Dr. Ángel Carracedo - Simposio Internacional 'La enfermedad de la duda: el TOC'
0505-0513
1. c Indian Academy of Sciences
RESEARCH ARTICLE
Maternal MTHFR polymorphism (677 C–T) and risk of Down’s
syndrome child: meta-analysis
AMANDEEP KAUR and ANUPAM KAUR∗
Department of Human Genetics, Guru Nanak Dev University, Amritsar 143 005, India
Abstract
Methylenetetrahydrofolate reductase (MTHFR) is the most important gene that participates in folate metabolism. Presence
of valine instead of alanine at position 677 and elevated levels of homocystein causes DNA hypomethylation which in turn
favours nondisjunction. In this study, we conducted a meta-analysis to establish link between maternal single-nucleotide
polymorphism (SNP) and birth of Down’s syndrome (DS) child. A total of 37 case–control studies were selected for analysis
including our own, in which we investigated 110 cases and 111 control mothers. Overall, the result of meta-analysis showed
significant risk of DS affected by the presence of maternal SNP (MTHFR 677 C–T OR = 0.816, 95% CI = 0.741–0.900,
P < 0.0001). Heterogeneity of high magnitude was observed among the studies. The chi-square value suggested a highly
significant association between homozygous mutant TT genotype and birth of DS child (χ2 = 23.63, P = 0.000). Genetic
models suggested that ‘T’ allele possesses high risk for DS whether present in dominant (OR = 1.23, 95% CI = 1.13–1.34);
codominant (OR = 1.17, 95% CI = 1.10–1.25) or recessive (OR = 1.21, 95% CI = 1.05–1.38) form. The analysis from all
37 studies combined together suggested that MTHFR 677 C–T is a major risk factor for DS birth.
[Kaur A. and Kaur A. 2016 Maternal MTHFR polymorphism (677 C–T) and risk of Down’s syndrome child: meta-analysis. J. Genet. 95,
505–513]
Introduction
Down’s syndrome (DS) is the most common chromosomal
abnormality that occurs with the prevalence rate of about 1 in
1200 live births (Patel and Adhia 2005), due to nondisjunc-
tion during meiosis I or II. Many studies have reported that
increased homocystein levels and presence of MTHFR 677
C–T polymorphism impairs the folate metabolism and results
in DNA hypomethylation, which in turn favours nondisjunc-
tion of chromosome 21 (James et al. 1999; Fenech 2001).
The results seen in literature are quite contradictory, may be
due to small sample size, making it difficult to elucidate the
role of single-nucleotide polymorphism (SNP) for the risk of
DS child. The present meta-analysis was conducted to under-
stand whether mutation in MTHFR gene is a risk factor for
DS or not.
Materials and methods
Literature regarding DS was searched on electronic PubMed
database and Google scholar from 1999 to January 2015 for
the studies on MTHFR 677 C–T polymorphism (figure 1).
∗For correspondence. E-mail: anupamkaur@yahoo.com.
All the selected reports were case–control studies. The key-
words used were: MTHFR 677 C–T, Down’s syndrome,
methylenetetrahydrofolate reductase and folate metabolism.
A total of 37 studies that evaluated the association between
the presence of MTHFR 677 C–T polymorphism and risk
of DS child were included. Only research article papers
were considred for this study. For each data, information
about author, country and year of publication along with total
number of cases and controls were extracted. Odds ratio, chi-
square values along with frequencies were calculated (table 1).
Inclusion criteria
The following criteria were considered. (i) Articles published
in English language; (ii) articles evaluated MTHFR 677 C–T
SNP; (iii) only those articles in which the mother had at least
two normal children, without miscarriages and abnormali-
ties; and (iv) reports with genotypic and allelic frequencies,
odds ratio and chi-square.
Statistical analysis
The meta-analysis was performed using StatsDirect 3.
Risk for the birth of DS child and presence of maternal
Keywords. MTHFR gene; Down’s syndrome; folate metabolism.
Journal of Genetics, DOI 10.1007/s12041-016-0657-7, Vol. 95, No. 3, September 2016 505
2. Amandeep Kaur and Anupam Kaur
polymorphism was assessed by odds ratio with 95% CI.
Presence of heterogeneity was estimated using chi-square
based Q-statistics and I2
metrics. In case of higher hetero-
geneity (I2
> 50%), random effect model (DerSimonian and
Laird 1986) was used else fixed effect model (Mantel and
Haenszel 1959) was applied. To estimate publication bias,
Begg and Mazumdar rank correlation (Begg and Mazumdar
1994) and Egger’s regression intercept (Egger et al. 1997)
tests were performed.
Results
In this study, we identified 37 eligible reports from a total of
182 articles published between 1999 and 2015 (figure 1). The
reports were from different ethnic populations, i.e. Asians,
Americans, Europeans and others. A total of 3401 moth-
ers having DS children and 5277 mothers having normal
children were included in the study. Meta-analysis indicated
a highly significant difference between cases and controls
(χ2
= 23.63, P = 0.000). The three genetic models (domi-
nant: OR = 1.23, 95% CI = 1.13–1.34; codominant: OR =
1.17, 95% CI = 1.10–1.25; recessive: OR = 1.21, 95% CI
= 1.05–1.38) suggested that presence of ‘TT’ genotype in
mothers significantly increased the risk of DS birth (table 2).
