Association Study Of X Chromosome Gene Polymorphisms With Susceptibility Of Rheumatoid Arthritis

ObjectiveRheumatoid arthritis(RA)is a systemic autoimmune disease with bone erosion and bone damage as the main manifestations.Its disability and irreversibility bring great pain to patients.The majority of RA patients are women,and genetic variations in the X chromosome gene may be the reason why its prevalence is much higher than that of men.Genomewide association study(GWAS)is used to conduct a comprehensive analysis to find single nucleotide polymorphisms(SNPs)related to the genetic susceptibility of X chromosome genes to RA,and to further study these loci Role in the course of RA.MethodsA case-control design was adopted to collect the research samples of RA GWAS in China.The cases were from the Department of Rheumatology and Immunology of the top three hospitals,and the controls were from the health examination center.A total of 1,027 cases and 2,879 healthy controls were collected.A questionnaire survey was conducted on the cases,including basic information,illness information and clinical examination results,age and gender information of the control group were collected,and Epi Data3.1 software was used for data entry.Collect blood samples of the research subjects,extract their DNA,and further perform genotyping.Samples were screened for quality control through a standardized process.For the genotyping data that met the inclusion criteria,haplotypes from the Thousand Human Genome Three-segment Database were used as reference.SHAPEIT was used to construct the haplotypes,and then IMPUTE2 was used to fill the genotypes.Finally,Odds ratio(OR)and 95%CI(Confidence interval)of individual genetic variation were calculated using the additive effect model.GWAS analysis was performed,and then the GWAS analysis was performed.A meta-analysis of the correlation between X chromosome SNPs and RA susceptibility was carried out by combining the Chinese RA GWAS study and the RA GWAS studies of two other Asian and European populations.The meta-analysis used a fixed-effect model and analyzed with METAL software.P<5.00E-08 was taken as the significance threshold to infer whether there was genetic variation in SNPs,in order to discover new or verify previously discovered SNPs related to RA susceptibility.The plasma was extracted,and the cytokines were detected by Luminex multi-factor liquid phase analysis technology.Linear regression analysis was used to analyze the association between the number of risk alleles of SNPs associated with RA susceptibility and plasma cytokines,and P<0.05 was considered to be statistically significant.Grph Pad Prism 7 was used for mapping to analyze the correlation between the genotype of risk SNPs and plasma cytokines.According to the above GWAS and meta-analysis results,three sites with potential RA-susceptible personality were selected for independent population verification in Anhui,China,and thex~2 test of dominant model,recessive model and additive model were conducted between case and control groups by gender,P<0.05 was considered to be significant.Thex~2 test,linear regression analysis,and the correlation analysis between genotype and clinical and laboratory indicators involved in this project were all carried out by STATA14.In the correlation analysis between genotype and cytokines,clinical and laboratory indicators,P<0.05 was considered statistically significant.ResultsThe Anhui China GWAS analysis included 1027 cases,177 males and 850females,and included 2879 healthy controls(502 males and 2377 females).The meta-analysis results of 3 GWAS studies in China,Asia and Europe did not find new genetic susceptibility sites related to RA,all of which were P>5.00E-08.However,the China GWAS analysis verified that the two genetic risk loci(rs201408742,rs5987194)in the Asian and European populations were related to the genetic susceptibility of the Han population in Anhui,with OR>1 and P<0.05.The number of rs201408742 and rs5987194 alleles in the two SNPs risk alleles and plasma cytokine levels were further analyzed by linear regression,and there was no statistical difference between the number of rs201408742 and rs5987194 alleles and the plasma levels of TNF-α,IL-6,IFN-αand IL-17A(P>0.05).Correlation analysis between rs5987194 and RA specific antibody showed that the G allele frequency of rs5987194 was related to ACPA antibody in RA patients(P<0.05).Three potentially related SNPs(rs151210035,rs9699328,rs9803219)found in GWAS analysis in Anhui,China,were independently verified,and it was found that the three loci may not be correlated with RA susceptibility in Anhui Han population.In addition,we analyzed whether the X chromosome SNPs associated with the susceptibility of Systemic Lupus erythematosus(SLE),another typical autoimmune disease,were also associated with the susceptibility of RA and SLE,found that rs5987194 was associated with the susceptibility of both RA and SLE.The other 5 X chromosome SNPs related to SLE were not related to RA genetic susceptibility[including TLR7(rs3853839),NAA10(rs2071128),PRPS2(rs7062536)].ConclusionsExcept for the two SNPs rs201408742 and rs5987194,the Meta analysis of the GWAS study of the three populations of China,Asia,and Europe did not find that new X chromosome SNPs were associated with RA genetic susceptibility.In addition,we found that the number of alleles at SNP locus rs201408742 and rs5987194 were not correlated with plasma levels of TNF-α,IL-6,IL-17A,and IFN-α.The G allele frequency of rs5987194 was may be related to the ACPA of RA patients.Rs5987194was not only related to the genetic susceptibility of RA,but also related to the genetic susceptibility of SLE,and may play a unique role in the course of RA and SLE.Rs3853839,rs7062536,rs887369,rs5914778,rs13440883,the five X chromosome SNPs related to SLE risk were not related to RA genetic susceptibility.