LncRNAs表达水平及其基因多态性与系统性红斑狼疮的关联性研究

发布时间：2018-05-06 14:43

本文选题：红斑狼疮 + 系统性RNA基因表达多态性　；参考：《安徽医科大学》2017年硕士论文

【摘要】：研究背景系统性红斑狼疮(systemic lupus erythematosus,SLE)是一种慢性多系统性的自身免疫性疾病,其以多种自身抗体产生和大量免疫复合物的沉积为主要特点,并可因此导致体内多种组织或器官的损伤。目前为止,SLE的确切发病机制尚未阐明。过去几十年中的实验和临床研究表明,遗传因素、环境因素、免疫调节因素及其之间的交互作用等共同参与了SLE的疾病进程。通过人类全基因组关联研究(genome-wide association studies,GWAS),发现了多个SLE的易感基因。既往SLE的研究主要集中在编码基因,而非编码基因的报道相对较少。和那些已经被广泛研究的短链非编码RNA(例如micro RNAs)不同的是,长链非编码RNA(long noncoding RNA,lnc RNA)是一类长度超过200个核苷酸的非编码RNA,且lnc RNAs可能在多种生物学功能中具有调节作用。虽然lnc RNAs的确切功能在很大程度上仍然不清楚,但一些研究已经表明lnc RNAs可能参与了多种关键的生物学过程,如染色质重塑、基因转录、RNA剪接和蛋白质转运等,这暗示它们可能对人类的多种疾病产生复杂的影响。最近,多种lnc RNAs被认为在人类的免疫系统调节中发挥着重要作用的,并且它们可能涉及一些由免疫介导的炎症性疾病的发病机理。因此,本研究推测lnc RNAs可能在SLE的发病机制中发挥了重要的作用。目的检测SLE病例和健康对照外周血单个核细胞(peripheral blood mononuclear cells,PBMCs)中lnc RNAs(GAS5,lnc-DC,linc0597 and linc0949)的表达水平,以及这些lnc RNAs基因多态性与SLE易感性之间的关联。方法本研究采用两阶段的病例对照设计。在第一阶段中,纳入85例SLE病例和71例健康对照,用于检测lnc RNAs在PBMCs中的表达水平;当发现有差异表达的lnc RNAs后,第二阶段纳入860例SLE病例和831例健康对照用于检测其单核苷酸多态性(single nucleotide polymorphisms,SNPs)。Lnc RNAs的表达水平采用实时荧光定量聚合酶链式反应(quantitative real-time polymerase chain reaction,q RT-PCR),基因分型实验通过Fluidigm EP1平台以及Taq Man SNP探针分型方法。对4种lnc RNAs表达水平以及4个SNP位点的统计分析进行Bonferroni多重校正,检验水准α=0.0125(0.05/4),其余统计分析检验水准α=0.05。结果(1)Linc0597,lnc-DC和GAS5在SLE病例PBMCs中的表达水平均低于健康对照中的水平,差异均有统计学意义(Z=-5.984,P0.001;Z=-3.703,P0.001;Z=-2.995,P=0.003);linc0949的表达水平在SLE和健康对照中差异无统计学意义(Z=-0.254,P=0.799)。(2)Linc0597在SLE合并狼疮肾炎(lupus nephritis,LN)组的表达水平要低于非LN组,差异有统计学意义(Z=-2.411,P=0.016)且linc0597在SLE合并蛋白尿阳性病例中的表达水平低于蛋白尿阴性病例中的表达水平,差异有统计学意义(Z=-2.865,P=0.004)。Linc0949,lnc-DC和GAS5的表达水平在SLE合并LN组与非LN组中差异均无统计学意义(均有P0.05)。(3)Linc0597的表达水平与SLE疾病活动度评分(systemic lupus erythematosus disease activity index 2000,SLEDAI-2K)成负相关(rs=-0.267,P=0.013);Linc0949,lnc-DC和GAS5的表达水平未发现与SLEDAI-2K评分存在相关性(均有P0.05)。(4)四个SNP位点与SLE的易感性被纳入分析:lnc-DC的rs10515177位点;linc0597(BZRAP1-AS1)的rs2070107,rs2632516和rs2877877位点。结果显示,linc0597的rs2070107位点基因型CC与GG在SLE病例和健康对照中的分布频率存在不同,差异有统计学意义(χ2=7.164,P=0.007)。调整性别、年龄后差异无统计学意义(χ2=5.000,P=0.025);遗传模型分析显示,SLE病例和健康对照在隐形模型(CC vs CG+GG)中差异有统计学意义(χ2=7.244,P=0.007),调整性别、年龄后差异无统计学意义(χ2=5.220,P=0.022)。其余lnc RNA的SNPs位点(rs10515177,rs2632516,rs2877877)基因多态性均未发现与SLE易感性存在关联(均有P0.0125)。(5)在34个SLE病例中的研究发现,rs2070107位点基因多态性(CC+CG vs GG)与linc0597在PBMCs中的表达水平有关,差异有统计学意义(Z=-2.236,P=0.025)。结论Linc0597,lnc-DC,GAS5在SLE病例中的表达水平较健康对照中的明显下调,但linc0949在SLE病例和健康对照中的表达水平无明显差异。进一步分析发现,SLE合并LN病例中的linc0597表达水平要低于非LN病例中的,且linc0597的表达水平可能与SLE疾病活动度呈负相关。基因分型结果显示,rs2070107位点基因多态性与SLE疾病易感性存在关联,但Bonferroni多重校正显示调整性别、年龄后rs2070107位点基因多态性与SLE疾病易感性之间的关联性消失。
[Abstract]:Background systemic lupus erythematosus (systemic lupus erythematosus, SLE) is a chronic, multisystemic autoimmune disease characterized by the deposition of a variety of autoantibodies and a large number of immune complexes, which can cause damage to various tissues or organs in the body. So far, the exact pathogenesis of SLE has not yet been explained. In the past several decades, experimental and clinical studies have shown that genetic, environmental, immunomodulatory factors and interactions among them have participated in the process of SLE disease. Through the genome-wide association studies (GWAS), many susceptible genes of SLE have been developed. Previous studies on SLE are the main collection. There are relatively few reports of coding genes, not coding genes. Unlike the widely studied short chain non coded RNA (such as micro RNAs), long chain non coded RNA (long noncoding RNA, LNC RNA) is a class of non coded RNA with a length of more than 200 nucleotides, and LNC RNAs may be regulated in a variety of biological functions. Use. Although the exact function of LNC RNAs remains largely unknown, some studies have shown that LNC RNAs may be involved in a variety of critical biological processes, such as chromatin remodeling, gene transcription, RNA splicing, and protein transport, suggesting that they may have a complex impact on a variety of human