HCV基因型分布及HLA-DP基因多态性与高危人群HCV感染转归的关系

发布时间：2018-05-26 18:25

本文选题：丙型肝炎病毒 + 基因型　；参考：《南京医科大学》2014年硕士论文

【摘要】：丙型肝炎病毒(Hepatitis C Virus, HCV)感染呈全球流行,据世界卫生组织(WHO)估计,全世界约有3%的人口(即1.7亿)感染HCV。目前,中国自然人群感染率约在0.9%-5.1%,平均约为3.2%,且呈增长趋势。HCV感染所致的丙型肝炎慢性化率高达75%～85%,可导致肝脏慢性炎症坏死和纤维化,部分患者可发展为肝硬化甚至肝细胞癌,对人类的健康和生命构成了严重威胁。HCV为单股正链RNA病毒且属于黄病毒家族成员。其基因组全长约9.5kb,分为5’非编码区(5'untranslated region,5'UTR)、编码区以及3’非编码区(3'UTR),其中5'UTR区、编码区中的核心区最保守,E1区、E2区最易变异。HCV高度变异导致其基因型和亚型的在全球呈区域性分布。我国大部分地区流行的HCV型别为1b型,其次为2a型,在南方城市1b型感染率占90%以上,从南向北2a型逐渐增多。HCV不同基因型、亚型与HCV感染者临床症状的严重程度、疾病的转归以及HCV的感染途径、地区分布规律等具有高度的相关性。此外,HCV基因型也是影响丙型肝炎患者干扰素治疗效果的重要因素之一。 HCV感染时,体液免疫在抗病毒感染中作用较小,而CD4+T淋巴细胞反应起很重要的作用。人类白细胞抗原(]human leukocyte antigen, HLA) Ⅱ类抗原是由人类主要组织相容性复合体(najor histocompatibility complex, MHC)中II类基因编码的HLA蛋白分子,主要分布于B淋巴细胞、巨噬细胞和活化的T淋巴细胞表面,其功能是将外源性抗原肽递呈给CD4+T淋巴细胞。因此,HLA-Ⅱ类基因的高度多态性是决定宿主免疫应答能力的最重要的遗传因素。HLA-Ⅱ类抗原基因主要包括HLA-DR、DQ和DP三个亚区,每个亚区的基因结构都呈高度多态性。HLA-DP分子是MHCⅡ类抗原,为人类的主要组织相容性复合体中一种蛋白质/肽抗原受体,HLA-DP在抗原提呈和免疫应答中发挥着重要作用,HLA-DP分子在宿主清除病毒过程中发挥重要的作用,提示HLA-DP基因型可能在HCV感染的慢性化过程中扮演重要角色。本研究拟以HLA-DP基因多态性为切入点,进一步采用病例-对照研究在中国HCV高危人群中,探讨不同感染途径丙型肝炎患者HCV基因型的分布和HLA-DP基因多态性与HCV感染及其转归的关系。第一部分江苏地区不同感染途径丙型肝炎患者HCV基因分型的研究丙型肝炎病毒(Hepatitis C Virus, HCV)为单股正链RNA,其基因组分为5’非编码区(5'untranslated region,5'UTR)、编码区以及3’非编码区(3UTR),其中5'UTR区、编码区中的核心区最保守,E1区、E2区最易变异。HCV高度变异导致其基因型和亚型在全球呈区域性分布。HCV基因型对HCV感染的流行病学研究具有重要意义。此外,HCV基因型也是影响丙型肝炎患者干扰素治疗效果的重要因素之一。为此,本研究于2006年5月——2009年12月在江苏省收集505例不同感染途径的丙型肝炎患者,并对其进行HCV常见基因型分型检测,探讨江苏省HCV基因型在不同感染途径人群中的分布情况,为丙型肝炎的临床治疗提供科学依据。 [目的]探讨江苏地区不同感染途径丙型肝炎患者病毒(HCV)基因型的分布及其与感染途径、肝功能的关系,为丙型肝炎的治疗策略和措施提供理论依据。 [方法]按照Simmonds分型方法对505例不同感染途径的丙型肝炎患者进行HCV基因型分型检测并分析年龄、性别、肝功能以及感染途径与HCV基因型的关系。 [结果] 1.HCV基因分型检测结果505例HCVRNA阳性标本中,1a型占1.6%,1b型占68.9%,2型占4.8%,3型占7.9%,其它分型占16.8%。 2.不同特征人群HCV基因型分布男性与女性的HCV基因型分布差异无统计学意义(χ2=4.94,P=0.08)；不同年龄人群中的HCV基因型分布差异亦无统计学意义(χ2=1.61,P=0.81)；不同HCV基因型人群的ALT.AST水平均无统计学差异(PALT＝0.64,PAsT=0.28)。 3.不同感染途径患者HCV基因型分布情况不同感染途径人群的HCV基因型分布存在统计学差异(χ2=73.35,P0.01)。 [结论]江苏地区丙型肝炎患者HCV基因型以1b型为主,1a型HCV在中国大陆地区流行程度有扩大趋势,HCV基因型与其感染途径有关。第二部分 HLA-DP基因多态性与HCV感染转归的关系丙型肝炎病毒(hepatitis C virus, HCV)是单链RNA病毒,人体感染HCV后,遗传因素参与了机体免疫,可能影响疾病的易感性及病程的进展。HLA是迄今所知最复杂、多态性最高的免疫相关遗传系统,在人类的免疫应答反应中起着十分重要的作用。HLA-Ⅱ类基因分为经典HLA-DP-DQ、-DR和非经典的HLA-DM.-DO两类。HLA-DP由组成2个亚基,DPα和DPp的抗原。α链和β链的远膜区有一个抗原结合槽(antigen binding cleft),因此可以推钡HLA-DP基因的多态性可导致了抗原结合槽的构象、结合及递呈抗原肽给T细胞的效率不同。从而影响着免疫反应的强弱。目前国内外大量研究表明HLA-DP基因区域的某些多态性位点与个体更易导致感染HCV慢性化。然而,未发现有关于HLA-DP基因多态性与HCV感染转归的研究出现。本研究探讨HLA-DP基因多态性与HCV感染转归的相关性,为研究HCV感染的发生机制和临床治疗提供新的线索。 [目的]探讨HLA-DP基因多态性与HCV感染转归的关系。 [方法]采用病例-对照研究设计,收集1653例HCV高危人群,其中包括401例HCV持续感染者(HCVRNA阳性),268例HCV自限清除者(抗-HCV抗体阳性,HCVRNA阴性)以及984例健康对照者(抗-HCV抗体阴性),采用TaqMan-MGB探针技术检测HLA-DP单核苷酸多态性,比较不同基因型与HCV感染慢性化关系。 [结果] 1.一般情况比较本研究有研究对象共1653人,分别为对照组984人、自限清除组268人和持续感染组401人。三组的性别和年龄构成差异均无统计学意义(P性别=0.24,P年龄=0.07),但是三组之间的谷丙转氨酶(ALT)和谷草转氨酶(AST)水平以及高危人群种类分布差异均有统计学意义。 2. HLA-DP基因多态性与HCV感染转归的关系将HCV感染组(自限清除组+持续感染组)与对照组进行比较,采用多因素Logistic回归分析,对性别、年龄和高危人群种类因素进行调整,估计HLA-DP突变基因型对HCV易感性的影响。rs3077位点分析结果显示：杂合基因型CT和突变型TT在感染组中的频率均高于对照组(调整OR和95%C1分别为1.31,1.05-1.64和1.93,1.36-2.73),提示杂合基因型CT和突变型TT可增加个体对HCV的易感性。rs2395309位点分析结果显示：杂合基因型GA和突变型AA在感染组中的频率均高于对照组(调整OR和95%C1分别为1.29,1.03-1.61和1.89,1.34-2.68),提示杂合基因型GA和突变型AA可增加个体对HCV的易感性。3. HLA-DP基因多态性与HCV自限清除的关系将HCV自限清除组与持续感染组进行比较,采用多因素Logistic回归分析,对性别、年龄和高危人群种类因素进行调整,估计HLA-DP基因多态性与HCV感染自限清除的关系。rs9277535位点分析结果显示：杂合基因型GA和突变型AA在持续性感染组中的频率均高于自限清除组(调整OR和95%C1分别为1.54,1.05-2.28和1.62,1.01-2.60),提示杂合基因型GA和突变型AA可降低个体自限清除病毒的能力。 4.单倍型分析以HLA-DP基因3个SNP位点(rs3077, rs9377535和rs2395309)中均成功进行基因型检测的样本为研究对象,用PHASE2.0推断可能的单倍型及其频率。与最常见的单倍型CGG相比,含三个位点突变等位基因的单倍型TAA可以增加HCV的易感性(调整OR和95%CI为1.31,1.09-1.55)。 [结论]HLA-DP基因多态性可能与丙型肝炎易感性及慢性化有关。
[Abstract]:Hepatitis C Virus (HCV) infection is a global epidemic. According to WHO (WHO), about 3% of the population (or 170 million) of the world is infected with HCV.. The infection rate of natural population in China is about 0.9%-5.1%, with an average of about 3.2%, and the chronicity of hepatitis C caused by.HCV infection is up to 75% ~ 85%, which can lead to Chronic inflammatory necrosis and fibrosis of the liver, some patients can develop into liver cirrhosis and even hepatocellular carcinoma, which poses a serious threat to human health and life..HCV is a single strand of RNA virus and belongs to the family of the yellow virus. The total length of the genome is about 9.5kb, divided into 5 'non coding region (5'untranslated region, 5'UTR), coding region and 3' non. The coding area (3'UTR), in which the core area of the 5'UTR region and the coding region is the most conservative, the E1 region and the most variable.HCV height variation in the E2 region lead to the regional distribution of the genotype and subtype in the world. The prevalent HCV types in most areas of China are 1B type, followed by 2 A, the 1b type infection rate in southern city is over 90%, and the 2A type is gradually increasing from south to north to.H. CV different genotypes, subtypes and the severity of the clinical symptoms of HCV infected people, the prognosis of the disease and the route of HCV infection, regional distribution, and so on. In addition, the HCV genotype is also one of the important factors affecting the effect of interferon therapy in hepatitis C patients.
