中国汉族人特应性皮炎全基因组关联研究
发布时间：2018-08-13 08:19
【摘要】：研究背景: 特应性皮炎(atopic dermatitis, AD) ,是一种反复发作的慢性炎症性皮肤病。其特征是皮肤瘙痒,皮损多形性并有渗出倾向,常伴有哮喘、过敏性鼻炎及血清IgE增高等,在儿童和青少年中较多见。近三十年来,AD发病率呈逐年上升趋势,较之前增长达2-3倍之多,在儿童中已达到15%-30%,在成人中达到2%-10%。本病病因和发病机制尚不清楚。国内外通过大量的人群、家系及双生子研究均表明遗传因素是构成AD易感性的主要因素,属多基因遗传病。 多年来,国内外学者主要应用全基因组连锁研究(Genome-wide linkage studies)、候选基因研究(Candidate genes studies)等研究策略搜寻AD的易感基因,并取得了一些进展。但这些研究均存在着一定的局限性,如全基因组连锁研究应用的微卫星标记信息量有限,所定位区域往往较大,很难从其中找出真正的易感基因。同时,全基因组连锁研究主要是针对家系研究,因而所发现的疾病易感位点是否能够代表散发病例尚不清楚。对于候选基因研究,其选取候选基因主要根据研究者们认为最有可能导致疾病发生的易感基因,其分析的位点过少,故不可避免的存在着局限性,不可能全面搜寻到AD的易感基因。 2005年以后,随着人类基因组计划(HGP)和人类基因组单体型图计划(HapMap)的相继完成,以及高通量基因分型技术的飞速发展和分型费用的降低,使得在大规模人群中开展全基因组关联研究(genome-wide association study, GWAS)成为可能。这种研究方法对全基因组范围内的SNP(single nucleotide polymorphism)进行总体的分析,能够更有效的发现和疾病有关联的基因。GWAS是目前寻找复杂疾病易感基因的最有效方法之一。多项研究的成功有力证明了GWAS搜寻复杂疾病易感基因的有效性,也为AD易感基因的搜寻提供了新的思路和方法。2009年,Esparza-Gordillo等对德国人群进行了大规模的GWAS研究,发现11q13.5(C11orf30)为AD的易感位点。 AD在不同人种间存在遗传异质性,主要表现在发病率和患病率以及临床表现存在一定的差异。由于多数AD研究主要是在欧洲高加索白人中进行的,因而这些AD遗传易感基因/位点与中国人群的关联性尚不清楚。因此对AD在中国人群中的易感基因研究很有必要。本研究首次在中国汉族人群中应用GWAS方法在全基因组范围内对AD的易感基因进行筛查,以期发现AD的易感基因。 目的: 在中国汉族人群中,利用全基因组关联分析研究方法在全基因组范围内搜寻与AD关联的遗传变异,鉴定中国汉族人AD的易感基因/位点。 方法: (1)初筛阶段:在中国汉族人群中,利用Illumina Human 610-Quad全基因组SNP分型芯片对1012个AD病例和1362个对照进行SNPs分型。 (2)验证阶段:①验证第一阶段:经过严格的数据质控和统计分析,从初筛阶段选择最有意义的67个SNPs,使用Sequenom平台在二个独立的中国人群中进行验证(北部人群:1119个病例和1203个对照;南部人群:2505个病例和6762个对照); ②验证第二阶段:从验证第一阶段中选取了19个SNPs,进一步加大样本量使用Sequenom平台在二个独立的中国人群中进行验证(北部人群:276个正常对照;南部人群:3956个正常对照)。之后,将上述附加的对照和之前已分型的对照数据及病例进行合并分析(验证第一、二阶段北部人群总计:1119个病例和1479个对照;验证第一、二阶段南部人群总计:2505个病例和10718个对照;GWAS初筛阶段人群总计:1012个病例和1362个对照),以发现了达到全基因组显著关联性水平SNPs(P5×10~(-8)); ③验证第三阶段:将上一阶段发现的有意义的SNPs进一步在德国的人群中使用TaqMan进行验证(1,806个病例和3,256个正常对照)。 结果: (1)在验证第一阶段中,在中国汉族人群的3624例患者和7965例对照中对67个SNPs进行基因分型。将验证第一阶段的结果和GWAS结果合并分析后显示,位于2个易感位点的3个SNPs达到了全基因组关联显著水平(P5×10~(-8)):①位于1q21.3(FLG)易感位点上的2个SNPs的rs3126085/rs11204971 (关联性分别为P_(combined)=5.75×10~(-12), OR=0.83; P_(combined)=5.02×10~(-9), OR=0.86);②位于5q22.1(TMEM232 /SLC25A46)易感位点上的1个SNP rs7701890 (P_(combined)=4.33×10~(-8), OR=1.23); (2)在验证第二阶段中,对从验证第一阶段中选取的19个SNPs在附加的4232对照中进行分型,之后与第一阶段的样本进行合并分析,最后再与GWAS数据综合分析后发现了位于3个易感位点区域(1q21.3, 