【作者】 周卫东；

【导师】 周智广；

【作者基本信息】 中南大学 ， 内科学， 2008， 博士

【摘要】 第一部分IL12B3’UTR多态性与1型糖尿病的关系目的:探讨ILl2B 3’UTR多态性与中国汉族1型糖尿病(T1DM)的关系设计:横断面病例-对照和核心家系研究方法:采用多聚酶链反应限制性片段长度多态性(PCR-RFLP)技术检测392例急性起病的T1DM患者、347例LADA患者、493例正常对照和115个核心家系IL12B 3’UTR基因型。根据是否携带胰岛细胞自身抗体,将急性起病的T1DM患者分为1A型糖尿病组和1B型糖尿病组;又根据起病年龄是否大于20岁,将急性起病的T1DM患者分为青少年急性起病的T1DM(JO)组和成人急性起病的T1DM(AO)组。根据GADA抗体滴度是否大于0.3,将LADA患者分为高滴度的LADAl组和低滴度的LADA2组。结果:1.成人急性起病的T1DM患者IL12B 3’UTR基因型AC+CC频率(76.6%vs.67.8%,P=0.026)和C等位基因频率明显高于正常对照组(50.2%vs.44.2%,P=0.047),前者与青少年急性起病的T1DM患者比较,IL12B 3’UTR基因型分布(P=0.032)和C等位基因(50.2%vs.41.7%,P=0.017)频率有显著性差异。2.仅携带GADA的1A型糖尿病患者IL12B 3’UTR基因型AC+CC频率(80.5%vs.67.8%,P=0.002)和C等位基因(51.2%vs.44.2%,P=0.027)频率明显高于正常对照组,前者与携带IA-2A或IAA或多个胰岛自身抗体1A型糖尿病患者比较,IL12B 3’UTR基因型AC+CC频率(80.5%vs.63.3%,P=0.001)和C等位基因51.2%vs.40.2%,P=0.008)频率有显著性差异。3.起病年龄＜40岁LADA患者AC基因型明显高于正常对照(64.1%vs.47.1%,P=0.018)和起病年龄≥40岁LADA患者(64.1%vs.45.5%,P=0.016),但前者C等位基因频率与正常对照和≥40岁LADA患者比较均无明显差异(P＞0.05)。4.以115个核心家系为基础的传递不平衡检验(TDT)未发现C等位基因优势传递给T1DM患者(P＞0.05);根据先证者起病年龄、性别及携带的HLA-DQ单体型分层核心家系,也未发现C等位基因优势传递给T1DM患者(P＞0.05)。结论:1.ILl2B 3’UTR多态性与中国汉族T1DM可能有关,与迟发的急性起病T1DM和早发的LADA有关。2.1A型糖尿病中,IL12B 3’UTR多态性可能与仅GADA阳性的T1DM遗传易感性有关。第二部分SUMO4(M55V)多态性与1型糖尿病的关系目的:探讨SUMO4(M55V)多态性与中国汉族1型糖尿病的关系。设计:横断面病例.对照和核心家系研究方法:采用多聚酶链反应限制性片段长度多态性(PCR-RFLP)技术检测389例急性起病的T1DM患者、359例LADA患者、532例正常对照和115个核心家系SUMO4(M55V)基因型。根据是否携带胰岛细胞自身抗体,将急性起病的T1DM患者分为1A型糖尿病组和1B型糖尿病组;又根据起病年龄是否大于20岁,将急性起病的T1DM患者分为青少年急性起病的T1DM(JO)组和成人急性起病的T1DM(AO)组。根据GADA抗体滴度是否大于0.3,将LADA患者分为高滴度的LADA1组和低滴度的LADA2组。结果:1.急性起病的T1DM患者SUMO4(M55V)基因型GG检出率明显高于正常对照(11.3%vs.7.1%,P=0.022)。2.1A型糖尿病患者SUMO4(M55V)基因型GG检出率明显高于正常对照(11.7%vs.7.1%,P=0.038)。携带IA-2A或IAA或多个胰岛自身抗体的T1DM患者GG频率明显高于正常对照(13.6%vs.7.1%,P=0.030)。3.起病年龄小于45岁LADA2患者与正常对照比较,基因型(P=0.037)分布及等位基因(P=0.047)频率均有统计学意义,前者AA基因型频率明显降低(32.8%vs.49.6%,P=0.016)。4.以核心家系为基础的传递不平衡检验(TDT)未发现G等位基优势传递给T1DM患者(P＞0.05)。根据先证者起病年龄、性别及携带的HLA-DQ单体型分层核心家系,也未发现G等位基因优势传递给T1DM患者(P＞0.05)。结论:1.SUMO4(M55V)多态性与中国汉族急性起病的T1DM可能有关联,这种关联性主要与携带IA-2A和(或)IAA的TIDM有关。2.SUMO4(M55V)多态性可能与GADA滴度低且年轻起病的LADA患者有关。第三部分HLA-DQ、IL12B及SUMO4基因在1型糖尿病中的相互作用目的:探讨HLA-DQ、IL12及SUMO4基因在1型糖尿病中的相互作用。设计:横断面、病例.对照研究方法:采用PCR-测序法检测HLA-DQ基因型,采用PCR-RFLP技术检测IL12B 3’UTR和SUMO4(M55V)多态性。结果:1.急性起病T1DM的HLA-DQ易感单体型是DQA1~*03-DQB1~*0303、DQA1~*05-DQB1~*0201及DQA1~*03-DQB1~*0401。DQA1~*05-DQB1~*0201易感性最强(OR=5.287),其次是DQA1~*03-DQB1~*0401(OR=3.579)和DQA1~*03-DQB1~*0303(OR=2.513)。保护性单体型是DQA1~*0102-DQB1~*0602(OR=0.328)。2.LADA的HLA-DQ易感单体型是DQA1~*03-DQB1~*0303、DQA1~*05-DQB1~*0201及DQA1~*03-DQB1~*0401,DQA1~*05-DQB1~*0201易感性最强(OR=2.939),其次是DQA1~*03-DQB1~*0401(OR=2.747)和DQA1~*03-DQB1~*0303(OR=1.465);LADA患者DQA1~*0102-DQB1~*0602频率明显低于正常对照组,但无统计学差异(P=0.088)。3.不携带DQA1~*05-DQB1~*0201和DQA1~*03-DQB1~*0401急性起病的T1DM患者,IL12B 3’UTR基因型AC+CC(76.4%vs.67.2%,P=0.044)和C等位基因频率(50.5%vs 41.7%,P=0.016)明显高于正常对照。4.携带HLA-DQ易感单体型急性起病的T1DM患者,同时携带SUMO4(M55V)基因型AA、GG、AG频率均明显高于正常对照组(P＜0.0001),其中,同时携带GG基因型(OR=5.851)相对危险度高于不携带GG基因型(OR=3.307)。5.