The frequency of heterozygote was highest among mothers
of Down’s syndrome children (MDS) (8.3–65.5%) as com-
pared to control mothers (0–55.3%) and a presence of TT
genotype was observed to be higher among cases (0–36.6%)
than the control mothers (0–21.5%) (table 1).
The present analysis revealed that occurrence of mater-
nal polymorphism 677 C–T significantly increases the risk
of birth of DS child (fixed effect model (OR = 0.816, 95%
CI = 0.741–0.900, P < 0.0001) (figure 2); random effect
model (OR = 0.776, 95% CI = 0.654−0.922, P = 0.003)
(figure 3)). The random model was used since the results
demonstrated high magnitude of heterogeneity among stud-
ies (I2
= 62.6%, P < 0.0001, Q = 96.26, df = 36). Sub-
group analysis revealed that MTHFR 677 C–T also found to
be significantly associated with the birth of DS child among
Asians (random model OR = 0.527, 95% CI = 0.355–0.782,
P = 0.001) (figure 4). Higher heterogeneity was observed
among Asian studies (I2
= 58.9%, Q = 29.17, df = 12,
Total articles searched=182
Mice models=5, Reviews=5, Case reports=7, Newsletters=10
Irrelevant articles, abstracts, foreign language articles,
Insufficient data=118
After exclusion
Articles included in the meta-analysis=37
Figure 1. Flow chart showing selection criteria for meta-analysis.
P = 0.003). Similar results were obtained from American
population suggesting association between 677 C–T SNP
and DS child birth (fixed model OR = 0.658, 95% CI =
0.551–0.786, P = 0.0001) (figure 5). However, heterogene-
ity was found to be of low magnitude (I2
= 39.9%, Q =
11.64, P = 0.113, df = 7). On the other hand, no association
and heterogeneity were found among European studies (fixed
model OR = 0.982, 95% CI = 0.851–1.133, P = 0.838,
I2
= 0%, Q = 8.264, P = 0.764, df = 12) (figure 6).
Publication bias
Publication bias was not observed among the studies as indi-
cated by Begg-Mazumdar bias P = 0.23 and Egger’s bias
P = 0.184 for CC versus CT+TT (figure 7).
Discussion
Literature search of individual reports suggest that the frequency
of MTHFR 677 TT distribution varies greatly worldwide. In
our previous study (Kaur and Kaur 2013), we observed
that 1.8% of cases exhibited ‘TT’ genotype, while con-
trol mothers did not exhibit homozygous mutant genotype.
No significant association was observed among cases and
controls (χ2
− 2.047, P = 0.359) due to the absence of
homozygous mutant allele. Similar findings were observed
in other studies by Cyril et al. (2009), Mohanty et al.
(2012), Divyakolu et al. (2013) and Pandey et al. (2013).
Further, studies from other countries also corroborates with
our study like James et al. (1999), Acacia et al. (2005),
Biselli et al. (2008), Boduroglu et al. (2004), Bozovic et
al. (2011), Brandalize et al. (2009), Chadefaux-Vekeman
et al. (2002), Chango et al. (2005), Coppede et al. (2006,
2009, 2010), Cretu et al. (2010), Da Silva et al. (2005),
El-Gharib et al. (2012), Elsayed et al. (2014), Kokotas et
al. (2009), Martinez-Frias (2008), Meguid et al. (2008),
O’Leary et al. (2002), Pozzi et al. (2009), Santos-Rebaucas
et al. (2008), Scala et al. (2006), Stuppia et al. (2002), Tayeb
(2012), Vranekovic et al. (2010), Liao (2010) and Zampeiri
et al. (2012). The nonsignificant associations in these stud-
ies could be due to small sample size and further a single
polymorphism was not sufficient to establish any association.
James et al. (1999) was the first to report that mutant geno-
type ‘TT’ is associated with the increased risk of DS. In
their study, the frequency of heterozygote was much higher
(59.6%) when compared to controls (44%). It was suggested
that predominance of heterozygote among MDS was due to
foetal viability that may be lower in mothers with the ‘TT’
genotype. Several studies observed nonsignificant associa-
tion but the presence of mutant allele increased the risk of
DS ranging from 0.4–2.7% (table 1). The high intake of
folate may neutralize the metabolic impact of MTHFR poly-
morphism. Thus, SNPs in other genes in folate pathway,
combined with MTHFR and homocystein levels need to be
evaluated to see the overall association.
506 Journal of Genetics, Vol. 95, No. 3, September 2016
4. Amandeep Kaur and Anupam Kaur
Indian reports by Kohli et al. (2008) (χ2
− 6.054, P −
0.048) and Rai et al. (2006) (χ2
− 9.664, P − 0.008) showed
a significant association among cases and controls. The
frequency of TT genotype in a study by Rai et al. (2006) was
8% and showed a 7.13-fold increase risk of DS birth in moth-
ers less than 31 years of age. Similarly, other reports also
Table 2. Genetic models showing odds ratio and chi-square values.