5q22.1和20q13.33)内的7个SNPs达到全基因组显著水平:①位于1q21.3(FLG)易感位点上的2个SNPs rs3126085/rs11204971 (关联性分别为P_(combined)=5.90×10~(-12), OR=0.82; P_(combined)=1.20×10~(-10), OR=0.85);②位于5q22.1(TMEM232/SLC25A46)易感位点上的4个SNPs rs7701890/rs10067777/rs13360927/rs13361382 (关联性分别为P_(combined)=3.15×10~(-9), OR=1.24; P_(combined)=1.20×10~(-8), OR=1.24; P_(combined)=2.45×10~(-8), OR=1.22; P_(combined)=4.02×10~(-8), OR=1.22);③位于20q13.33(TNFRSF6B/ZGPAT)易感位点上的1个SNP rs6010620 (P_(combined)=3.0×10~(-8), OR=1.17)。此外,尚有1个提示性的易感位点,即位于10q21.2(ZNF365)易感位点上的SNP rs2393903 (P_(combined)=1.05×10~(-7), OR=1.15); (3)在验证第三阶段中,发现20q13.33(TNFRSF6B/ZGPAT)也与德国AD存在关联性(rs6010620, P=2.87×10~(-5), OR=1.25)。 结论: 本研究通过大样本量的全基因组关联研究搜寻中国汉族人群AD易感基因,构建了第一个中国汉族人群AD病例-对照的全基因组关联分析数据库。在中国人群中发现了2个新的AD易感基因/位点:5q22.1 (TMEM232/SLC25A46),20q13.33 (TNFRSF6 / ZGPAT);验证出既往欧洲人群和亚洲人群报道的1q21.3(FLG)易感位点;发现了AD一个提示易感位点10q21.2(ZNF365)。此外,发现20q13.33 (TNFRSF6B / ZGPAT)与德国人群AD发病也存在关联性。
[Abstract]:Research background:
Atopic dermatitis (AD) is a recurrent chronic inflammatory skin disease characterized by itching, pleomorphic skin lesions and exudative tendencies, often accompanied by asthma, allergic rhinitis and increased serum IgE, more common in children and adolescents. In the past 30 years, the incidence of AD has increased year by year, compared with the previous increase. As many as 2-3 times, it has reached 15-30% in children and 2-10% in adults. The etiology and pathogenesis of AD are still unclear.
Over the years, scholars at home and abroad have made some progress in searching for the susceptible genes of AD by using the strategies of Genome-wide linkage studies and Candidate genes studies. However, these studies have some limitations, such as the application of microsatellite markers in whole-genome linkage studies. It is not clear whether the susceptibility loci can represent sporadic cases. For candidate gene research, the candidate genes are selected mainly according to the researchers'opinion. The susceptibility genes most likely to lead to disease are too few to analyze, so there are inevitable limitations and it is impossible to search for the susceptibility genes of AD comprehensively.
