the study of Shaw et al14 and in our analysis. MATERIALS AND METHODS Both the Hungarian RCT (from February 1, 1984 until April 30, 1991) and the two-cohort controlled study (from May 1, 1993 until April 30, 1996) were based on the participants of the Hungarian periconceptional service,15,16 and they were supplied with the same multivitamin (Elevit Pronatal, Roche, Basel, Switzerland) during the periconceptional period (at least 1 month before and 2 months after conception). Elevit Pronatal contains 12 vitamins (vitamin A 4000 IU, but 6000 IU until the end of 1989, B1 1.6 mg, B2 1.8 mg, nicotinamide 19.0 mg, B6 2.6 mg, calcium pantothenate 10 mg, biotin 0.2 mg, B12 4.0 g, C 100.0 mg, D 500.0 IU, E 15.0 mg, and folic acid 0.8 mg), four minerals (calcium 125 mg, phosphorous 125 mg, magnesium 100 mg, and iron 6.0 mg), and three trace elements (copper 1 mg, manganese 1 mg, and zinc 7.5 mg). The change in the vitamin A content of the supplement was connected with the possible teratogenic effect of vitamin A.17 However, the teratogenic effect of vitamin A can be excluded under the dose of 10,000 IU.18 The placebo-like “trace element” included copper 1 mg, manganese 1 mg, zinc 7.5 mg, and vitamin C 7.5 mg in the RCT. The participants were advised to take a single tablet each day. The method of the RCT, including the “blind” use of one of two kinds of tablets, has been described previously.4 In the two-cohort, controlled study, the recruitment of routine care subjects for an unsupplemented cohort took place at their first visit in the regional antenatal care clinics between the 8th and 12th weeks of gestation. There were two criteria for preliminary recruitment: 1) appropriate matching to a pregnant woman in the supplemented cohort with regard to age (plus or minus 1 birth year), socioeconomic status (number of school grades and/or employment status), and residence, and 2) no multivitamin and/or folic acid was used in the periconceptional period before the first visit. This and the diet were checked on the basis of data obtained by a questionnaire through personal interview. The final recruitment of unsupplemented cohort subjects occurred at the 14th week of gestation with existing pregnancy. For the reduction of matched control pair losses, each
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participant in the supplemented cohort had two unsupplemented subjects. Compliance with the regimen of supplementation was verified 1) verbally in discussion with the women, 2) by evaluating the check marks on the form for the basal body temperature measurement, and 3) by counting unused tablets when boxes were returned. A full course of the supplement was completed when women took the supplement on each day or only 1 day was missing during 1 month before and at least 2 months after conception. Women who received a partial course of the supplement were defined as those who failed for more than one day to take the supplement during the preand/or postconceptional periods (in general, these women failed to take the supplement for only a few days). Women who conceived before or during the first month of multivitamin administration were considered as unsupplemented in the RCT. The rate of congenital abnormalities was calculated with a denominator including the three groups of the so-called informative offspring: 1) antenatally diagnosed malformed fetuses in electively terminated pregnancies, 2) stillborn fetuses (ie, late fetal death after the 28th week of gestation and/or greater than 1000 g of fetus), and 3) live-born infants. Each congenital abnormality had welldefined diagnostic criteria, and isolated and multiple congenital abnormalities were differentiated. The evaluation of congenital abnormalities had three time windows in both the RCT and the two-cohort, controlled study: 1) antenatally diagnosed fetal defects in terminated pregnancies. All available medical records (ultrasound films, fetal pathologic description, etc) were collected to have an accurate diagnosis of fetal defects, 2) at birth in all newborn infants. In Hungary, all deliveries took place in hospitals, and neonates were examined by pediatricians. Autopsy was obligatory