Model Odds ratio 95% CI Chi-square
Dominant (CT/TT versus CC) 1.23 1.13–1.34 22.313, P = 0.000
Codominant (TT/CT versus CT/CC) 1.17 1.10–1.25 22.533, P = 0.000
Recessive (TT versus CC/CT) 1.21 1.05–1.38 7.047, P = 0.008
0.01 0.1 0.2 0.5 1 2 5 10 100
)97.1,44.0(98.0ydutsruo)aidnI(3102ruaKdnaruaK
Elsayed et al. )01.2,72.0(67.0tpygE,)4102(
)70.5,32.0(00.1Divyakolu et al. (2013), India
)56.1,62.0(66.0Pandey et al. (2013), India
)00.1,53.0(06.0Zampeiri et al. (2012), Brazil
Mohanty et al. )35.1,50.0(43.0aidnI,)2102(
)96.2,33.0(49.0)aibarAiduaS(2102beyaT
Gharib et al. (2012), )30.1,41.0(83.0tpygE
)43.1,15.0(38.0Bozovic et al. (2011), Crotia
Sadiq 2011 (Jordan) 0.20 (0.06, 0.62)
Lio et al. (2010), China 0.49 (0.20, 1.17)
)35.1,35.0(09.0Vrenekovic et al. (2010), Crotia
Cretu et al. )73.5,26.0(18.1emoR,)0102(
)71.2,56.0(02.1Pozzi et al. (2009), Danish
)52.1,65.0(48.0Brandalize et al. (2009), Brazil
Coppede et al. )52.1,53.0(66.0ylatI,)9002(
)94.1,77.0(70.1Kokotas et al. (2009), Denmark
Cyril et al. (2009), India 0.08 (0.00, 1.42)
)01.2,66.0(81.1Santos et al. (2008), Brazil
Martinez-Frias et al. (2008), )86.1,76.0(60.1niapS
Biselli et al. )41.1,53.0(36.0lizarB,)8002(
Wang et al. (2007), C )06.0,11.0(62.0anih
0.44 (0.17, 1.12)Meguid et al. (2008), Egypt
Kohli et al. (2008), India 1.37 (0.73, 2.56)
Wang et al. (2007), C )97.0,22.0(24.0anih
Rai et al. (2006), India 0.62 (0.37, 1.03)
Scala et al. (2006), Italy 1.21 (0.70, 2.06)
Liang et al. )90.3,13.0(30.1anihC,)6002(
Acacio et al. )57.0,02.0(93.0lizarB,)5002(
Da Silva et al. )38.81,32.3(83.7lizarB,)5002(
Chango et al. (2005), )14.1,64.0(18.0ecnarF
Boduroglu et al. (2004), Turkey 0.74 (0.42, 1.31)
Chadeaux-Vekemans et al. )80.2,25.0(40.1ecnarF,)2002(
)89.2,86.0(34.1Stuppia et al. (2002), Italy
O’Leary et al. )48.1,24.0(98.0dnalerI,)2002(
Hobbs et al. )68.0,23.0(25.0ASU,)0002(
)49.0,61.0(93.0(1999), USAl.atesemaJ
combined [fixed] 0.82 (0.74, 0.90)
Figure 2. Forest plot (fixed effect) showing significant association between maternal
SNP MTHFR 677 C–T and birth of DS child (CC vs CT+TT).
508 Journal of Genetics, Vol. 95, No. 3, September 2016
5. MTHFR 677 C–T: a meta-analysis
0.01 0.1 0.2 0.5 1 2 5 10 100
0.89 (0.44, 1.79)
0.76 (0.27, 2.10)
)70.5,32.0(00.1
0.66 (0.26, 1.65)
)00.1,53.0(06.0
0.34 (0.05, 1.53)
0.94 (0.33, 2.69)
0.38 (0.14, 1.03)
)43.1,15.0(38.0
0.20 (0.06, 0.62)
0.49 (0.20, 1.17)
)35.1,35.0(09.0
)73.5,26.0(18.1
)71.2,56.0(02.1
)52.1,65.0(48.0
0.66 (0.35, 1.25)
)94.1,77.0(70.1
)24.1,00.0(80.0
)01.2,66.0(81.1
1.06 (0.67, 1.68)
)41.1,53.0(36.0
0.26 (0.11, 0.60)
0.44 (0.17, 1.12)
)65.2,37.0(73.1
0.42 (0.22, 0.79)
0.62 (0.37, 1.03)
)60.2,07.0(12.1
1.03 (0.31, 3.09)
0.39 (0.20, 0.75)
)38.81,32.3(83.7
0.81 (0.46, 1.41)
)13.1,24.0(47.0
1.04 (0.52, 2.08)
)89.2,86.0(34.1
0.89 (0.42, 1.84)
0.52 (0.32, 0.86)
)49.0,61.0(93.0
combined [random] 0.78 (0.65, 0.92)
Kaur and Kaur 2013 (India)our study
Elsayed et al. (2014), Egypt
Divyakolu et al. (2013), India
Pandey et al. (2013), India
Zampeiri et al. (2012), Brazil
Mohanty et al. (2012), India
Tayeb 2012 (Saudi Arabia)
Gharib et al. (2012), Egypt
Bozovic et al. (2011), Crotia
Sadiq 2011 (Jordan)
Lio et al. (2010), China
Vrenekovic et al. (2010), Crotia
Cretu et al. (2010), Rome
Pozzi et al. (2009), Danish
Brandalize et al. (2009), Brazil
Coppede et al. (2009), Italy
Kokotas et al. (2009), Denmark
Cyril et al. (2009), India
Santos et al. (2008), Brazil
Martinez-Frias et al. (2008), Spain
Biselli et al. (2008), Brazil
Wang et al. (2008), China
Meguid et al. (2008), Egypt
Kohli et al. (2008), India
Wang et al. (2007), China
Rai et al. (2006), India
Scala et al. (2006), Italy
Liang et al. (2006), China
Acacio et al. (2005), Brazil
Da Silva et al. (2005), Brazil
Chango et al. (2005), France
Boduroglu et al. (2004), Turkey
Chadeaux-Vekemans et al. (2002), France
Stuppia et al. (2002), Italy
O’Leary et al. (2002), Ireland
Hobbs et al. (2000), USA
(1999), USAl.atesemaJ
Figure 3. Forest plot (random effect) showing significant association between MTHFR 677 C–T and DS child (CC vs CT+TT).
