Research Of Genetic Susceptibility To Primary Biliary Cirrhosis In China

BackgroundPBC (Primary Biliary Cirrhosis) is a kind of progressive chronicnonsuppurative cholangitis, mediated by disorder of the immune system.Pathological changes of PBC include biliary duct distruction, portal inflammation,and liver necrosis. It may cause intrahepatic bile duct injury and intrahepaticcholestasis, which finally leads to hepatic fibrosis and cirrhosis. The incidence ofPBC increased in recent years due to the lifestyle changes and cognitive rised ofdoctors. It is thought that the morbidity of PBC is affected by environmentalfactors and genetic factors. Many reports showed that there was an obviousfamilial aggregation of PBC. The incidence of PBC remarkably increased amongtwins, and the relatives of PBC patients has a100times higher risk than healthypeople, indicating that gene susceptibility played an important role in thepathogenesis of PBC. Gene susceptibility loci had been widely screened byresearchers worldwide. Polymorphisms of different genes seemed to beassociated with the development of PBC among different populations, such asEuropeans and Asians. Different loci or hepalotypes of these genes wereconfirmed to be associted with PBC susceptibility. The disparity of genes among Europeans and Asians indicated that the results of Western countries could notapply to Chinese population. To fill the blank in this research field, we seek todiscover the genes associated with PBC susceptibility among Chinese population.AimsThe aims are identifying genetic susceptibility to PBC in China andexploring the underlying mechanisms of susceptibility genes contributing toPBC.MethodsCarry out the overwhelming literature search to identify the possible genesthat contributed to PBC susceptibility. Analyze single nucleotide polymorphismsof the genes that involved in autoimmune diseases according to literatures andcarefully screened. Select the genes which may participate in the pathogenesis ofPBC for further research. Collect the blood of PBC patients and healthy controlsfor susceptibility gene locus tests using SNP chips. Mass spectmetry analysis wasused to identify the PBC susceptibility gene phenotype. Multi-layer analysis wasused to analyze the data according to the clinical statistics of the participants.Results(1) The forty-five SNPs were selected in nineteen genes which wereassociated with autoimmune liver diseases and other liver diseases. Those genesalso involved in immune regulation, bile excretion and biliary epithelial injurymechanism. For example, CTLT-4, PTPN22and HLA are involved in T cellproliferation; IRF5, STAT4, IL-12, IFN-gamma and Tyk2are involved in Th1celldifferentiation, MDR1, MDR3, BSEP, MRP2, CYP and AE2are involved in bilesynthesis, transport and excretion.(2) The study subjects comprised134Chinese patients with PBC and314relational, healthy Chinese control subjects. (3) A chi-square test revealed that:1) The frequency of the A variant allele atrs473351was significantly increased in PBC patients compared with controlsubjects [70.0%versus30.4%; P=0.004; odds ratio (OR)5.33;95%CI=1.98–14.36].2) The frequency of the variant allele at rs6748358was significantlyincreased in PBC patients compared with control subjects (40.9%versus26.9%;P=0.009; OR,1.88;95%CI,1.11–3.17).3) The frequency of the G variant alleleat rs738409was significantly higher in PBC patients than that in control subjects(51.1%versus37.5%; P=0.02; OR=1.74;95%CI,1.05–2.89). No evidence ofassociation of other SNPs to PBC was observed.(4) All the loci selected were analyzed in dominant, recessive andco-dominant genetic models, respectively. The rs473351locus of BSEP wasassociated with PBC susceptibility. Its variant of A allele (dominant model, OR=2.063;95%CI,1.254-3.393, P=0.004) was highly associated with PBCsusceptibility. The rs2287618locus was associated with PBC susceptibility. Itsvariant of A allele (dominant model, OR=0.617;95%CI,0.411-0.928; P=0.020)is highly associated with PBC susceptibility. The rs6748358locus of STAT4wasassociated with susceptibility to PBC. Its variant of A allele (dominant model, OR,1.567;95%CI,1.044-2.353, P=0.030) was highly associated with PBCsusceptibility.(5) Based on the genotypic concordance, two SNPs in BSEP gene (rs473351and rs2287618) were detected in linkage disequilibrium (D’>0.9). According tothe MDR analysis, two significant interactions were observed for variant allelesin the following loci: rs738409、 rs2235048、 rs2280714、 rs10181656(STAT4-PNPLA3)(P<0.01) and rs280519、rs473351、rs2280714、rs2461823rs10181656(BSEP-STAT4)(P<0.01).(6)96patients were included to analysis the relationship between clinical indexes such as ALP、GGT、TBIL and BSEP SNPs. Logistic regression analysesrevealed that:1) the frequency of variant allele at rs2287618was significantlyassociated with ALP (P<0.05).2) the frequency of variant allele at rs2287618was significantly decreased in UDCA-responsive PBC patients compared withUDCA-non-responsive PBC patients (38.6%versus53.4%; P=0.021; OR=0.55;95%CI,0.33-0.91).(7) The further analysis of PBC specific antibodies showed that theAMA-positive and negative population has a significant difference between thetwo groups with the rs2287618mutation. At the same time, the positive rate ofSp100antibody significantly increased in individuals carrying the rs2287618riskfactor. We conducted a Wilcoxon rank sum test analysis in21PBC patients and21normal controls (the family members of PBC patients) with respect to themajor cytokines. IL-4, IL-10has a significant difference (P=0.04; P=0.033). Theresults of IL-2, IL-12, and IL-17tests showed that the PBC and control group didnot have statistical significance (P=0.24,0.12,0.07; repectively). Besides, therewas no association between IL-4and BSEP SNPs by Logistic analysis (P>0.05),neither as IL-10and BSEP SNPs.Conclusions(1) We have found the relationship between PBC and some SNPs from theperspective of genetic susceptibility such as rs473351, rs2287618, rs6748358,rs738409, and so on. The results may indicate that the locus participate theprogress of PBC.(2) It is the first time to investigate the relationship between BSEP genepolymorphisms and PBC susceptibility. A variety of statistical models and geneticmodels have demonstrated that the BSEP mutations may be a candidate moleculein the process of PBC occurred. (3) SNPs of rs473351and rs2287618in BSEP gene linked closely. Haplotypeof GG was the risk allele, which indicated that individuals carring GG haplotypemay suffer from PBC easier. Besides, we should pay more attention on minoralleles of BSEP-STAT4and STAT4-PTPN22genes respectively. They may have aclose linkage with PBC.(4) It was showed a significantly higher serum ALP, GGT, and TBIL levelsin the early stage of PBC. ALP-regression analysis showed that rs2287618corelated with it. PBC patients carrying rs2287618risk allele have a significantlyhigher level of ALP than the population carried the protective allele of samegenes. Patients carrying rs2287618protective allele were significantly better thanthose carrying risk alleles to respond to UDCA. The result showed that BSEPplayed an important role in the biliary excretion.(5) It was firstly found that gene polymorphisms of rs2287618have animportant role on the expression of the AMA. The results suggested that positiverates of AMA and Sp100were significantly higher in the groups of BSEPrs2287618mutation.(6) CD4+cells, especially Th1cells, are mainly infiltrating lymphocytes inliver lesion. In this experiment, we first conducted that BSEP genepolymorphisms and IL-4, IL-10expression level by logistic regression analysis;but didn’t find any association between them. It may be due to the small samplesize.