stillborn fetuses and infant deaths. Furthermore, antenatal (minimum four) and neonatal ultrasound screening became routine care during the study period. If the completed pregnancy outcome certificate indicated any congenital abnormality, all available medical records (detailed physician’s description, autopsy record, karyotype, etc) were obtained to evaluate as much clinical and pathologic information as possible, and 3) at approximately the 12th month of life, all infants were invited for an examination to the coordinating or countryside centers of the Hungarian periconceptional service, and infants were “blindly” checked by one pediatrician. If families did not take part in this examination of infants after the first invitation, we repeated the invitation. If there was still no response, we contacted the pediatrician of infants studied and obtained the case history, particularly data concerning congenital abnormalities diagnosed after birth, recent
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Table 1. Data Sets and Distribution of Informative Offspring in the Supplemented and Unsupplemented Pregnant Women of Two Intervention Studies: Randomised Controlled Trial and Two-Cohort, Controlled Study Intervention studies, informative offspring RCT Confirmed pregnancies Drop-outs Evaluated pregnancies Noninformative offspring* Informative offspring Malformed fetuses in electively terminated pregnancies Stillborn fetuses Live-born infants Two-cohort, controlled study Recruited pregnancies Drop-outs Evaluated pregnancies Unevaluated offspring† Informative offspring Malformed fetuses in electively terminated pregnancies Stillborn fetuses Liveborn infants
Supplemented
Unsupplemented
2819 (100.0) 26 (0.9) 2793 (100.0) 322 (11.5) 2471 (100.0) 3 (0.1) 11 (0.5) 2457 (99.4)
2683 (100.0) 23 (0.9) 2660 (100.0) 269 (10.1) 2391 (100.0) 13 (0.5) 9 (0.4) 2369 (99.1)
3123 (100.0) 54 (1.7) 3069 (100.0) 13 (0.4) 3056 (100.0) 20 (0.7) 7 (0.2) 3029 (99.1)
3123 (100.0) 47 (1.5) 3076 (100.0) 20 (0.7) 3056 (100.0) 9 (0.3) 8 (0.3) 3039 (99.4)
RCT ⫽ randomized, controlled trial. Data are presented as n (%). * Early fetal death: ectopic pregnancy, miscarriage, induced abortion. † Minus: late miscarriage, unmatched pairs; plus: twins and triplets.
death, serious and/or chronic disorders, etc. Thus, all live-born infants except cases of infant death had a double check (birth and follow-up data concerning congenital abnormality). For the fetal and infant deaths, however, autopsy reports were available. Student t and 2 tests were used for the comparison of demographic data. Odds ratios with their 95% CIs were used to estimate effects of multivitamin supplementation. On the other hand, the Manzel-Haenszel test seemed to be appropriate to check the possible heterogeneity of data sets in the RCT and two-cohort, controlled study.
RESULTS The data sets and the distribution of informative offspring in supplemented and unsupplemented women of the two intervention studies are shown in Table 1. The total numbers of supplemented and unsupplemented women were 5527 and 5447, respectively. Among confounders, all were similar between the two study groups in the RCT (Table 2). Mean birth order was higher, thus the proportion of primiparas was lower, and the mean school grades were lower in the unsupplemented group of the two-cohort, controlled study.
Table 2. Demographic Data of Supplemented and Unsupplemented Pregnant Women in Two Intervention Studies Intervention studies, variables
Supplemented
Unsupplemented
RCT Maternal age (y) Birth order Proportion of primiparous Pregnancy order School grades Two-cohort, controlled study Maternal age (y) Birth order Proportion of primiparous Pregnancy order School grades
n ⫽ 2471 26.9 (3.4) 1.31 (0.80) 2182 (88.3) 1.52 (0.91) 14.4 (2.9) n ⫽ 3056 27.4 (3.9) 1.55 (0.75) 1742 (57.0) 1.91 (1.05) 13.5 (2.3)
n ⫽ 2391 26.9 (3.4) 1.29 (0.80) 2150 (89.9) 1.50 (0.89) 14.5 (2.7) n ⫽ 3056 27.4 (4.0) 1.75 (0.88) 1424 (46.6) 1.90 (1.04) 13.1 (2.2)
Difference t or 21 0.00 0.87 3.27 0.77 1.49
P 1.00 .39 .07 .44 .15
0.00 8.45 66.70 0.33 2.19
1.00 ⬍.01 ⬍.01 .74 .04
Abbreviation as in Table 1. Data are presented as mean (standard deviation) or n (%).