diseases. Recently, a variety of LNC RNAs It is considered to play an important role in the regulation of human immune system, and they may be involved in the pathogenesis of some immune mediated inflammatory diseases. Therefore, this study suggests that LNC RNAs may play an important role in the pathogenesis of SLE. Objective to detect SLE disease and healthy control of peripheral blood mononuclear cells (periphe The expression level of LNC RNAs (GAS5, lnc-DC, linc0597 and linc0949) in ral blood mononuclear cells, PBMCs, and the association between these polymorphisms and susceptibility. Methods the two stage case control design was used. Expression level in MCs; after the discovery of differentially expressed LNC RNAs, the second stage was included in 860 cases of SLE and 831 healthy controls to detect the expression level of the single nucleotide polymorphisms (single nucleotide polymorphisms, SNPs).Lnc RNAs using real time fluorescent quantitative polymerized enzyme chain reaction (quantitative real-time) Reaction, Q RT-PCR), the genotyping experiment was carried out by the Fluidigm EP1 platform and the Taq Man SNP probe typing method. The statistical analysis of 4 LNC RNAs expressions and the statistical analysis of the 4 SNP loci were performed on the Bonferroni multiplicity correction, the remaining statistical analysis test level alpha results (1). The expression level in BMCs was lower than that in the healthy control, and the difference was statistically significant (Z=-5.984, P0.001; Z=-3.703, P0.001; Z=-2.995, P=0.003); the expression level of linc0949 was not statistically significant in SLE and healthy controls (Z=-0.254, P=0.799). (2) the expression level of Linc0597 in the group of lupus nephritis (lupus nephritis) The difference was statistically significant (Z=-2.411, P=0.016) and the expression level of linc0597 in SLE combined proteinuria positive cases was lower than that of proteinuria negative cases, the difference was statistically significant (Z=-2.865, P=0.004).Linc0949, and the expression level of lnc-DC and GAS5 was not statistically significant in SLE combined with non LN groups. Learning significance (all P0.05). (3) the expression level of Linc0597 was negatively correlated with the SLE disease activity score (systemic lupus erythematosus disease activity index 2000, SLEDAI-2K). The rs10515177 locus of lnc-DC; rs2070107, rs2632516 and rs2877877 loci of linc0597 (BZRAP1-AS1). The results showed that the frequency of rs2070107 loci genotype CC and GG in linc0597 were different in SLE cases and healthy controls. Statistical significance (x 2=5.000, P=0.025); genetic model analysis showed that the difference between SLE and CC vs CG+GG (CC vs CG+GG) was statistically significant (x 2=7.244, P=0.007), and there was no statistical significance in adjusting sex (x 2=5.220, P=0.022) after age (P=0.022). No association was found to be associated with SLE susceptibility (all P0.0125). (5) in the study of 34 SLE cases, the rs2070107 locus gene polymorphism (CC+CG vs GG) was associated with the expression level of linc0597 in PBMCs (Z=-2.236, P=0.025). There was no significant difference in the expression level of linc0949 in SLE cases and healthy controls. Further analysis showed that the expression level of linc0597 in the cases of SLE combined with LN was lower than that in non LN cases, and the expression level of linc0597 may be negatively correlated with the degree of SLE disease activity. The result of the genotyping showed that the polymorphism of the rs2070107 loci was polymorphic. Sex is associated with the susceptibility to SLE disease, but the Bonferroni multiplex correction shows that the association between the polymorphism of the rs2070107 locus after age and the susceptibility to SLE disease is disappearing.

【学位授予单位】：安徽医科大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：R593.241

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