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  本文選題：紅斑狼瘡 + 系統(tǒng)性RNA基因表達(dá)多態(tài)性�。� 參考：《安徽醫(yī)科大學(xué)》2017年碩士論文

【摘要】：研究背景系統(tǒng)性紅斑狼瘡(systemic lupus erythematosus,SLE)是一種慢性多系統(tǒng)性的自身免疫性疾病,其以多種自身抗體產(chǎn)生和大量免疫復(fù)合物的沉積為主要特點,并可因此導(dǎo)致體內(nèi)多種組織或器官的損傷。目前為止,SLE的確切發(fā)病機制尚未闡明。過去幾十年中的實驗和臨床研究表明,遺傳因素、環(huán)境因素、免疫調(diào)節(jié)因素及其之間的交互作用等共同參與了SLE的疾病進(jìn)程。通過人類全基因組關(guān)聯(lián)研究(genome-wide association studies,GWAS),發(fā)現(xiàn)了多個SLE的易感基因。既往SLE的研究主要集中在編碼基因,而非編碼基因的報道相對較少。和那些已經(jīng)被廣泛研究的短鏈非編碼RNA(例如micro RNAs)不同的是,長鏈非編碼RNA(long noncoding RNA,lnc RNA)是一類長度超過200個核苷酸的非編碼RNA,且lnc RNAs可能在多種生物學(xué)功能中具有調(diào)節(jié)作用。雖然lnc RNAs的確切功能在很大程度上仍然不清楚,但一些研究已經(jīng)表明lnc RNAs可能參與了多種關(guān)鍵的生物學(xué)過程,如染色質(zhì)重塑、基因轉(zhuǎn)錄、RNA剪接和蛋白質(zhì)轉(zhuǎn)運等,這暗示它們可能對人類的多種疾病產(chǎn)生復(fù)雜的影響。最近,多種lnc RNAs被認(rèn)為在人類的免疫系統(tǒng)調(diào)節(jié)中發(fā)揮著重要作用的,并且它們可能涉及一些由免疫介導(dǎo)的炎癥性疾病的發(fā)病機理。因此,本研究推測lnc RNAs可能在SLE的發(fā)病機制中發(fā)揮了重要的作用。目的檢測SLE病例和健康對照外周血單個核細(xì)胞(peripheral blood mononuclear cells,PBMCs)中l(wèi)nc RNAs(GAS5,lnc-DC,linc0597 and linc0949)的表達(dá)水平,以及這些lnc RNAs基因多態(tài)性與SLE易感性之間的關(guān)聯(lián)。方法本研究采用兩階段的病例對照設(shè)計。在第一階段中,納入85例SLE病例和71例健康對照,用于檢測lnc RNAs在PBMCs中的表達(dá)水平;當(dāng)發(fā)現(xiàn)有差異表達(dá)的lnc RNAs后,第二階段納入860例SLE病例和831例健康對照用于檢測其單核苷酸多態(tài)性(single nucleotide polymorphisms,SNPs)。Lnc RNAs的表達(dá)水平采用實時熒光定量聚合酶鏈?zhǔn)椒磻?yīng)(quantitative real-time polymerase chain reaction,q RT-PCR),基因分型實驗通過Fluidigm EP1平臺以及Taq Man SNP探針分型方法。對4種lnc RNAs表達(dá)水平以及4個SNP位點的統(tǒng)計分析進(jìn)行Bonferroni多重校正,檢驗水準(zhǔn)α=0.0125(0.05/4),其余統(tǒng)計分析檢驗水準(zhǔn)α=0.05。結(jié)果(1)Linc0597,lnc-DC和GAS5在SLE病例PBMCs中的表達(dá)水平均低于健康對照中的水平,差異均有統(tǒng)計學(xué)意義(Z=-5.984,P0.001;Z=-3.703,P0.001;Z=-2.995,P=0.003);linc0949的表達(dá)水平在SLE和健康對照中差異無統(tǒng)計學(xué)意義(Z=-0.254,P=0.799)。(2)Linc0597在SLE合并狼瘡腎炎(lupus nephritis,LN)組的表達(dá)水平要低于非LN組,差異有統(tǒng)計學(xué)意義(Z=-2.411,P=0.016)且linc0597在SLE合并蛋白尿陽性病例中的表達(dá)水平低于蛋白尿陰性病例中的表達(dá)水平,差異有統(tǒng)計學(xué)意義(Z=-2.865,P=0.004)。Linc0949,lnc-DC和GAS5的表達(dá)水平在SLE合并LN組與非LN組中差異均無統(tǒng)計學(xué)意義(均有P0.05)。(3)Linc0597的表達(dá)水平與SLE疾病活動度評分(systemic lupus erythematosus disease activity index 2000,SLEDAI-2K)成負(fù)相關(guān)(rs=-0.267,P=0.013);Linc0949,lnc-DC和GAS5的表達(dá)水平未發(fā)現(xiàn)與SLEDAI-2K評分存在相關(guān)性(均有P0.05)。(4)四個SNP位點與SLE的易感性被納入分析:lnc-DC的rs10515177位點;linc0597(BZRAP1-AS1)的rs2070107,rs2632516和rs2877877位點。結(jié)果顯示,linc0597的rs2070107位點基因型CC與GG在SLE病例和健康對照中的分布頻率存在不同,差異有統(tǒng)計學(xué)意義(χ2=7.164,P=0.007)。調(diào)整性別、年齡后差異無統(tǒng)計學(xué)意義(χ2=5.000,P=0.025);遺傳模型分析顯示,SLE病例和健康對照在隱形模型(CC vs CG+GG)中差異有統(tǒng)計學(xué)意義(χ2=7.244,P=0.007),調(diào)整性別、年齡后差異無統(tǒng)計學(xué)意義(χ2=5.220,P=0.022)。其余l(xiāng)nc RNA的SNPs位點(rs10515177,rs2632516,rs2877877)基因多態(tài)性均未發(fā)現(xiàn)與SLE易感性存在關(guān)聯(lián)(均有P0.0125)。(5)在34個SLE病例中的研究發(fā)現(xiàn),rs2070107位點基因多態(tài)性(CC+CG vs GG)與linc0597在PBMCs中的表達(dá)水平有關(guān),差異有統(tǒng)計學(xué)意義(Z=-2.236,P=0.025)。結(jié)論Linc0597,lnc-DC,GAS5在SLE病例中的表達(dá)水平較健康對照中的明顯下調(diào),但linc0949在SLE病例和健康對照中的表達(dá)水平無明顯差異。進(jìn)一步分析發(fā)現(xiàn),SLE合并LN病例中的linc0597表達(dá)水平要低于非LN病例中的,且linc0597的表達(dá)水平可能與SLE疾病活動度呈負(fù)相關(guān)。基因分型結(jié)果顯示,rs2070107位點基因多態(tài)性與SLE疾病易感性存在關(guān)聯(lián),但Bonferroni多重校正顯示調(diào)整性別、年齡后rs2070107位點基因多態(tài)性與SLE疾病易感性之間的關(guān)聯(lián)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.

【學(xué)位授予單位】：安徽醫(yī)科大學(xué)
【學(xué)位級別】：碩士
【學(xué)位授予年份】：2017
【分類號】：R593.241

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