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Table 3. Number and Rate (per 1000) of Informative Offspring With Multiple Congenital Abnormalities Supplemented
Categories of multiple congenital abnormalities, name of congenital abnormalities
Unsupplemented
Two-cohort Two-cohort OR (95% CI) RCT study Total RCT study Total Mantel-Haenszel (n ⫽ 2471) (n ⫽ 3056) (n ⫽ 5527) (n ⫽ 2391) (n ⫽ 3056) (n ⫽ 5447) P
Monogenic syndromes Alagille (Au.D.) Collodion fetus (Au.R.) Exostoses, multiple (Au.D.) Situs inversus viscerum (Au.R.) Treacher-Collins (Au.D.) Subtotal No. Rate Chromosomal syndromes Down (⫹21) Edwards (⫹18) Deletions (18q⫺) Turner (45, X) Sex chromosomes (47XXY, 46XX/92XXXX) Subtotal No. Rate Teratogenic/maternal syndromes Hydantoin Diabetic Subtotal No. Rate Recognizable multiple congenital abnormalityentities GAM* Non-immune hydrops Postural* Subtotal No. Rate Unidentified multiple congenital abnormalities Components 2 3 4 6 7 8 Subtotal No. Rate Total No. Rate
1 1 0 0
0 0 0 1
1 1 0 1
0 0 0 0
0 0 1 1
0 0 1 1
0
1
1
0
0
0
2 0.81
2 0.65
4 0.72
0 0.00
2 0.65
2 0.37
2 0 0 0 0
8 1 2 1 2
10 1 2 1 2
5 0 0 1 0
8 1 0 0 0
13 1 0 1 0
0.76 (0.76, 1.73)
16 2.89
6 2.51
9 2.94
15 2.75
1.06 (0.50, 2.26) 0.87
2 0.81
14 4.58
0 0
1 0
1 0
0 0
0 1
0 1
0 0.00
1 0.33
1 0.18
0 0.00
1 0.33
1 0.18
3 1 4
1 1 2
4 2 6
4 0 6
3 0 2
7 0 8
8 3.24
4 1.31
12 2.17
3 3 0 0 0 0
9 0 0 1 1 1
12 3 0 1 1 1
2 2 1 0 0 0
10 4.18
5 1.64
1.98 (0.31, 1.55) 0.42
1.00 (0.10, 9.62) 1.00
15 2.75
0.78 (0.34, 1.72) 0.53
10 2 3 0 0 0
12 4 4 0 0 0
1.00 (0.53, 2.46)
6 2.43
12 3.93
18 3.26
5 2.09
15 4.91
20 3.67
0.89 (0.45, 1.68) 0.72
18 7.28
33 10.80
51 9.23
21 8.78
32 10.47
53 9.73
0.95 (0.63, 1.42) 0.79
OR ⫽ odds ratio; CI ⫽ confidence interval; Au.D. ⫽ autosomal dominant; Au.R. ⫽ autosomal recessive; GAM ⫽ genital anomalies of the male; other abbreviation as in Table 1. * See definition in text.
