Fine-mapping Study Of The MHC Region For Henoch-sch（?）nlein Purpura In Chinese Han Population

Background Henoch-Sch（?）nlein purpura（HSP）,also known as Ig A vasculitis,is an autoimmune vasculitis characterized by extensive deposition of Ig A1 immune complexes in capillaries,arterioles,and venules.Most of them have a good prognosis and are self-limitations:Children aged 2-10 years are more likely to have the disease,and adults are relatively rare.The annual incidence of Ig A vasculitis in children is about 3-26,7/100.000,and in adults is about 0.8-1.8/100.000,with more men than women.HSP takes non-thrombocytopenic purpura as the main clinical manifestation,which may be combined with joint swelling and pain,joint cavity effusion,gastrointestinal symptoms（abdominal pain,blood in the stool）,kidney involvement（hematuria,proteinuria）,and a small number of patients may involve the heart and lungs and other important organs.According to the site of involvement and clinical manifestations,HSP can be divided into five subtypes:simple,abdominal,articular,renal,and mixed.The current view is that renal involvement is often associated with poor prognosis.The pathogenesis of HSP has not yet been fully elucidated.The higher incidence in Asian populations,the higher risk of first-degree relatives of patients,and the incidence of family clusters all indicate that HSP has a strong genetic background,while pathogenic microorganism infection,mosquitoes Bites,drugs,vaccinations,and food are currently considered to be closely related to induced diseases.The above phenomenon indicates that the disorder of the autoimmune system caused by the joint action of genetic and environmental factors may play an important role in the development of the disease.Human leukocyte antigen（HLA）gene located in the human major histocompatibility complex（MHC）region is currently considered to be one of the genes most closely related to the pathogenesis of HSP.Based on the traditional candidate gene method A series of research results have suggested that multiple HLA alleles are closely related to the pathogenesis of HSP.Due to the limitations of the candidate gene method,in order to find the HLA region susceptibility genes of HSP patients in the Chinese Han population,we conducted a GWAS study in the HSP patients in the Chinese Han population for the first time,and then used the HLA genotype developed by Jia et al.in 2013 to fill in Methods（SNP2HLA）The relationship between HLA alleles and their corresponding amino acid positions in HSP patients and the relationship between HSP patients and the disease was studied by referring to the MHC region database constructed based on the deep sequencing data of 10689 normal Chinese Han population genetic variation in 2016 by the Institute of Dermatology,Anhui Medical University,Lay the foundation for further understanding of the mechanism of HLA gene genetic variation in the occurrence of HSP disease.Object HSP is the most common subtype of primary autoimmune vasculitis in childhood,and it can also develop in adults.Since the disease has a strong genetic background,we refer to the Chinese Han population-specific MHC region database based on the data obtained from the GWAS institute,and use the HLA genotype filling method to fine-locate the MHC region of HSP patients and control populations and further explore the entire The role of MHC regional genetic variation in the pathogenesis of HSP in Chinese Han population.Method With reference to the sample collection standards,we first used the Illumina Infinium Global Screening Array chip to perform genome-wide genotyping of 514Chinese Han population HSP patients and 7186 healthy controls,and after quality control,we extracted expanded MHC region SNPs（Single-nucleotide polymorphism,SNP）locus typing data,using SNP2HLA software,refer to the newly constructed MHC region reference data set based on 10689 cases of Chinese Han population to fill the MHC region with HLA genotypes,and then study HLA alleles,HLA The corresponding amino acid positions of genes and the correlation between SNPs and HSP,and the stepwise regression analysis of HLA alleles and HLA gene corresponding amino acid positions were used to further determine the independent HSP susceptibility sites in the MHC region.For the discovered susceptible sites in the MHC region,we can analyze whether there is a correlation between the different clinical phenotypic characteristics of patients and the susceptible sites through HSP cases with detailed clinical data.Result After genome-wide genotyping and quality control filtering data,we found 17genotyping SNPs in the MHC region（rs9275407,rs9275440,rs9275332,rs9275393,rs9275428,rs9275371,rs9275439,rs9275390,rs9275377,rs9272346,rs9275464,rs660895,rs521539,Rs9275365,rs17427599,rs9269110,rs9275327）and 27 genotyped SNPs in non-MHC regions were associated with HSPs at the level of genome-wide significance(P<5×10^-8).Then we used the newly constructed Chinese Han population MHC reference data set to fill in HLA genotypes and performed quality control and correlation analysis on the data obtained after filling.As a result,we found 143 SNPs and3 HLA amino acid sites（HLA-DRB1 Amino acid Asn120,HLA-DRB1 amino acid Ser120,HLA-DRB1 amino acid Val11）reached the significance level of this study（P<2.21×10-6）,and 3 HLA alleles（HLA-DRB1*04,HLA-DRB1*06,HLA-DRB1*06:02）are infinitely close to the significance level of this study and are related to the onset of HSP.Further stepwise regression analysis of HLA alleles and amino acid sites found 6 independent HLA allele abnormal signals in the entire MHC region（HLA-DRB1*04,HLA-DRB1*16,HLA-DRB1*01,HLA-DRB1*12:02,HLA-DRB1*10,HLA-DRB1*15:02）and three independent abnormal signals of HLA gene amino acid positions（HLA-DRB1 amino acid positions 120,26,96）HSP susceptibility sites signal.In our study,the abnormal sites associated with the onset of HSP were all located in the HLA-DRB1 gene,and no abnormal signals of other HLA genes in the MHC region were found.A total of 216 HSP patients can obtain detailed clinical data.HLA-DRB1*04 and HLA-DRB1 amino acid Asn120 are grouped according to homozygous,heterozygous,and non-carrying patients.Refer to the patient’s age（≤10 years old,>10 years old）,whether the kidney is Involvement,whether the white blood cell count is normal,whether the neutrophil count is increased,whether the lymphocyte count is increased,whether hemoglobin is decreased,whether the platelet count is increased,whether the ASO is increased,whether the complement C3 is decreased,whether the complement C4 is decreased,whether the CRP is Elevated,serum Ig A level,serum total Ig E level,DD dimer,ESR,ESR,and Chi-square test.The results show that patients are homozygous for the HLA-DRB1 amino acid Ser120 amino acid site There was a statistical difference with the abnormal ASO level（P=0.043）,and no other clinical features were found to be statistically different from the above sites.Conclusion This study is the first time to conduct a GWAS study on HSP in a Chinese Han population and use a large sample MHC regional reference data set for the Chinese Han population to conduct a further fine positioning study through HLA genotype filling,and 6 HLAs were found in the entire MHC region.The DRB1 alleles and three HLA-DRB1 amino acid sites have abnormal signals.The research results suggest that the onset of HSP has a high genetic background and is closely related to the HLA-DRB1 gene,which lays a foundation for the elucidation of the mechanism of MHC regional genetic variation in the Chinese Han HSP population The preliminary theoretical basis is established.Our study did not find a significant correlation between HLA-DRB1 related sites and disease severity.