suggested that homozygous mutant (TT) genotype is a risk
factor for DS and its presence increases the risk of DS by
1.4–5.0 folds (Hobbs et al. 2000; Wang et al. 2007, 2008;
Sadiq et al. 2011).
Some Indian and international reports failed to show any
significant association between cases and controls when
analysed individually. However, the genetic models were
best fit for this analysis, demonstrating the presence of
MTHFR 677 C–T polymorphism either in dominant or reces-
sive form, playing a significant role as one of the risk factor
for the birth of DS (table 2). The variation in frequency of
‘T’ allele may be due to ethnicity, lifestyle and availability of
dietary folate that minimizes the effect of MTHFR 677 C–T
polymorphism. Large-scale studies are needed to rule out
the exact mechanism behind the role of homocystein, folate,
vitamin B levels in combination with other polymorphisms
in folate pathway.
Literatures of various meta-analysis suggest significant
association between presence of maternal polymorphism and
birth of DS child. Wu et al. (2014) provided the result on
28 publications that included 2806 cases and 4597 control
mothers for MTHFR 677 C–T analysis. They also performed
Journal of Genetics, Vol. 95, No. 3, September 2016 509
6. Amandeep Kaur and Anupam Kaur
0.01 0.1 0.2 0.5 1 2 5 10
)71.0,00.0(20.0ydutsruo)aidnI(3102ruaKdnaruaK
)70.5,32.0(00.1Divyakolu et al. (2013), India
)56.1,62.0(66.0Pandey et al. (2013), India
Mohanty et al. 0aidnI,)2102( .34 (0.05, 1.53)
)96.2,33.0(49.0)aibarAiduaS(2102beyaT
Sadiq 2011 (Jordan) 0.20 (0.06, 0.62)
Lio et al. 0anihC,)0102( .49 (0.20, 1.17)
)24.1,00.0(80.0Cyril et al. (2009), India
Wang et al. )06.0,11.0(62.0anihC,)8002(
)65.2,37.0(73.1Kohli et al. (2008), India
Wang et al. 0anihC,)7002( .42 (0.22, 0.79)
Liang et al. )90.3,13.0(30.1anihC,)6002(
Rai et al. 0aidnI,)6002( .62 (0.37, 1.03)
combined [random] 0.53 (0.36, 0.78)
Figure 4. Forest plot (random plot) showing significant association between MTHFR
677 C–T and DS child among Asians (CC vs CT+TT).
0.1 0.2 0.5 1 2 5
)00.1,53.0(06.0Zamoeiri et al. (2012), Brazil
)52.1,65.0(48.0Brandalize et al. (2009), Brazil
)01.2,66.0(81.1Santos et al. (2008), Brazil
Biselli et al. (2008), Brazil 0.63 (0.35, 1.14)
Acacio et al. (2005), Brazil 0.39 (0.20, 0.75)
Da Silva et al. (2005), Brazil 0.68 (0.42, 1.09)
Hobbs et al. (2000), USA 0.52 (0.32, 0.86)
James et al. (1999), USA 0.39 (0.16, 0.94)
combined [fixed] 0.66 (0.55, 0.79)
Figure 5. Forest plot (fixed effect) showing significant association between
MTHFR 677 C–T and risk of DS child in American studies (CC vs Ct+TT).
subgroup analysis and revealed significant association. Other
analyses by Medica et al. (2009), Rai et al. (2014) and
Victorino et al. (2014) have suggested similar findings. Yang
et al. (2013) also reported significant association but did not
perform subgroup analysis. In our meta-analysis, a strong
association of MTHFR 677 C–T was observed and we have
also performed subgroup analysis and genetic models to esti-
mate the association. On the other hand, Zintazaras (2007)
did not observe significant involvement of MTHFR 677
C–T polymorphism. Similarly, Costa-Lima et al. (2013)
conducted meta-analysis on 20 publications and reported
moderate relationship for maternal MTHFR 677 C–T using
codominant model only. The difference in the results may be
due to difference in the number of publications included and
various approaches used to demonstrate the association.