急性起病的T1DM患者同时携有SUMO4(M55V)基因型GG和IL12B 3’UTR基因型AC频率明显高于正常对照组(7.4%vs.2.2%,OR=3.486,P=0.001):急性起病的T1DM(9.7%vs.14.4%,OR=0.641,P=0.049)和LADA(9.3%vs.14.4%,OR=0.606,P=0.035)患者同时携有SUMO4(M55V)基因型AA和IL12B 3’UTR基因型CC频率明显低于正常对照组。结论:1.DQA1~*03-DQB1~*0303、DQA1~*05-DQB1~*0201及DQA1~*03-DQB1~*0401是中国汉族T1DM的HLA-DQ易感单体型。DQA1~*0102-DQB1~*0602是急性起病的T1DM保护性单体型。2.在急性起病T1DM患者中,IL12B 3’UTR遗传易感性是对HLA-DQ单体型危险性的补充。3.SUMO4(M55V)对急性起病T1DM的易感性不是独立的,而是与携带HLA-DQ易感单体型有关。4.SUMO4(M55V)和IL12B 3’UTR存在相互作用。SUMO4(M55V)基因型GG与IL12B 3’UTR基因型AC对T1DM易感性具有协同作用,SUMO4(M55V)基因型AA抑制IL12B 3’UTR基因型CC对T1DM的易感性。更多还原

【Abstract】 Part1 The relationship between IL12B3’UTR polymorphism and type 1 diabetes mellitusObjective:To investigate the association of the 3’ untranslated region(UTR) polymorphism of interleukin-12 p40 gene(IL12B) with type 1 diabetes mellitus(T1DM) in Chinese Han population.Design:Cross-sectional case-control and family-based association studyMethods:A total of 392 acute-onset type 1 diabetes patients,347 LADA patients,493 nondiabetic unrelated controls and 115 trio families comprising both parents and an affected offspring were typed for the IL12B3’UTR polymorphisms by polymerase chain reaction restriction fragment length polymorphism(PCR-RFLP) method.The patients with acute-onset type 1 diabetes were divided into type 1A diabetes and type 1B diabetes according to islet cell autoantibodies positive or negative, juvenile-onset group(JO) and adult-onset group(AO) according to the age at onset higher than 20 years or not,and LADA patients were divided into LADA1 and LADA2 according to the GADA titers higher than 0.3 or not.Results:1.The AC and CC genotypes had a higher frequency in AO patients than in control subjects(76.6%vs.67.7%,P=0.026),and C allele frequency was significantly different from controls(50.2%vs.44.2%,P =0.047).The genotype distribution(P=0.032) and C allele frequency (50.2%vs.41.7%,P=0.017) was significantly different between AO and JO patients.2.The patients only with GADA had a higher frequency of the AC and CC genotypes than controls(80.5%vs.67.8%,P=0.002) and those with IA-2A or IAA or at least two islet cell autoantibodies(80.5%vs. 63.3%,P=0.001),and the C allele had a higher frequency in the former than the latter two(51.2%vs.44.2%,51.2%vs.40.2%,respectively,all P＜0.05).3.LADA patients,who were younger than 40 years old at onset age, had a higher frequency in AC genotype than those older than 40 years old at onset age(64.1%vs.45.5%,P=0.016) and controls(64.1%vs. 47.1%,P=0.018),but the C allele frequency in the former was no difference from the latter two(all P＞0.05).4.Transmission disequilibrium test analysis,was based on 115 trio families,revealed no preferential transmission in C allele(P＞0.05), stratifying the families according to HLA-DQ haplotype,sex,and age-at-onset of the proband,no significant distortion was found in allele transmissions for the 3’UTR