Since 2005, with the completion of the Human Genome Project (HGP) and the HapMap, as well as the rapid development of high-throughput genotyping techniques and the reduction of typing costs, it has become possible to conduct genome-wide association studies (GWAS) in large populations. GWAS is one of the most effective methods to find susceptible genes for complex diseases. Successful studies have proved the effectiveness of GWAS in searching susceptible genes for complex diseases. In 2009, Esparza-Gordillo et al. conducted a large-scale GWAS study on the German population and found that 11q13.5 (C11orf30) was the susceptible site of AD.
The genetic heterogeneity of AD among different races is mainly manifested by differences in morbidity, prevalence and clinical manifestations. Since most AD studies have been conducted mainly among Caucasian Europeans, the association of these AD susceptibility genes/loci with Chinese populations is not yet clear. It is necessary to study the susceptibility genes of AD. GWAS method was used to screen the susceptibility genes of AD in Chinese Han population for the first time.
Objective:
In order to identify the susceptible gene/locus of AD in Chinese Han population, genome-wide association analysis was used to search for the genetic variation associated with AD in the whole genome.
Method:
(1) Preliminary screening stage: 1 012 AD cases and 1 362 controls were classified by using the Illumina Human 610-Quad genome-wide SNP typing chip.
(2) Verification phase: 1. Verification phase: After strict data quality control and statistical analysis, 67 most significant SNPs were selected from the initial screening phase and validated in two independent Chinese populations using the Sequenom platform (northern population: 1119 cases and 1203 controls; southern population: 2505 cases and 6762 controls);
(2) The second phase of validation: 19 SNPs were selected from the first phase of validation, and further enlarged the sample size using the Sequenom platform to validate in two independent Chinese populations (northern population: 276 normal controls; southern population: 3956 normal controls). Consolidated analysis (validation of 1,119 cases and 1,479 controls in the northern population of the first and second stages; validation of 2,055 cases and 10,718 controls in the southern population of the first and second stages; total of 1,012 cases and 1,362 controls in the GWAS preliminary screening stage) was performed to identify SNPs (P5 *10~(-8)) reaching the genome-wide significant association level.
(3) Verification Phase 3: The meaningful SNPs found in the previous phase were further validated with TaqMan in the German population (1,806 cases and 3,256 normal controls).
Result:
(1) In the first stage of validation, 67 SNPs were genotyped in 3 624 Chinese Han patients and 7 965 controls. The results of the first stage of validation and GWAS analysis showed that three SNPs located at two susceptibility loci reached a significant level of genome-wide association (P5 *10-8):1 at the 1q21.3 (FLG) susceptibility locus The rs3126085 / rs11204971 of the two SNPs were P_ (combined) = 5.75 * 10 ~ (- 12) and OR = 0.83; P_ (combined) = 5.02 * 10 ~ (- 9), OR = 0.86; and 1 SNP rs7701890 (P_ (combined) = 4.33 * 10 ~ (- 8), OR = 1.23, located at 5q22.1 (TMEM232 / SLC25A46) susceptibility site.
(2) In the second stage of validation, 19 SNPs selected from the first stage of validation were typed in an additional 4232 control, and then combined with the first stage of the sample analysis. Finally, comprehensive analysis with GWAS data revealed that 7 SNPs located in three susceptibility locus regions (1q21.3, 5q22.1 and 20q13.33) reached genome-wide significance. Levels: 1) Two SNPs at 1q21.3 (FLG) susceptibility locus rs3126085/rs11204971 (P_ (combined) = 5.90 *10 ~(-12), OR = 0.82; P_ (combined) = 1.20 *10 ~(-10), OR = 0.85); 2) Four SNPs at 5q22.1 (TMEM232/SLC25A46) susceptibility locus rs7701890/rs10067777/rs13360927/rs13362 (comb P_ 13362, respectively) (combined) =2.45 x 10 ~ (-8), OR = 1.22; P (combined) = = 2.45 x 10 ~ (-8), OR = 1.22; P (combined) = = 4.02 x 10 ~ (-8), OR = 1.22; P (combined) = = 4.02 x 10 ~ (-8), OR = 1.22); 3) 20 q13.33 (TNFRSF6B / ZGPAT) at the susceptibility site of 20q13.33 (TNFRSF6B / ZGPAT) susceptibility site of 1 SNP 6010 620 (combined) =3.0 (combined) =3.0 (combined) =3.0 ~ (-10 ~ (-8 Suggestive susceptibility locus, i.e. at 1 0q21.2 (ZNF365) SNP rs2393903 (P_ (combined) =1.05 * 10~ (-7), OR=1.15) on susceptible loci.