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Table 4. Phenotypes of 18 Infants Affected With Unidentified Multiple Congenital Abnormalities Whose Mothers Consumed Multivitamins in the Periconceptional Period Number of congenital abnormalities
RCT
Two-cohort, controlled study
Two
Urethral stenosis ⫹ undescended testis, bilateral Ventricular septal defect ⫹ congenital inguinal hernia, bilateral Congenital hydrocephalus ⫹ hydronephrosis, bilateral
Three in the RCT
Cleft lips, bilateral and cleft palate ⫹ talipes equinovarus, bilateral ⫹ minor: spina bifida occulta Oesophageal atresia ⫹ duodenal atresia with annular pancreas ⫹ anal atresia Malrotation of right kidney ⫹ congenital inguinal hernia, right ⫹ undescended testis, right
Six–eight in the two-cohort study
Pyelon duplex, left ⫹ congenital inguinal hernia Short limbs ⫹ hydronephrosis, bilateral Renal agenesis, right ⫹ undescended testis, left Situs inversus abdominal ⫹ complex cardiovascular malformations: common truncus and ventricular septal defect, stenosis of pulmonary and renal arteries Microcephaly ⫹ congenital dislocation of the hip Tracheoesophageal atresia ⫹ ventricular septal defect Poly/syndactyly of hands ⫹ hypospadias, penile Trigonocephaly (needed surgery) ⫹ undescended testis, bilateral Achondroplasia ⫹ renal agenesis, left
Microcephalus ⫹ complex cardiovascular malformation: ventricular septal defect and patent ductus arteriosus ⫹ kidney hypoplasia and ren arcuatus ⫹ diaphragmatic defect: relaxation ⫹ undescended testis, bilateral, ⫹ minors: lowseat ear, simian crease. Arhinencephalus ⫹ cleft lip, bilateral and cleft palate ⫹ microphthalmos, bilateral ⫹ complex cardiovascular malformations: atrial septal defect and ventricular septal defect ⫹ pyelon ureter duplex ⫹ doubling of uterus and vagina ⫹ polydactyly, hands, bilateral, postaxial Complex cardiovascular malformations: atrial septal defect, ventricular septal defect and aortic valve stenosis ⫹ choanal atresia ⫹ oesophageal atresia ⫹ doubling of uterus and vagina ⫹ polydactyly, hands, preaxial ⫹ congenital dislocation of hips, bilateral ⫹ dysmorphic face ⫹ minors: Meckel diverticulum, clinodactyly
Abbreviation as in Table 1.
The numbers and rates of informative offspring affected with multiple congenital abnormalities per 1000 are shown in Table 3. Five categories of multiple congenital abnormalities were differentiated. The first category included mendelian–monogenic syndromes. Each of them occurred only once. Chromosomal syndromes are summarized in the second category. Karyotype analysis was performed in most malformed fetuses and all surviving multiple congenital abnormality cases. Of 16 supplemented and 15 unsupplemented women, 10 (57.1%) and 13 (75.0%) had Down syndrome. Only one case was diagnosed as teratogenic syndrome, such as fetal hydan-
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toin syndrome, and as maternal syndrome, such as diabetic embryopathy. The fourth category included the so-called recognizable multiple congenital abnormalityentities without known etiology. The genital anomalies of the male include at least two congenital abnormalities of male genital organs from the following three: hypospadias, undescended testis, and congenital inguinal hernia with or without minor anomalies of genital organ as phimosis, hydrocele, etc.19 Postural associations represent the combinations of two or more postural deformities as congenital dislocation of the hip, different manifestations of clubfoot, torticollis, and other deformities
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without nonpostural abnormalities.20 There was no difference in the occurrence of recognizable multiple congenital abnormality-entities between supplemented and unsupplemented groups. Finally, the fifth category of multimalformed subjects comprised the unidentified multiple congenital abnormalities differentiating according to the number of component congenital abnormalities. The total number of multiple congenital abnormality cases, and each category within them, did not show difference between supplemented and unsupplemented groups. Mantel-Haenszel test did not indicate a significant heterogeneity of multiple congenital abnormality cases between the RCT and the two-cohort, controlled study; thus it is possible to evaluate them together. The list of component congenital abnormalities in 18 infants affected with unidentified multiple congenital abnormalities whose mothers consumed multivitamins in the periconceptional period is shown in Table 4. Some informative offspring had the combination of congenital abnormalities that were similar to previously delineated multiple congenital abnormality-entities. For