Association Analysis Of MtDNA Haplogroup M/N And STATt4rs3821236to Systemic Lupus Erythematosus Risk In Han Females From Hunan Province

Systemic lupus erythematosus (SLE) is a complex disease influencd by interaction among multigenic and environmental factors. Functional genetic screening method and genome-wide association study (GWAS) were two main methods exploring the susceptibility genes by analyzing the differential genetic polymorphisms between the case-contol groups, and the single nucleotide polymorphism (SNP) was the most important polymorphism. However, the methods presented some problems as conflicting results among different studies, which required repeated validation in independent multicenters, and the functional confirmation biologically. Up to date, a large number of susceptibility genes had been reported in SLE, among which the signal transducer and activator of transcription4(STAT4)gene got so much attention for its contribution to the immune disorders in SLE, however, the mitochondria DNA having great impact on the apoptosis and inflammation did not acquire enough attention. To verify the association of STAT4SNPs with SLE in more independent groups and detect its biological function is an important task. And to discover more mtDNA polymorphisms associated with SLE is another fascinating work.Objective:To detect the association between mtDNA M/N haplogroup, STAT4rs3821236G/A and the risk of SLE in Han females from Hunan Province. And to analyze the expression levels of total STA4、STAT4α、STAT4β3between different STAT4rs3821236genotype carriers.Methods:The mtDNA M/N haplogroup, and STAT4rs3821236genotyping were underwent by using ASA (the alleles specific amplification) and PCR-RFLP (PCR-restriction fragment length polymorphism) in all the subjects. The statistical significance of mtDNA M/N haplogroup and STAT4SNPrs3821236frequency was assessed with SPSS18.0. The expression levels of total STAT4, STAT4a and STAT4β mRNA in all the subjects was detected using PCR qRT-PCR (quantitative real-time).Results:1) Among the868SLE patints, the mtDNA M/N haplogroup maked up429cases (49.4%) of M type and439cases (50.6%) of N type, while in the control group, the composition was456cases (52.4%) and414cases (47.6%) respectively, which was of no significane (P=0.213). But when stratified by the age more than50years, the mtDNA N type displayed higher frequence (59.6%vs41.7%, P=0.042) with a statistical significane. And the mtDNA N haplogroup also predicted risk for the hematologic disorder (P=0.000, OR=2.128), skin rash (P=0.000, OR=1.873), the neuropsychiatric disorder (P=0.000, OR=3.956) and the alopecia (P=0.044, OR=1322).2) The STAT4rs3821236G/A genotyping was successfully performed by PCR-RFLP(Ban Ⅰ).3) Among the475affected patients, the STAT4rs3821236genotyping maked up125cases (26.3%) of AA genotype,238cases (50.1%) of AG genotype and112cases (23.6%) of GG genotype, while in the control group with the composition of105cases (19.2%),249cases (45.4%) and194cases (35.4%) correspondingly, which meaned that the STAT4rs3821236AA genotype and A allele frequency between the SLE and control group was26.3%vs19.2%, and51.4%vs41.9%, respectively, with a good significance (P=8×10-5, P=1.76×10-5). And the STAT4rs3821236A allele up-regulated the risk of lupus nephritis (P=0.026, OR=1.345) and alopecia (P=0.000, OR=1.684).4) The total STAT4and STAT4a mRNA expression levels was up-regulated by1.044-fold and1.103-fold, with STAT4β down-regulated by0.819times in the active SLE patients, when compared to the control group. Howerver, the total STAT4mRNA, STAT4a and STAT4β mRNA were all up-regulated by2.288-fold,1.763-fold,1.981-fold, respectively, in the AA and AG genotype carriers of STAT4rs3821236than the GG genotype group.Conclusion:1) The mtDNA N haplogroup up-regulated the risk for late-onset SLE in Han females form Hunan Province. And the mtDNA N haplogroup also predicted risk for the hematologic disorder, skin rash, the neuropsychiatric disorder and the alopecia.2) STAT4rs3821236A allele and AA alleles were risk factors for SLE in the Han female carriers from Hunan Province, and up-regulated the risk of lupus nephritis and alopecia.