Our study have few limitations: we analysed only sin-
gle polymorphism, unadjusted OR in the analysis was used,
no environmental factors were considered, foreign language
and unpublished reports were not included. However, advan-
tages were: there were more number of studies in this meta-
analysis (37 studies), overlapping studies were excluded and
subgroup analysis was also conducted.
In conclusion, our meta-analysis suggests that exhibit-
ing TT genotype significantly increases the risk for DS.
However, this risk varies across different ethnicities and DS
results from the interaction of genetic and environmental
510 Journal of Genetics, Vol. 95, No. 3, September 2016
7. MTHFR 677 C–T: a meta-analysis
0.2 0.5 1 2 5 10
)43.1,15.0(38.0Bozovic et al. (2011), Crotia
Vrenekovic et al. (2010), Crotia 0.90 (0.53, 1.53)
Cretu et al. (2010), Rome 1.81 (0.62, 5.37)
Pozzi et al. (2009), Danish 1.20 (0.65, 2.17)
Coppede et al. (2009), Italy 0.66 (0.35, 1.25)
Kokotas et al. (2009), Denmark 1.07 (0.77, 1.49)
Martinez-Frias et al. (2008), Spain 1.06 (0.67, 1.68)
Scala et al. (2006), Italy 1.21 (0.70, 2.06)
Chango et al. (2005), France 0.81 (0.46, 1.41)
)13.1,24.0(47.0Boduroglu et al. (2004), Turkey
Chadeaux-Vekemans et al. (2002), France 1.04 (0.52, 2.08)
Stuppia et al. (2002), Italy 1.43 (0.68, 2.98)
O’leary et al. (2002), Ireland 0.89 (0.42, 1.84)
combined [fixed] 0.98 (0.85, 1.13)
Figure 6. Forest plot (fixed effect) showing significant association between MTHFR
677 C–T and birth of DS child among European studies (CC versus CT+TT).
Figure 7. Standard error by log odds ratio for dominant model.
factors as well. Considering all such factors in future stu-
dies will lead to better understanding of association between
SNPs and birth of DS child.
Acknowledgements
We gratefully acknowledge the support from DST grant and fellow-
ship (SR/WOS-A/LS-348/2013) awarded to Amandeep Kaur and in
part (UGC) grant no. F.37190/2009 (SR) awarded to Anupam Kaur.
References
Acacia G. L., Barini R., Bertuzzo C., Couto E. C., Annichino-
Bizzachi J. M. and Junior W. P. 2005 Methylenetetrahydrofolate
reductase gene polymorphisms and their association with trisomy
21. Prenat. Diagn. 25, 1196–1199.
Begg C. B. and Mazumdar M. 1994 Operating characteristics of a
rank correlation test for publication bias. Biometrics 50, 1088–
1101.
Biselli J. M., Golonani-Betollo E. C., Zampieri B. L.,
Haddad R., Eberlin M. N. and Pavarino-Bertelli E. C. 2008
Genetics polymorphisms involved in folate metabolism and
elevated plasma concentration of homocystein: maternal risk
factors for Down syndrome in Brazil. Genet. Mol. Res. 7,
33–42.
Boduroglu K., Alanay Y., Koldan B. and Tuncbilek E. 2004
Methylenetetrahydrofolate reductase enzyme polymorphisms as
maternal risk for Down syndrome among Turkish women. Am. J.
Med. Genet. A 127A, 5–10.
Bozovic I. B., Vranekovic J., Cizmarevic N. S., Mahulja-
Stamenkovic V., Prpic I. and Brajenovic-Milic B. 2011 MTHFR
C677T and A1298C polymorphisms as a risk factor for con-
genital heart defects in Down syndrome. Pediatr. Int. 53, 546–
550.
Brandalize A. P., Bandinelli E., Dos Santos P. A., Roisenberg I.
and Schuller-Faccini L. 2009 Evaluation of C677T and A1298C
polymorphisms of the MTHFR gene as maternal risk factor for
Journal of Genetics, Vol. 95, No. 3, September 2016 511
8. Amandeep Kaur and Anupam Kaur
Down syndrome and congenital heart defects. Am. J. Med. Genet.
A 149A, 2080–2087.
Chadefaux-Vekeman B., Caude M., Muller F., Oury J. F., Chabli A.,
Jais J. et al. 2002 Methylenetetrahydrofolate reductase polymor-
phism in the etiology of Down syndrome. Pediatr. Res. 51, 766–
767.
Chango A., Fillon-Emery N., Mircher C., Blehaut H., Lambert D.,
Herbeth B. et al. 2005 No association between common polymor-
phisms in genes of folate and homocystein metabolism and risk
of Down syndrome among French mothers. Br. J. Nutr. 96, 166–
169.
Coppede F., Margini G., Bargagna S., Stuppia L., Minichilli F.,
Fontana I., Coloqnato R. et al. 2006 Folate gene polymorphisms
an risk of Down syndrome pregnancies in young Italian women.
Am. J. Med. Genet. A 140, 1083–1091.
Coppede F., Migheli F., Bargagna S., Siciliano G., Antonucci I.,
Stuppia L. et al. 2009 Association of maternal polymorphisms in
folate metabolizing genes with chromosome damage and risk of
Down syndrome offspring. Neurosci. Lett. 499, 15–19.