(3) In the third stage of verification, 20q13.33 (TNFRSF6B/ZGPAT) was also found to be associated with German AD (rs6010620, P = 2.87 x 10-5, OR = 1.25).
Conclusion:
In this study, we searched for AD susceptibility genes in Chinese Han population and constructed the first Chinese Han population case-control genome-wide association analysis database. Two new AD susceptibility genes/loci were found in Chinese population: 5q22.1 (TMEM232/SLC25A46), 20q13.33 (TNFRSF6/ZGPAT). In addition, 20q13.33 (TNFRSF6B/ZGPAT) was also found to be associated with AD in the German population.
【学位授予单位】：安徽医科大学
【学位级别】：博士
【学位授予年份】：2011
【分类号】：R758.2
本文编号：2180409
[Abstract]:Research background:
Atopic dermatitis (AD) is a recurrent chronic inflammatory skin disease characterized by itching, pleomorphic skin lesions and exudative tendencies, often accompanied by asthma, allergic rhinitis and increased serum IgE, more common in children and adolescents. In the past 30 years, the incidence of AD has increased year by year, compared with the previous increase. As many as 2-3 times, it has reached 15-30% in children and 2-10% in adults. The etiology and pathogenesis of AD are still unclear.
Over the years, scholars at home and abroad have made some progress in searching for the susceptible genes of AD by using the strategies of Genome-wide linkage studies and Candidate genes studies. However, these studies have some limitations, such as the application of microsatellite markers in whole-genome linkage studies. It is not clear whether the susceptibility loci can represent sporadic cases. For candidate gene research, the candidate genes are selected mainly according to the researchers'opinion. The susceptibility genes most likely to lead to disease are too few to analyze, so there are inevitable limitations and it is impossible to search for the susceptibility genes of AD comprehensively.
Since 2005, with the completion of the Human Genome Project (HGP) and the HapMap, as well as the rapid development of high-throughput genotyping techniques and the reduction of typing costs, it has become possible to conduct genome-wide association studies (GWAS) in large populations. GWAS is one of the most effective methods to find susceptible genes for complex diseases. Successful studies have proved the effectiveness of GWAS in searching susceptible genes for complex diseases. In 2009, Esparza-Gordillo et al. conducted a large-scale GWAS study on the German population and found that 11q13.5 (C11orf30) was the susceptible site of AD.
The genetic heterogeneity of AD among different races is mainly manifested by differences in morbidity, prevalence and clinical manifestations. Since most AD studies have been conducted mainly among Caucasian Europeans, the association of these AD susceptibility genes/loci with Chinese populations is not yet clear. It is necessary to study the susceptibility genes of AD. GWAS method was used to screen the susceptibility genes of AD in Chinese Han population for the first time.
Objective:
In order to identify the susceptible gene/locus of AD in Chinese Han population, genome-wide association analysis was used to search for the genetic variation associated with AD in the whole genome.
Method:
(1) Preliminary screening stage: 1 012 AD cases and 1 362 controls were classified by using the Illumina Human 610-Quad genome-wide SNP typing chip.