polymorphism(all P＞0.05). Conclusion:1.In Chinese Han population,IL12B 3’UTR polymorphism may be associated with T1DM,and it is associated with late-onset diabetes in acute-onset T1DM patients and is related to early-onset diabetes in LADA patients.2.In the acute-onset type 1A diabetes patients,the C allele of IL12B 3’UTR may be associated with patients only with GADA. Part2 The relationship between SUMO4(M55V) polymorphism and type 1 diabetes mellitusObjective:To investigate the association of SUMO4(M55V) polymorphism with type 1 diabetes(T1DM) in Chinese Han population.Design:Cross-sectional case-control and family-based association studyMethods:A total of 389 acute-onset type 1 diabetes patients,359 LADA patients,532 nondiabetic unrelated controls and 115 trio families comprising both parents and an affected offspring were typed for the SUMO4(M55V) polymorphisms by polymerase chain reaction restriction fragment length polymorphism(PCR-RFLP) method.The patients with acute-onset type 1 diabetes were divided into type 1A diabetes and type 1B diabetes according to islet cell autoantibodies positive or negative,juvenile-onset group(JO) and adult-onset group(AO) according to the age at onset higher than 20 years or not,and LADA patients were divided into LADA1 and LADA2 according to the GADA titers higher than 0.3 or not.Result:1.The GG genotype of SUMO4(M55V) had a higher frequency in acute-onset T1DM patients than in control subjects(11.3%vs.7.1%, P=0.022). 2.In type 1A diabetes patients,the GG genotype had a higher frequency than in control subjects(11.7%vs.7.1%,P=0.038).There was a significant difference in the GG genotype frequency between the patients with IA-2A or(and) IAA and control subjects(13.6%vs.7.1%, P=0.030).3.In LADA2 patients diagnosed before age 45 years,the genotype distribution(P=0.037) and G allele frequency(P=0.047) was significantly different from control subjects,and the AA genotype frequency was lower than controls(32.8%vs.49.6%,P=0.016).4.Transmission disequilibrium test analysis,was based on 115 trio families,revealed no preferential transmission in G allele(P＞0.05), stratifying the families according to HLA-DQ haplotype,sex,and age-at-onset of the proband,no significant distortion was found in allele transmissions for SUMO4(M55V) polymorphism(all P＞0.05).Conclusions:1.SUMO4(M55V) polymorphism may be associated with acute-onset type 1 diabetes in Chinese Han population,and the association is concentrated in patients with IA-2A and/or IAA.2.SUMO4(M55V) polymorphism may be associated with LADA patients with GADA of lower titters and younger age at onset. Part 3 The interactive effects of HLA-DQ,IL12B 3’UTR and SUMO4(M55V) on type 1 diabetes mellitusObjective:To investigate the interactive effects of HLA-DQ,IL12B 3’UTR and SUMO4(M55V) on type 1 diabetes mellitus(T1DM).Design:Cross-sectional and case-control