Coppede F., Grossi E., Migheli F. and Migliore L. 2010 Poly-
morphisms in folate metabolizing genes, chromosome damage
and risk of Down syndrome in Italian mothers: identification of
key factors using artificial neural networks. BMC Med. Genet.
3, 42.
Costa-Lima M. A., Amorim M. R. and Orioli I. M. 2013 Associa-
tion of methylenetetrahydrofolate reductase gene 677 C>T poly-
morphism and Down syndrome. Mol. Biol. Rep. 40, 2115–2125.
Cretu R., Neagos D., Tutulan-Cunita A., Bohiltea L.C. and Stolian
V. 2010 MTHFR gene polymorphisms and prenatal risk of Down
syndrome. Alexandra Loan Guza. Genet. Mol. Biol. 19, 157–
163.
Cyril C., Rai P., Chandra N., Gopinath P. M. and Satyamoorthy
K. 2009 MTHFR gene variants C677T, A1298C and association
with Down syndrome: a case control study from South India. Ind.
J. Hum. Genet. 15, 60–64.
Da Silva L., Vergani N., Galdieri L. C., Ribeiro P. M., Longhitano
S. B., Brunoni D. et al. 2005 Relationship between polymor-
phisms in genes involved in homocystein metabolism and mater-
nal risk factor for Down syndrome in Brazil. Am. J. Med. Genet.
A 135, 263–267.
DerSimonian R. and Laird N. 1986 Meta-analysis in clinical trials.
Control. Clin. Trials 7, 177–188.
Divyakolu S., Tejaswani Y., Thomas W., Thumoju S., Sreekanth
V. R., Vasavi M. et al. 2013 Evaluation of C677T polymorphism
of the methylenetetrahydrofolate reductase (MTHFR) gene in
various neurological disorders. J. Neurol. Disord. 2, 142.
Egger M., Davey S.G., Schneider M. and Minder C. 1997 Bias in
meta-analysis detected by a simple, graphical test. BMJ 315,
629–634.
El-Gharib M. N., Mahfouz A. E. and Hammoudah S. A. 2012
Maternal MTHFR gene polymorphisms and risk of Down syn-
drome offspring. Int. J. Med. Biomed. Res. 1, 199–205.
Elsayed G. M., Elsayed S. M. and Ezz-Elarab S. S. 2014 Maternal
MTHFR C677T genotype and septal defects in offsprings with
Down syndrome: a pilot study. Egyptian J. Med. Hum. Genet. 15,
39–44.
Fenech M. 2001 The role of folic acid and vitamin B12 in genomic
stability of human cells. Mutat. Res. 475, 56–67.
Hobbs C. A., Sherman S. L., Yi P., Hopkins S. E., Torfs C. P.,
Hine R. J. et al. 2000 Polymorphisms in genes involved in folate
metabolism as maternal risk factors for Down syndrome. Am. J.
Hum. Genet. 67, 623–630.
James S. J., Pogribna M., Pogribny I. P., Melnyk S., Hine R. J.,
Gibson J. B. et al. 1999 Abnormal folate metabolism and muta-
tion in methylenetetrahydrofolate reductase gene may be mater-
nal risk factor for Down syndrome. Am. J. Clin. Nutr. 70, 495–
501.
Kaur A. and Kaur A. 2013 Prevalence of methylenetetrahydrofo-
late reductase 677 C–T polymorphism among mothers of Down
syndrome children. Ind. J. Hum. Genet. 19, 1412–1414.
Kohli U., Arora S., Kabra M., Ramakrishanan I., Gulati S. and
Pandey R. M. 2008 Prevalence of MTHFR C677T polymorphism
in north Indian mothers having babies with Trisomy 21 Down
syndrome. Downs Syndr. Res. Pract. 12, 133–137.
Kokotas H., Grigoriadou M., Mikkelson M., Gannaoulia-Karantana
A. and Petersen M. B. 2009 Investigating the impact of Down
syndrome related common MTHFR 677 C>T polymorphism in
Danish population. Dis. Markers 27, 279–285.
Liang X., Feng Z., Lan-Fang Z., Guo X. X., Xiao G. F. et al. 2005
Analysis of Down syndrome screening and antenatal diagnosis of
3195 cases in the middle period of pregnancy. China J. Modern
Med. 20, 11.
Liao Y. P., Bao M. S., Liu C. Q., Liu H. and Zhang D. 2010 Folate
gene polymorphism and the risk of Down syndrome pregnancies
in young Chinese women. Yi Chuan. 32, 461–466.
Mantel N. and Haenszel W. 1959 Statistical aspects of the analysis
of data from retrospective studies of disease. J. Natl. Cancer Inst.
22, 719–748.
Martinez-Frias M. L. 2008 The biochemical structure and func-
tion of methylenetetrahydrofolate reductase provide the rationale
to interpret the epidemiological results on the risk for infants
with Down syndrome. Am. J. Med. Genet. A 146A, 1477–
1482.
Medica I., Maver A., Augusto G. and Peterlin B. 2009 Polymor-
phisms in genes involved in folate metabolism as maternal risk
factors for Down syndrome-meta analysis. Open Med. 4, 395–
408.
