The Construction And Applications Of The SNP Database For Uveitis

Background:Uveitis is an intraocular inflammation leading to blindness worldwide.Although the inflammation can be caused by a variety of factors(infections,systemic diseases,organspecific autoimmune processes,trauma,and primary or secondary ocular neoplasms),the exact pathogenesis of the disease remains unclear.Accumulating evidence indicates that genetic factors are involved in the pathogenesis of a number of uveitis entities.With the advantage of next-generation sequencing(NGS)technologies,many associations between single nucleotide polymorphisms(SNPs)have been identified as susceptibility factors for a number of diseases.To date,several databases of SNPs have been constructed,based on published data for a variety of diseases,such as PESNPdb for Pre-eclampsia(http://bejerano.stanford.edu/pesnpdb)and RAvariome for rheumatoid arthritis(RA)(http://hinv.jp/hinv/rav/).To the best of our knowledge,though some existing publicly available variant databases provide variant associated with different disease(such as LOVD: http://grenada.lumc.nl/LSDB_list/lsdbs and ClinVar: http://www.ncbi.nlm.nih.gov/clinvar),thereis no variant database that specifically addresses uveitis-related genes and SNPs.We therefore found it necessary to construct a specific database for uveitis,thereby facilitating the research on geneticmechanisms involved in uveitis.Vogt-Koyanagi-Harada(VKH)syndrome and Behcet's disease(BD)are two of the most common uveitis entities with high blindness in the Chinese population.VKH syndrome mainly occurs in Asia and is characterized by bilateral granulomatous panuveitis with alopecia,vitiligo,poliosis and deafness.BD is a multi-systemic autoimmune disease and high incidence in Far East and Mediterranean5.The main clinical manifestations of BD were oral ulcer,uveitis,skin lesions and genital ulceration.Although the clinical symptoms and epidemiological characteristics of these two diseases were different,there are some similarities in their genetic background.Recently,some disease-related SNPs have been reported to mediate disease risk through modulating the expression of long non-coding RNAs(LINCRNAs)which are one type of non-coding RNAs regulating gene expression.According to the background information mentioned above,this study mainly focused on the following parts:(1)The construction of a new SNP database for investigations on genetic factors associated with uveitis,UVEOGENE,(2)The applications of the UVEOGENE for investigations on thegenetic mechanisms involved in uveitis.Based on these associations,we investigated the genetic relationship among three widely studied uveitis entities including Behcet's disease(BD),Vogt–Koyanagi–Harada(VKH)disease,and acute anterior uveitis(AAU).Furthermore,"UVEOGENE" can be used as a reliable and informative resource to facilitate the research on geneticmechanisms involved in uveitis.PART ? THE CONSTRUCTION OF A NEW SNP DATABASE FOR INVESTIGATIONS ON GENETIC FACTORS ASSOCIATED WITH UVEITIS,UVEOGENEObjective:To facilitate the research on genetic mechanisms involved in uveitis,we constructed the first SNP database for uveitis,"UVEOGENE." In this database,we provide a reliable set of comprehensive genetic associations for researchers focused on uveitis.Methods:1.Data sources: We collected 289 English-language original articles from the PubMed database(http://www.ncbi.nih.gov/pubmed).In addition,we constructed the criteria of studies included.2.Identification of disease-related genes and pathways in Behcet'sdisease(BD),Vogt–Koyanagi–Harada(VKH)disease,and acute anterior uveitis(AAU): The SNPs judged as "significant" between patients and healthy individuals were considered as disease-related SNPs.To move the related SNPs into a functional pathway,we assigned these SNPs to genes which were considered as disease-related genes through KEGG pathway analysis.Based on these associations,we investigated the genetic relationship among three widely studied uveitis entities including BD,VKH disease,and AAU.3.Comparison with data from other databases focused on autoimmune disease: To investigate differences and overlap concerning disease-related pathways,we compared the disease-related genes and pathways of BD,VKH,and RA groups using KEGG pathway.Results:The mRNA and protein expression of DAB2 in DC from active VKH patients were lower than normal controls.In inactive VKH patients,the expression of DAB2 was increased when compared to the active patients.The promoter methylation level of DAB2 in active VKH patients was marginally upregulated when compared with healthy controls and inactive VKH patients.Conclusions:"UVEOGENE" can be used as a reliable and informative resource to identify similarities as well as differences in the genetic susceptibilityamong uveitis and other autoimmune diseases,thereby facilitating the research on geneticmechanisms involved in uveitis.PART ? THE APPLICATIONS OF THE UVEOGENE FOR INVESTIGATIONS ON THE GENETIC MECHANISMS INVOLVED IN UVEITIS.Objective:The rs12569232 single nucleotide polymorphism(SNP)has been reported associated with both Vogt-Koyanagi-Harada(VKH)syndrome and Behcet's disease(BD)in Chinese population.However,the mechanism underlying the involvement of the SNP in the diseases is unclear.A LINCRNA,LINC00467,was identified by expression quantitative trait loci(eQTL)analysis of the rs12569232 SNP.This study aimed to investigate the role of LINC00467 in VKH syndrome,BD and CD4+T cells.Methods:1.A case-control association study between rs12569232 and the two diseases was validated in 550 patients with VKH syndrome,749 patients with BD and 920 healthy controls using Sequenom MassARRAY system.2.The mRNA expression of LINC00467 was detected by real-time PCR.The protocol and localization of LINC00467 in CD4+T cells was detected by fluorescent in situ hybridization(FISH).3.An adenovirus carrying the LINC00467 was transduced into CD4+T cells from healthy individuals,and the effect on cell viability was measured by the CCK-8 test.4.The human proteome microarray and starBase 2.0 were used to identify the binding proteins of LINC00467 and RNA Immunoprecipitation(RIP)was used to confirm the correlated binding.Results:The GG genotype and G allele of rs12569232 in VKH and BD were increased as compared with normal controls.The normal subjects with GG genotype of rs12569232 showed an increased expression of LINC00467 in PBMCs.The expression of LINC00467 was up-regulated in PBMCs and CD4+T cells from active VKH syndrome and BD patients.The over-expression of LINC00467 could promote the cell viability of CD4+T cells.HUR was confirmed to be the binding proteins of LINC00467 by RIP.Conclusion:The study suggests that the expression of LINC00467 may be correlated with VKH syndrome and BD,and the LINCRNA is modulated by rs12569232 SNP associated with the two diseases.In addition,the LINC00467 probably promotes the viability of CD4+T cells through binding HUR.