study.Methods:The polymorphisms of HLA-DQ were typed by PCR sequencing-based typing method,IL12B 3’UTR and SUMO4(M55V) were typed by PCR-RFLP method.Results:1.In acute-onset T1DM patients,the susceptible haplotypes of HLA-DQ gene were DQA1~*03-DQB1~*0303,DQA1~*05-DQB1~* 0201 and DQA1~*03-DQB1~*0401,and DQA1~*05-DQB1~*0201 was the highest risk haplotype(OR=5.278),DQA1~*03-DQB1 ~*0401 and DQA1~*03-DQB1~*0303 were moderate risk(OR= 3.579) and low risk haplotype (OR=2.513),respectively.The protective haplotype was DQA1~*0102-DQB1~*0602.2.In LADA patients,the susceptible haplotypes of HLA-DQ gene were DQA1~*03-DQB1~*0303,DQA1~*05-DQB1~*0201 and DQA1~*03-DQB1~*0401,and DQA1~*05-DQB1~*0201 was the highest risk haplotype (OR=2.939),DQA1~*03-DQB1~*0401 and DQA1~*03-DQB1~* 0303 were moderate risk(OR=2.747) and low risk haplotype(OR=1.465),respecttively. The protective DQA1~*0102-DQB1~*0602 was non-significant decrease than control subjects(P=0.088).3.In the acute-onset T1DM patients without DQA1~*05-DQB1~*0201 or DQA1~* 03-DQB1~*0401,the AC and CC genotype frequency of IL12B 3’UTR was significantly higher than control subjects without DQA1~*05-DQB1~*0201 or DQA1~*03-DQB1~*0401(76.4%vs.67.2%, P=0.044),and C allele frequency was higher than controls(50.5%vs. 41.4%,P=0.016).4.In acute-onset T1DM patients with HLA-DQ susceptibility haplotypes, the frequencies of SUMO4(M55V) genotype AA,GG,AG were higher than controls,respectively(P＜0.0001),and compared with the patients without GG(OR=3.307),there is an increase in the relative risk (OR=5.851) in the patients with GG.5.In the acute-onset T1DM patients,the concurrence frequency of SUMO4(M55V) GG genotype and IL12B 3’UTR AC genotype was significantly higher than controls(7.4%vs.2.2%,OR=3.486,P=0.001), and the concurrence frequency of SUMO4(M55V) AA genotype and IL12B 3’UTR CC genotype was significantly lower(9.7%vs.14.4%, OR=0.641,P=0.049).In LADA patients,the concurrence frequency of SUMO4(M55V) AA genotype and IL12B 3’UTR CC genotype was significantly lower than controls(9.3%vs.14.4%,OR=0.606,P=0.035).Conclusion:1.DQA1~*03-DQB1~*0303,DQA1~*05-DQB1~*0201 and DQA1~*03- DQB1~*0401 of HLA-DQ gene are susceptible haplotypes in T1DM patients of Chinese Han population,and DQA1~*0102-DQB1~*0602 is protective haplotype in the acute-onset T1DM patients.2.In the acute-onset T1DM patients,IL12B 3’UTR polymorphism may be a supplementary risk factor to T1DM in conjunction with HLA-DQ haplotypes.3.SUMO4 M55V is associated with TIDM in association with HLA-DQ susceptibility haplotypes,but not by itself.4.The interactive effects of SUMO4(M55V) polymorphism and IL12B 3’UTR polymorphism on T1DM are not independent.SUMO4 (M55V) genotype GG and IL12B 3’UTR genotype AC is synergistic on susceptibility to T1DM,SUMO4(M55V) genotype AA changes genetic susceptibility conferred by IL12B 3’UTR genotype CC to T1DM.更多还原