Associations Of Rs3740677 Within GAB2 Gene With LOAD In Chinese Han Population

Objective:Our clinical study aim to investigate the possible associations between the common single nucleotide polymorphism(SNP)rs3740677 within “Growth factor receptor bound protein-2 associated binding protein-2” gene(GAB2)with late onset Alzheimer's disease(LOAD).This SNP is identified that sitting on the special sequence which is targeted by 3'UTR mi RNAs-185.Method: By the large sample study,we recruited 992 patients with LOAD as the case group from Oingdao Municipal Hospital and other several public hospital in Shandong province,1358 healthy people were selected as control group from the examination center of Oingdao Municipal Hospital and other severally cooperative hospitals,elderly Centers.The DNA samples were extracted from peripheral blood of all subjects.Through the "Biological information science" approach,the rs3740677 was screened from the binding-site sequence of 3'UTR mi RNA-185 in its target gene GAB2.The screening progression bases on five mainly criterions which must be all coincided: a,the SNPs sites that was aimed must at the 3'UTR of relative gene;b,the objective SNPs sites at the binding-site of mi RNAs in relative gene through the matching standard according to the typical "Seed region" 7mer-m8 which have been described by Bartel,David P in 2009;c,the relative gene where the objective SNPs sites in must be the target gene of mi RNAs which has different expression in the brains of AD patients;d,the gene where objective SNPs sites in must have been documented that is associated with AD onset;e,the minimum allele frequency(MAF)of SNPs must ?0.05(In our study,the value was identified as 0.233 to the minimum allele frequency T of SNPs rs3740677).Using the Wizard genomic DNA purification kit(Cat.#A1125,Promega,USA)which have been described by Yu,J.T.et al,we extracted the DNA sample from the peripheral blood.Subsequently,we carried out the rs3740677 Single nucleotide polymorphism that in the GAB2 gene through Shanghai Genesky Bio-Tech Co.,Ltd.(http://biotech.geneskies.com/index.html)using the improved multiplex ligase detection reaction(i MLDR)technology,which have been implicated by Chen,Bifeng in 2014,the primers that were designed for rs3740677 were F:CTGTCGTATGGGGCCCCTTGTA;R: GCTTGGGTCCAGTGGAGGAAGA.In addition,allele specific multiplex PCR (Multi-ARMS)technique that has been described by Donohoe et al was used to identify the APOE genotype.The laboratory staffs were all blind to the status of every participant.The Hardy–Weinberg equilibrium between observed genotype distributions and expected was tested by the chi-square test.The characteristic differences between healthy controls and LOAD patients were examined using the the chi-square test(for the gender and Apo E?4 status)or Student t-test(for the age and MMSE score).The distributional distinctions of Genotype and the relative allele between the two groups were analyzing through the chi-square test.We further stratified the differences between LOAD cases and healthy controls basing on the Apo E ?4 status,which were also tested using the chi-square test.Using logistic regression adjusted by age,gender and Apo E ?4 status we further calculated and analyzed the differences in genotype and allele distribution between patients and controls by three differently genetic models,which were individually named dominant,recessive and additive model.The p value,odds ratios(OR)and 95% confidence intervals(95% CI)under every model were all calculated by the logistic regression analysis.The possible interactions between SNP rs3740677 with APOE?4 were also examined by logistic regression that was also adjusted by age,gender.All the statistical analyses were performed by the SPSS 19.0 software.The standard for significant difference is "?<0.05".Results: The T allele was discovered and identified as the lower frequencies allele in both group,comparing the higher allele G.The significant differences between LOAD and healthy control group have been revealed in the allele frequencies(P=0.008)and genotype(P=0.024)by the significance level(??0.05)through the Chi-square test analysis.Additionally,as the expected risk allele T of GAB2 SNP rs3740677,we further discovered a possibly protective role of SNP rs3740677 to the LOAD onset by the OR=0.833 and 95%CI=0.728-0.952.Furthermore,we stratified the two groups(LOAD patients and control subjects)by APOE ?4 status(carrier or non-carrier).Neither in the APOE?4 allele carriers(APOE ?4+)nor in the APOE?4 allele non-carriers(APOE ?4-)subgroup,the significant differences of allele and genotype distributer have been found between the LOAD patients and controls(APOE ?4+:genotype P=0.437,allele P=0.208;APOE ?4-:genotype P=0.149,allele P=0.064).Additionally,we used the Logistic Regression approach to exam the probable distributer differences of SNP rs3740677 genotype between the two groups adjusted with the relevant factor including age at onset or at examination,gender and APOE ?4 status in three separately genetic models.Eventually,the statistic results showed significant differences in two of the three separately genetic models(Dominant: OR = 0.831,95%CI = 0.702–0.983,P = 0.031;Additive: OR = 0.855,95%CI = 0.745–0.983,P=0.027;Recessive: P= 0.252).In addition,we further to test the possible interactions between the SNP rs3740677 genotypes with APOE ?4 allele to LOAD onset respectively to the three genetic models,whereas failed to find any statistic significations.Conclusion: Our clinical study first confirmed the significant associations between the SNP rs3740677 sites in the 3'UTR specially targeted sequence of mi RNAs-185 within the AD relevant GAB2 gene in Chinese northern Han populations,and further proposed a protective effect of SNP rs3740677 to LOAD onset.