Meguid N. A., Dardir A. A., Khass M., Hossieny L. E., Ezzat A.
and El-Awady M. K. 2008 MTHFR genetic polymorphism as a
risk factor in Egyptian mothers in Down syndrome children. Dis.
Markers 28, 19–26.
Mohanty P. K., Kapoor S., Dubey A. P., Pandey S., Shah R.,
Nayak H. K. et al. 2012 Evaluation of C677T polymorphism
of the methylenetetrahydrofolate reductase gene and its associ-
ation with levels of serum homocystein folate and vitamin B12
as maternal risk factors for Down syndrome. Ind. J. Hum. Genet.
18, 285–289.
O’Leary V. B., Paele-Mcdermott A., Molloy A. M., Kirke P. N.,
Johnson Z., Conley M. et al. 2002 MTRR and MTHFR polymor-
phisms: link to Down syndrome? Am. J. Med. Genet. 10, 388–
390.
Pandey S. K., Mohanty P. K., Polipalli S. K. and Kapoor S. 2013
Genetic polymorphisms of MTHFR (C677T and A1298C) and
homocystein metabolism as a risk factor for Down’s syndrome
patients in north Indian mothers. Int. J. Pharma. Biosci. 4, 249–
256.
Patel Z. M. and Adhia R. A. 2005 Birth defects surveillance study.
Ind. J. Pediatr. 72, 489–491.
Pozzi E., Vergani P., Dalpra L., Combi R., Silvestri D., Crosti
F. et al. 2009 Maternal polymorphism for methylenetetrahydro-
foalte reductase and methionine synthase reductase and risk of
children with Down syndrome. Am. J. Obstet. Gynecol. 200,
e1–e6.
Rai A. K., Singh S., Mehra S., Kumar A., Pandey I. K. and Raman
R. 2006 MTHFR C677T and A1298C polymorphisms are risk
factors for Down syndrome in Indian mothers. J. Hum. Genet.
51, 278–283.
Rai V., Yadav U., Kumar P., Yadav S. K. and Mishra O. P. 2014
Maternal methylenetetrahydrofolate reductase C677T polymor-
phism and Down syndrome risk: a meta-analysis from 34 studies.
PLoS One 9, e108552.
Sadiq M. F., Al-Refai E. A., Al-Nasser A., Khassawneh M. and
Al-Batayneh Q. 2011 Methylenetetrahydrofolate reductase poly-
morphisms C677T and A1298C as maternal risk factors for
512 Journal of Genetics, Vol. 95, No. 3, September 2016
9. MTHFR 677 C–T: a meta-analysis
Down syndrome in Jordan. Genet. Test Mol. Biomarkers 15,
51–57.
Santos-Rebaucas C. B., Correa J. C., Bonomo A., Fintelman-
Rodrigues N., Moura K. C., Rodriguez C. S. et al. 2008 The
impact of folate pathway polymorphisms combined to nutritional
deficiency as maternal predisposition factor for Down syndrome.
Dis. Markers 25, 149–157.
Scala I., Granese B., Sellitto M., Salome S., Sammartino A., Pepe A.
et al. 2006 Analysis of seven maternal polymorphisms of genes
involved in homocystein/folate metabolism and risk of Down
syndrome offspring. Genet. Med. 8, 409–416.
Stuppia L., Gatta V., Gaspari A. R., Antonucci I., Morizio E.,
Calabrese G. et al. 2002 C677T mutation in the 5,10 MTHFR
gene and risk of Down syndrome in Italy. Eur. J. Hum. Genet. 10,
388–390.
Tayeb M. T. 2012 The methylenetetrahydrofolate reductase gene
variant (C677T) in risk mothers with Down syndrome among
Saudi population. Egyptian J. Med. Hum. Genet. 13, 263–
268.
Victorino D. B., Godoy M. F., Goloni-Bertollo E. M. and Pavarino
E. C. 2014 Meta-analysis of methylenetetrahydrofolate reduc-
tase maternal gene in Down syndrome: increased susceptibility
in women carriers of the MTHFR 677T allele. Mol. Biol. Rep. 41,
5491–5504.
Vranekovic J., Babic Bozovic I., Starcevic Cizmarevic N., Buretic-
Tomljanovic A., Ristic S., Petrovic O. et al. 2010 Func-
tional inference of methylenetetrahydrofolate reductase gene
polymorphisms on enzyme stability as a potential risk factor for
Down syndrome in Croatia. Dis. Markers 28, 293–298.
Wang W., Xei W. and Wang X. 2007 The relationship between poly-
morphism of gene involved in folate metabolism, homocystein
level and risk of Down syndrome. Zhonghua Yi Xue Yi Chuan
Xue Za Zhi 24, 533–537.
Wang S. S., Qiao F. Y. and Lv J. J. 2008 Polymorphisms in genes
involved in folate metabolism as maternal risk factors for Down
syndrome in China. J. Zhejiang Univ. Sci. B 9, 93–99.
Wu X., Wang X., Chan Y., Jia S., Luo Y and Tang W. 2014 Folate
metabolism gene polymorphisms MTHFR C677T and A1298C
and risk of for Down syndrome offspring: a meta analysis. Eur.