example, a multimalformed boy who died in infancy was affected with eight component congenital abnormalities in the two-cohort, controlled study and seemed to be CHARGE (coloboma, heart defect, atresia choanae, retarded growth and mental development, genital anomalies, ear anomalies) association. However, the lack of coloboma and ear anomalies, in addition to the existence of preaxial polydactyly in hands, refused this “registrydiagnosis.” There was a cluster of urinary tract defects among component congenital abnormalities because these congenital abnormalities were recorded in eight multiple congenital abnormalities, whereas complex cardiovascular malformations and undescended testis were found in 4-4 multiple – congenital abnormality cases. Poly/syndactyly, esophageal atresia, and congenital inguinal hernia in 3-3 multiple congenital abnormality cases. However, a specific pattern of component congenital abnormalities within multiple congenital abnormalities was not recognized. DISCUSSION The objective of this analysis was to evaluate the occurrence of multiple congenital abnormalities after periconceptional folic acid– containing multivitamin supplementation in the pooled materials of two Hungarian intervention studies. Our findings did not indicate either a protective or a triggering effect of this treatment for multiple congenital abnormalities. The strength of study designs of these two intervention studies lies in their prospective approach, whereby the same multivitamin was used in a racially homoge-
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neous European–Caucasian population. The periconceptional use of the multivitamin was achieved effectively in both intervention studies, and there was a low proportion of drop-outs. Moreover, the diet of women recruited was checked, and it corresponded with that of the Hungarian population.21 Finally, the diagnosis of well-defined congenital abnormalities had a double check: medically recorded and personally examined, or based on autopsy record. However, some methodologic limitations were found at the evaluation of the studies. Most participants in the RCT were primiparous, with low rates of comorbidity. Many women with previous unsuccessful pregnancy outcomes and maternal disorders took part in the Hungarian periconceptional service, because they expected a higher level of medical care. For ethical reasons, it was not possible to make any discrimination among voluntary participants in the Hungarian periconceptional service. These factors might explain the higher number of chromosomal syndromes and unidentified multiple congenital abnormalities in the two-cohort, controlled study compared with the RCT. Although the sample sizes of our studies appear adequate, the rates of multiple congenital abnormality groups are based on small numbers. Hence, larger sample sizes are necessary for a final conclusion. There was no difference in the rate of total multiple congenital abnormalities between the supplemented and unsupplemented groups; thus our data do not confirm the finding of Shaw et al14 concerning a higher occurrence of multiple congenital abnormalities after periconceptional intake of multivitamin supplements. The definition of multiple congenital abnormality was same in the Hungarian and US studies, but the study design was different. The Hungarian RCT and two-cohort, controlled study were intervention studies, the dropout was very low, all participants used the same multivitamin, and the diagnosis of congenital abnormalities was controlled personally. The US observational study had a prospective case– control approach, approximately 75% of eligible mothers were evaluated, different multivitamins were used, and the congenital abnormality diagnosis was based on the report to the California Birth Defects Monitoring Program. In addition, the Hungarian and Northern Californian populations might differ substantially on the basis of racial origin, diet (eg, American populations currently have a tendency to indulge in herbal supplements), and lifestyle. Nevertheless, the major conclusion was similar in both countries: We did not find multimalformed informative offspring with recognizable patterns of component congenital abnormalities. In general, environmental factors induce specific multiple congenital abnormality patterns, because nearly all
