The Research Of The Relationship Between MTHFR, ENOS Gene Polymorphism Of Mother As Well As Environmental Factors And NTDs

Objective: The purpose was to find out some suspectable risk factors of neural tubedefects (NTDs)in Guizhou province. Research the relationship between NTDs occuring inthe offspring,methylene-tetrahydrofolate reductase (MTHFR) and endothelial nitricsynthase (eNOS)gene polymorphism of mother as well as environmental factors. At thesame time, explore whether there is interaction between risk factors, in order to provide thecorresponding theoretical basis for preventing the taking place of NTDs effectively.Methods: Monitor the children with NTDs in63hospitals belong to9differentvenues,cities and states of Guizhou province from October of2010to November of2013Or155mothers of induced labor who are diagnosed with NTDs child by ultrasound as casegroup, select155cases of pregnant women as the same gestational age as the of the samein the same period or mothers who give birth to normal children as the control goup,1:1matched case-control study was used. Questionnaire survey were conducted on two groupsof research object. At the same time, collect5ml elbow venous blood on each group,extract DNA. Apply the methods of polymerase chain reaction-restriction fragment lengthpolymorphism (PCR-RFLP) and PCR product sequencing to verify the related genepolymorphisms, detect677locus of MTHFR gene and894and4ab locus of eNOS gene.Use SPSS13.0software to analyze datas, compare gene frequency between case group andcontrol group. At the same time, combine with the environmental impact factors to carryon multifactor conditional logistic regression analysis, analyze interactions betweengene-gene、gene-environment by applying GMDR vo.7software.Results: There are three types of genes on the677locus of MTHFR gene.They are Wildtype CC, homozygous mutant TT type and heterozygous mutations CT, The results of PCRproduct sequencing is consistent with the results of restriction enzyme digestion.Distribution of the genotype frequencies of CC、TT、CT in case group are47.7％、21.3％、31.0％，while in the control group are63.9％,7.1％,29.0％.There is statisticallydifference between case group and control group which confirmed by chi-square test（χ2＝14.69, P＝0.001）；distribution of allele frequencies about (C、T) are64.8％and35.2％, in control group, there are78.4％and21.6％. There is statistically difference between casegroup and control group which confirmed by chi-square tes（tχ2＝13.99, P＝0.000）. Thereare three types of genes on the894locus of eNOS gene: wild GG, heterozygous mutationGT and homozygous mutant TT, the results of PCR product sequencing is consistent withthe results of restriction enzyme digestion. The distribution of the genotype frequencies of（GG、GT、TT）in case group are61.3％、29.7％、9.0％, in the control group are76.1％、20.7％、3.2％, there is statistically difference between case group and control groupwhich confirmed by chi-square test（χ2＝9.26, P＝0.010); the distribution of allelefrequencies about (G、T) in case group are76.1％and23.9％, there are86.5％and13.5％inthe control group, there is statistically difference between case group and control groupwhich confirmed by chi-square test（χ2＝10.86, P＝0.001）.4a/bVNTR of eNOS gene is made up by a allele and b alleles gene, divided into a/a、a/b、b/b three genotypes. In this experiment, there is no a/a genotypes in control group,whilie only one case discovered in the case group. The distribution of the genotypefrequencies about （a/a、a/b、b/b）in case group are0.6％、12.3％、87.1％, there are0.0％、10.3％、98.7％in the control group, there is no statistically difference between casegroup and control group which confirmed by chi-square test（χ2＝0.14, P＝0.704）；thedistribution of allele frequencies about (a、b) in case group are93.2％and6.8％, in controlgroup are94.8％and5.2％, there is no statistically difference between case group andcontrol group which confirmed by chi-square test（χ2＝0.72, P＝0.397）.High performance liouid of chromatography (HPLC) is used to determine HbAlc. Thelevel of HbAlc in case group (5.141.36) is more than the control group（4.830.38）,provinging that there was significant difference in statistics (t=2.09, P＝0.040）.Bring11items variables with statistically difference confirmed by Single factorconditional logistic regression analysis in to Multifactor conditional logistic regressionanalysis, there are6items variables finally can be used in models: passive smoking,cultural degree, edible germination potatoes, hyperemesis gravidarum, MTHFR C677Tmutation and eNOS G894T mutation, their values of OR and95％CI are3.430（1.608～7.317）、0.488（0.320～0.744）、2.017（1.155～3.523）、2.868（1.407～5.849）、2.051（1.258～3.344）、3.650（1.902～7.003）respectively. The results of interaction analysis are showed as follows: type related genes which areconfirmed meaningfully by multiple factors analysis into GMDR v0.7software, analyzeinteraction between gene-gene and gene-environment. K-fold cross-validation in onemodel is consistently10/10, balance accuracy is0.6439, explaining that there areinteraction between MTHFR, eNOS, hyperemesis gravidarum, cultural degree, passivesmoking and eating germination potatoes.Conclusion: MTHFRC677T and eNOSG894T nucleotide gene polymorphism may beassociated with the occurrence of NTDs in Guizhou. The level of HbAlc related to theoccurring of NTDs. Mother with high cultural degree is independent protection factorsabout NTDs happening. There are interaction between MTHFR, eNOS, hyperemesisgravidarum, cultural degree, passive smoking and eating germination potatoes.