In HCV infection, humoral immunity plays a small role in antiviral infection, and CD4+T lymphocyte reaction plays an important role. Human leukocyte antigen (]human leukocyte antigen, HLA) class II antigen is a HLA protein molecule encoded by the II class of human major histocompatibility complex (najor histocompatibility complex, MHC). Distributed in B lymphocyte, macrophage and activated T lymphocyte, its function is to send exogenous antigen peptide to CD4+T lymphocyte. Therefore, the high polymorphism of HLA- class II gene is the most important genetic factor determining host immune response,.HLA- class II anti primordia mainly includes three subregions of HLA-DR, DQ and DP, each subregion. The genetic structure of the region is highly polymorphic and.HLA-DP is a MHC class II antigen. It is a protein / peptide antigen receptor in the major histocompatibility complex of human beings. HLA-DP plays an important role in antigen presentation and immune response. HLA-DP molecules play an important role in the process of virus removal in the host, suggesting that the HLA-DP genotypes are available. It plays an important role in the chronicity of HCV infection.
In this study, we use the HLA-DP gene polymorphism as the breakthrough point to further use case control study to investigate the distribution of HCV genotypes and the relationship between the polymorphism of HLA-DP gene and the HCV infection and the outcome of the hepatitis C patients with different infection routes in the high risk population of HCV in China.
Part one
HCV genotyping of hepatitis C patients with different routes of infection in Jiangsu
Hepatitis C Virus (HCV) is a single strand positive chain RNA whose genome is divided into 5 'non coding region (5'untranslated region, 5'UTR), coding region and 3' non coding region (3UTR). The 5'UTR region, the core region of the coding region is the most conservative, E1 region, and the largest variation in the E2 region causes its genotypes and subtypes to be regional in the global region. The distribution of.HCV genotype is of great significance to the epidemiological study of HCV infection. In addition, the HCV genotype is one of the important factors affecting the effect of interferon therapy in hepatitis C patients. In this study, 505 cases of hepatitis C in different routes of infection were collected in Jiangsu province in May 2006 and December 2009, and the common HCV was common. Genotypic typing was used to investigate the distribution of HCV genotypes in Jiangsu among different routes of infection, so as to provide a scientific basis for clinical treatment of hepatitis C.
[Objective] to investigate the distribution of virus (HCV) genotype of hepatitis C virus (HCV) in different infection routes in Jiangsu and its relationship with the way of infection and liver function, and provide a theoretical basis for the treatment strategy and measures of hepatitis C.
[Methods] HCV genotyping was carried out in 505 hepatitis C patients with different infection routes according to the Simmonds typing method and the relationship between age, sex, liver function and the infection pathway and HCV genotype was analyzed.
[results]
The results of 1.HCV genotyping test showed that among 505 HCVRNA positive specimens, 1.6% were 1a, 68.9% were 1b, 2 were 4.8%, 3 were 7.9%, and the others were 16.8%..
2. there was no significant difference in the distribution of HCV genotypes between men and women in different HCV genotypes (x 2=4.94, P=0.08), and there was no statistical difference in the distribution of HCV genotypes in different age groups (x 2=1.61, P=0.81), and there was no statistical difference in the level of ALT.AST among the different HCV genotypes (PALT = 0.64, PAsT=0.28).
3. the distribution of HCV genotypes in patients with different routes of infection was statistically different from those in the infected group (HCV 2=73.35, P0.01).