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teratogens cause well-delineated syndromes (as rubella, alcohol, hydantoin, etc). On the other hand, periconceptional folic acid– containing multivitamin supplementation indicated a protective effect for the occurrence of isolated neural tube defects, obstructive defects of urinary tract, and cardiovascular malformations, particularly ventricular septal defects in both Hungarian intervention studies.4 – 6,13 However, a protective effect was not found for these congenital abnormalities as component congenital abnormalities (particularly urinary tract defects) of multiple congenital abnormalities in these studies. It is understandable, because isolated and multiple congenital abnormalities have different etiologies.1 The rate of offspring with Down syndrome was also similar in the supplemented and unsupplemented groups, though recent publications have shown a possible association between polymorphism in genes involved with folate metabolism and maternal risk for Down syndrome.22,23 In conclusion, we found no evidence that periconceptional folic acid– containing multivitamin supplementation either prevents or induces multiple congenital abnormalities. REFERENCES 1. Czeizel AE, Telegdi L, Tusnady G. Multiple congenital abnormalities. Budapest, Hungary: Akade´miai Kiado´, 1988. 2. Spranger J, Benirschke K, Pinsky L, Schwarzacher HG, Smith DW. Errors of morphogenesis: Conceptional terms. J Pediatr 1982;100:160–5. 3. Opitz JM, Czeizel AE, Evans JA, Hall JG, Lubinsky MS, Spranger JW. Nosologic grouping of birth defects. In: Vogel F, Sperling K, eds. Human genetics. Berlin: Springer Verlag, 1987:382–5. 4. Czeizel AE, Duda´s I. Prevention of the first occurrence of neural-tube defects by periconceptional vitamin supplementation. N Engl J Med 1992;327:1832–5. 5. Czeizel AE. Prevention of congenital abnormalities by periconceptional multivitamin supplementation. Br Med J 1993;306:1645–8. 6. Czeizel AE. Reduction of urinary tract and cardiovascular defects by periconceptional multivitamin supplementation. Am J Med Genet 1986;62:179–83. 7. Li DK, Daling JR, Mueller B, Hickok DE, Fantel AG, Weiss NS. Periconceptional multivitamin use in relation to the risk of congenital urinary tract anomalies. Epidemiology 1995;6:212–8. 8. Werler MW, Hayes C, Louik C. Multivitamin use and risk of birth defects. Am J Epidemiol 1999;150:675–82. 9. Shaw GM, O’Malley CD, Wasserman CR, Tolarova HM, Lammer EJ. Maternal periconceptional use of multivita-
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mins and reduced risk for conotruncal heart defects and limb deficiencies among offspring. Am J Med Genet 1995; 59:536–45. Botto LD, Khoury MJ, Mulinare J, Erickson JD. Periconceptional multivitamin use and the occurrence of conotruncal heart defects. Results from a population-based case-control study. Am J Med Genet 1995;59:536–45. Botto LD, Mulinare J, Erickson JD. Occurrence of congenital heart defects in relation to maternal multivitamin use. Am J Epidemiol 2000;151:878–84. Yang Q, Khoury MJ, Olney RS, Mulinare J. Does periconceptional multivitamin use reduce the risk for limb deficiency in offspring? Epidemiology 1997;8:157–61. Czeizel AE. Folic acid containing multivitamin and primary prevention of birth defects. In: Bendich A, Deckelbaum RJ, eds. Preventive Nutrition, 3rd ed. Totowa: Humana Press (in press). Shaw GM, Croen LA, Todoroff K, Tolarova MM. Periconceptional intake of vitamin supplements and risk of multiple congenital anomalies. Am J Med Genet 2000;93: 188–93. Czeizel AE, Dobo´ M, Duda´s I, Gasztonyi Z, Lantos I. The Hungarian periconceptional service as a model for community genetics. Community Genet 1998;1:252–9. Czeizel AE. Ten years of experience in periconceptional care. Eur J Obstet Gynecol Reprod Biol 1999;84:43–9. Rothman KJ, Moore LL, Singer MR, Nguyen US, Mannino S, Milunsky A. Teratogenicity of high vitamin A intake. N Engl J Med 1995;346:393–6. Czeizel AE, Rockenbauer M. Prevention of congenital abnormalities by vitamin A. Int J Vit Nutr Res 1998;68: 219–31. Czeizel AE. Genital anomialies of males: GAM-complex. Eur J Pediatr 1987;146:181–3. Pazonyi I, Kun A, Czeizel AE. Congenital postural deformity association. Acta Paediatr Acad Sci Hung 1982;23: 431–45. Czeizel AE, Susza´nszky E. Diet intake and vitamin supplement use of Hungarian women during the periconceptional period. Int J Vitam Nutr Res 1994;64:300–5. James SJ. Abnormal folate metabolism and mutation in the methylenetetrahydrofolate reductase gene may be maternal risk factors for Down syndrome. Am J Clin Nutr 1999;70:495–501. Hobbs CA. Polymomorphisms in genes involved in folate metabolism as maternal risk factors for Down syndrome. Am J Hum Genet 2000;67:623–30.
Address reprint requests to: Andrew E. Czeizel, MD, DSc, To¨ro¨kve´sz lejto¨ 32, 1026 Budapest, Hungary; E-mail: [email protected]. Received October 3, 2002. Received in revised form May 14, 2003. Accepted June 5, 2003.
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