(2) Verification phase: 1. Verification phase: After strict data quality control and statistical analysis, 67 most significant SNPs were selected from the initial screening phase and validated in two independent Chinese populations using the Sequenom platform (northern population: 1119 cases and 1203 controls; southern population: 2505 cases and 6762 controls);
(2) The second phase of validation: 19 SNPs were selected from the first phase of validation, and further enlarged the sample size using the Sequenom platform to validate in two independent Chinese populations (northern population: 276 normal controls; southern population: 3956 normal controls). Consolidated analysis (validation of 1,119 cases and 1,479 controls in the northern population of the first and second stages; validation of 2,055 cases and 10,718 controls in the southern population of the first and second stages; total of 1,012 cases and 1,362 controls in the GWAS preliminary screening stage) was performed to identify SNPs (P5 *10~(-8)) reaching the genome-wide significant association level.
(3) Verification Phase 3: The meaningful SNPs found in the previous phase were further validated with TaqMan in the German population (1,806 cases and 3,256 normal controls).
Result:
(1) In the first stage of validation, 67 SNPs were genotyped in 3 624 Chinese Han patients and 7 965 controls. The results of the first stage of validation and GWAS analysis showed that three SNPs located at two susceptibility loci reached a significant level of genome-wide association (P5 *10-8):1 at the 1q21.3 (FLG) susceptibility locus The rs3126085 / rs11204971 of the two SNPs were P_ (combined) = 5.75 * 10 ~ (- 12) and OR = 0.83; P_ (combined) = 5.02 * 10 ~ (- 9), OR = 0.86; and 1 SNP rs7701890 (P_ (combined) = 4.33 * 10 ~ (- 8), OR = 1.23, located at 5q22.1 (TMEM232 / SLC25A46) susceptibility site.
(2) In the second stage of validation, 19 SNPs selected from the first stage of validation were typed in an additional 4232 control, and then combined with the first stage of the sample analysis. Finally, comprehensive analysis with GWAS data revealed that 7 SNPs located in three susceptibility locus regions (1q21.3, 5q22.1 and 20q13.33) reached genome-wide significance. Levels: 1) Two SNPs at 1q21.3 (FLG) susceptibility locus rs3126085/rs11204971 (P_ (combined) = 5.90 *10 ~(-12), OR = 0.82; P_ (combined) = 1.20 *10 ~(-10), OR = 0.85); 2) Four SNPs at 5q22.1 (TMEM232/SLC25A46) susceptibility locus rs7701890/rs10067777/rs13360927/rs13362 (comb P_ 13362, respectively) (combined) =2.45 x 10 ~ (-8), OR = 1.22; P (combined) = = 2.45 x 10 ~ (-8), OR = 1.22; P (combined) = = 4.02 x 10 ~ (-8), OR = 1.22; P (combined) = = 4.02 x 10 ~ (-8), OR = 1.22); 3) 20 q13.33 (TNFRSF6B / ZGPAT) at the susceptibility site of 20q13.33 (TNFRSF6B / ZGPAT) susceptibility site of 1 SNP 6010 620 (combined) =3.0 (combined) =3.0 (combined) =3.0 ~ (-10 ~ (-8 Suggestive susceptibility locus, i.e. at 1 0q21.2 (ZNF365) SNP rs2393903 (P_ (combined) =1.05 * 10~ (-7), OR=1.15) on susceptible loci.
(3) In the third stage of verification, 20q13.33 (TNFRSF6B/ZGPAT) was also found to be associated with German AD (rs6010620, P = 2.87 x 10-5, OR = 1.25).
Conclusion:
In this study, we searched for AD susceptibility genes in Chinese Han population and constructed the first Chinese Han population case-control genome-wide association analysis database. Two new AD susceptibility genes/loci were found in Chinese population: 5q22.1 (TMEM232/SLC25A46), 20q13.33 (TNFRSF6/ZGPAT). In addition, 20q13.33 (TNFRSF6B/ZGPAT) was also found to be associated with AD in the German population.
【学位授予单位】：安徽医科大学
【学位级别】：博士
【学位授予年份】：2011
【分类号】：R758.2
【参考文献】
相关期刊论文 前1条
1 ;Inhibition of histamine release from human mast cells by natural chymase inhibitors[J];Acta Pharmacologica Sinica;2004年06期
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