[Conclusion] the HCV genotype of hepatitis C patients in Jiangsu area is mainly 1b type, and the prevalence of type 1 a HCV in the mainland of China has a tendency to expand, and the HCV genotype is related to the infection pathway.
The second part
The relationship between the polymorphism of HLA-DP gene and the outcome of HCV infection
Hepatitis C virus (hepatitis C virus, HCV) is a single chain RNA virus. After human infection of HCV, genetic factors participate in the immune system, which may affect the susceptibility and progress of the disease..HLA is the most complex and polymorphic immune related genetic system known to date, which plays a very important role in the response of human immune response.HLA-. Class II genes are divided into classical HLA-DP-DQ, -DR and non classical HLA-DM.-DO two.HLA-DP, which consist of 2 subunits, DP alpha and DPp antigens. The alpha and beta chains have an antigen binding slot (antigen binding cleft). Therefore, the polymorphism of the barium HLA-DP gene can lead to the conformation of antigenic grooves, binding and presenting antigen peptides to T fines. At present, a large number of studies at home and abroad have shown that some polymorphic loci and individuals in the HLA-DP gene region are more likely to lead to the chronicity of HCV. However, there is no discovery of the HLA-DP gene polymorphism and the HCV infection.
The aim of this study was to investigate the association between HLA-DP gene polymorphism and the outcome of HCV infection, and to provide new clues for studying the pathogenesis and clinical treatment of HCV infection.
[Objective] to explore the relationship between HLA-DP gene polymorphism and the prognosis of HCV infection.
[Methods] a case control study was used to collect 1653 high risk groups of HCV, including 401 cases of HCV persistent infection (HCVRNA positive), 268 HCV self limiting scavengers (anti -HCV antibody positive, HCVRNA negative) and 984 healthy controls (anti -HCV negative). The HLA-DP single nucleotide polymorphisms were detected by TaqMan-MGB probe technique. The chronicity of the genotype and HCV infection.
[results]
1. general situation comparison
There were 1653 subjects in this study, 984 in the control group, 268 in the self clearance group and 401 in the continuous infection group. There was no significant difference in gender and age between the three groups (P sex =0.24, P age =0.07), but the level of alanine transaminase (ALT) and gluten transaminase (AST) between the three groups and the differences in the distribution of high risk groups There were statistical significance.
Relationship between 2. HLA-DP gene polymorphism and HCV infection outcome
The HCV infection group (self limited clearance group + persistent infection group) was compared with the control group. The multiple factor Logistic regression analysis was used to adjust the factors of sex, age and high risk population, and the effect of HLA-DP mutation genotype on the susceptibility of HCV was estimated. The results of.Rs3077 site analysis showed that the heterozygous genotype CT and the mutant TT were in the infected group. The frequency was higher than that of the control group (adjusted OR and 95%C1 respectively 1.31,1.05-1.64 and 1.93,1.36-2.73), suggesting that heterozygous genotype CT and mutant TT could increase the susceptibility of individual to HCV, which showed that the frequency of heterozygous genotype GA and mutant AA in infected group was higher than that of the control group (OR and 95%C1) were higher than those of the control group. 1.61 and 1.89,1.34-2.68), suggesting that heterozygous genotype GA and mutant AA can increase the susceptibility of individuals to HCV, and the relationship between.3. HLA-DP polymorphism and HCV self limited clearance.
The HCV self limited clearance group was compared with the continuous infection group. The multiple factor Logistic regression analysis was used to adjust the sex, age and high risk population, and the relationship between the HLA-DP gene polymorphism and the HCV infection self limiting clearance was estimated. The results of the.Rs9277535 site analysis showed that the heterozygous genotype GA and the mutant AA were in the persistent infection group. The frequency was higher than that of the self limited scavenging group (adjusting OR and 95%C1 for 1.54,1.05-2.28 and 1.62,1.01-2.60 respectively), suggesting that heterozygous genotype GA and mutant AA could reduce the ability of individual self limiting virus clearance.
4. haplotype analysis
The samples of the 3 SNP loci (rs3077, rs9377535 and rs2395309) of the HLA-DP gene (rs3077, rs9377535 and rs2395309) were studied. The possible haplotype and its frequency were deduced by PHASE2.0. The haplotype TAA containing three loci mutation alleles could increase the susceptibility to HCV (adjusted OR and 95%CI 1.31, compared with the most common haplotype CGG. " 1.09-1.55).
[conclusion]HLA-DP gene polymorphism may be associated with susceptibility to hepatitis C and chronicity.
【学位授予单位】：南京医科大学
【学位级别】：硕士
【学位授予年份】：2014
【分类号】：R512.63

【参考文献】