J. Obstet. Gynec. Rep. Biol. 167, 154–159.
Yang M., Gong T., Lin X., Qi L., Guo Y., Cao Z. et al. 2013
Maternal gene polymorphisms involved in folate metabolism and
the risk of having Down syndrome offspring: a meta analysis.
Mutagenesis 28, 661–671.
Yaping Liao M. P. C. L. 2010 Folate gene polymorphism and the
risk of Down syndrome pregnancies in young Chinese women.
Yi Chuan 32, 461–466.
Zampeiri B. L., Biselli L. M., Goloni-Bertollo E. M., Vannuchi H.,
Carvatho V. M., Cordeiro J. A. et al. 2012 Maternal risk for Down
syndrome is modulated by genes involved in folate metabolism.
Dis. Markers 32, 73–81.
Zintazaras E. 2007 Maternal gene polymorphisms involved in
folate metabolism and risk of Down syndrome offspring: a meta
analysis. J. Hum. Genet. 52, 943–953.
Received 23 June 2015, in revised form 23 September 2015; accepted 24 November 2015
Unedited version published online: 30 November 2015
Final version published online: 4 July 2016
Journal of Genetics, Vol. 95, No. 3, September 2016 513
![c Indian Academy of Sciences
RESEARCH ARTICLE
Maternal MTHFR polymorphism (677 C–T) and risk of Down’s
syndrome child: meta-analysis
AMANDEEP KAUR and ANUPAM KAUR∗
Department of Human Genetics, Guru Nanak Dev University, Amritsar 143 005, India
Abstract
Methylenetetrahydrofolate reductase (MTHFR) is the most important gene that participates in folate metabolism. Presence
of valine instead of alanine at position 677 and elevated levels of homocystein causes DNA hypomethylation which in turn
favours nondisjunction. In this study, we conducted a meta-analysis to establish link between maternal single-nucleotide
polymorphism (SNP) and birth of Down’s syndrome (DS) child. A total of 37 case–control studies were selected for analysis
including our own, in which we investigated 110 cases and 111 control mothers. Overall, the result of meta-analysis showed
significant risk of DS affected by the presence of maternal SNP (MTHFR 677 C–T OR = 0.816, 95% CI = 0.741–0.900,
P < 0.0001). Heterogeneity of high magnitude was observed among the studies. The chi-square value suggested a highly
significant association between homozygous mutant TT genotype and birth of DS child (χ2 = 23.63, P = 0.000). Genetic
models suggested that ‘T’ allele possesses high risk for DS whether present in dominant (OR = 1.23, 95% CI = 1.13–1.34);
codominant (OR = 1.17, 95% CI = 1.10–1.25) or recessive (OR = 1.21, 95% CI = 1.05–1.38) form. The analysis from all
37 studies combined together suggested that MTHFR 677 C–T is a major risk factor for DS birth.
[Kaur A. and Kaur A. 2016 Maternal MTHFR polymorphism (677 C–T) and risk of Down’s syndrome child: meta-analysis. J. Genet. 95,
505–513]
Introduction
Down’s syndrome (DS) is the most common chromosomal
abnormality that occurs with the prevalence rate of about 1 in
1200 live births (Patel and Adhia 2005), due to nondisjunc-
tion during meiosis I or II. Many studies have reported that
increased homocystein levels and presence of MTHFR 677
C–T polymorphism impairs the folate metabolism and results
in DNA hypomethylation, which in turn favours nondisjunc-
tion of chromosome 21 (James et al. 1999; Fenech 2001).
The results seen in literature are quite contradictory, may be
due to small sample size, making it difficult to elucidate the
role of single-nucleotide polymorphism (SNP) for the risk of
DS child. The present meta-analysis was conducted to under-
stand whether mutation in MTHFR gene is a risk factor for
DS or not.
Materials and methods
Literature regarding DS was searched on electronic PubMed
database and Google scholar from 1999 to January 2015 for
the studies on MTHFR 677 C–T polymorphism (figure 1).
∗For correspondence. E-mail: anupamkaur@yahoo.com.
All the selected reports were case–control studies. The key-
words used were: MTHFR 677 C–T, Down’s syndrome,
methylenetetrahydrofolate reductase and folate metabolism.
A total of 37 studies that evaluated the association between
the presence of MTHFR 677 C–T polymorphism and risk
of DS child were included. Only research article papers
were considred for this study. For each data, information
about author, country and year of publication along with total
number of cases and controls were extracted. Odds ratio, chi-
square values along with frequencies were calculated (table 1).
Inclusion criteria
The following criteria were considered. (i) Articles published
in English language; (ii) articles evaluated MTHFR 677 C–T
SNP; (iii) only those articles in which the mother had at least
two normal children, without miscarriages and abnormali-
ties; and (iv) reports with genotypic and allelic frequencies,
odds ratio and chi-square.
Statistical analysis
The meta-analysis was performed using StatsDirect 3.
Risk for the birth of DS child and presence of maternal
Keywords. MTHFR gene; Down’s syndrome; folate metabolism.
Journal of Genetics, DOI 10.1007/s12041-016-0657-7, Vol. 95, No. 3, September 2016 505](https://image.slidesharecdn.com/bed1e901-589c-4070-a1ce-5316c2060c44-170208120949/85/05050513-1-320.jpg)
