Association Study Of JARID2 Polymorphisms And Environmental Risk Factors With Nonsyndromic Orofacial Clefts

Background and PurposeNonsyndromic orofacial clefts?NSOC?are the most common human craniofacial malformation in all worldwide populations.There are various complications such as difficulties with feeding,speech,hearing,and psychologic development,and usually require surgical repair were suffered by patients,as well as a large number of other therapies and specialist treatments.A large number of studies suggested that nonsyndromic orofacial clefts?NSOC?result from the complex interplay of both genes and environmental factors.Folic acid deficiency,maternal cigarette smoking,and maternal drinking are a part of the environmental risk factors that have been associated with NSOC.Recently,the jumoji AT-rich interaction domain 2?JARID2?locating on chromosome 6p had been reported to be a novel candidate gene for nonsyndromic cleft lip with or without cleft palate?CL/P?by Luca Scapoli.JARID2is a transcriptional repressor and a member of the JumonjiC?JmjC?and ARID domain protein family,which is essential for early embryonic development.A family based linkage disequilibrium?LD?study in an Italian population confirmed the pivotal role of JARID2 in orofacial development.Three SNPs rs2076056,rs2237138 and rs2299043 in JARID2 were highly significant in Italian families.Thus,two independent evaluations were conducted in our study.First,we evaluated association of JARID2 polymorphisms with NSOC in northern Chinese Han population.Second,association of environmental exposures with risk of NSOC was expolored.MethodsPART ONE:To evaluate association of JARID2 polymorphisms with NSOC in northern Chinese Han populationA case–control study was conducted to examine the association between these three SNPs and NSOC in a northern Chinese Han population.A sample of 299 patients who affected with NSOC and 325 normal individuals without any other serious illness,especially hereditary diseases,and no family history of NSOC was utilized in this study.Genomic DNA was extracted from blood samples.We selected three single-nucleotide polymorphisms?SNPs?including rs2076056,rs2237138 and rs2299043 in JARID2 in this study by the following criteria:prior evidence of association with CL/P and minor allele frequency of the SNP at least 20%in the Chinese population according to HapMap data.Genotyping of the three SNPs were performed using SNaPshot minisequencing technique.Hardy–Weinberg equilibrium of the genotype distributions of controls was estimated.The differences in the genotype and allele frequencies of the three SNPs between cases and controls were evaluated using chi-square test.Genotypic odds ratios?ORs?and 95%confidence intervals?CIs?for genotypes were obtained from logistic regression models.All of above statistical analysis was performed using SAS v9.1.3 statistical software Package?SAS Institute,Inc.?.Haplotype constructions of the three polymorphisms were analyzed using SHEsis online software.D'and r² values were analyzed to evaluate linkage disequilibrium.A P-value of<0.05?two-sided?was used as the criterion of statistical significance,and the odds ratio?OR?>1 was regarded as an increased risk of disease.PART TWO:To explore association of parental environmental exposures and maternal supplementation intake with risk of NSOC.We performed a retrospective hospital-based case-control study that involved 499cases and 480 controls in Heilongjiang Province of China during the period from 2009to 2014.We extracted information of case and control mothers from interviewer-administered questionnaires.The interview was administered by a trained interviewer and addressed exposures from one month before conception through the end of the first trimester.The content of the questionnaire mainly comprised sociodemographic characteristics such as maternal gestational age,gender of infant,parental education,annual household income,gravidity,maternal body mass index?BMI?,history of negative reproduction parental exposures such as maternal history of illnesses,use of medication,lifestyle behaviors such as smoking,alcohol consumption,negative life events,possible toxic occupational exposures,and use of supplements.Single-factor logistic regression analysis was used to compare frequencies of demographic characteristics and potential exposures between CL/P group,CPO group and control group,respectively.Then the significant data were included in multivariable logistic regression model for further analysis to compare these groups.Analyses were performed using SAS v9.1.3.P-values less than or equal to 0.05 were considered to be significant.ResultsPart one:Distribution of rs2237138 genotypes in CL/P?cleft lip with or without palate?group was different from those in the control group?P=0.04?,but significant results did not persist after Benjamini and Hochberg false discovery rate?FDR?correction for multiple tests.Further logistic regression analysis showed that rs2237138 GG genotypes were associated with decreased CL/P susceptibility?OR=0.34,95%CI=0.13–0.84?,compared with the AA wild-type homozygote.For the haplotype CGT,a statistically difference was identified between the CL/P group and controls?P=0.04?.And carriers of GAT haplotype were considered to be less frequent among cleft palate only group as compared to controls?P=0.02?.Part two:The results showed that maternal history of fever and the common cold without fever(OR_CL/P=3.11 and 5.56,95%CI:1.67–5.82 and 2.96–10.47,OR_CPO=3.31 and 8.23,95%CI:1.58–6.94 and 4.08–16.95),paternal smoking and alcohol consumption(OR_CL/P=2.15 and 5.04,95%CI:1.37–3.38 and 3.00–8.46,OR_CPO=1.82 and 4.40,95%CI:1.06–3.13 and 2.50–7.74),maternal exposure to organic solvents,heavy metals,or pesticides(OR_CL/P=6.07,5.67 and 5.97,95%CI:1.49–24.76,1.34–24.09 and 2.10–16.98,OR_CPO=10.65,7.28 and 3.48,95%CI:2.54–44.67,1.41–37.63 and 1.06–11.46)and multivitamin use during the preconception period(OR_CL/P=0.06,95%CI:0.02–0.23,OR_CPO=0.06,95%CI:0.01–0.30)were associated with cleft lip or without cleft palate?CL/P?and cleft palate only?CPO?.Maternal history of skin disease and negative life events(OR_CL/P=12.07 and 1.67,95%CI:1.81–80.05 and 1.95–2.67)were associated with CL/P.ConclusionsFirst,we got a weak association between JARID2 gene polymorphisms and NSOC in both single-marker and haplotype analyses.Our data further strengthen the conclusion that JARID2 polymorphisms are associated with NSOC susceptibility.Second,some potential parental hazardous exposures during the periconception period and maternal use of multivitamins during the preconception period were associated with risk of NSOC.