HLA-A33表型、TNF-α/TNF-RⅡ基因多态性和肠道病毒71型脑炎的相关性研究

Association of HLA-A33 phenotype,TNF-α/TNF-RⅡ promoter polymorphisms with the susceptibility to enterovirus 71 infection

目的研究HLA-A33表型,TNF-α/TNFRⅡ单核苷酸多态性与肠道病毒71型(EV71)感染遗传易感性的关系,探讨不同基因表型对EV71感染患病风险的影响。方法收集2009年9月至2010年10月EV71检测为阳性的急性期中枢神经系统感染患儿123例,根据病情轻重分成轻症组、重症组;提取患儿外周血白细胞的基因组DNA,用序列特异性引物-聚合酶链反应(PCR-SSP)技术检测EV71感染患儿及正常对照儿童HLA-A33表型、TNF-α-308位点的多态性和限制性片段长度多态性-聚合酶链反应(PCR-RFLP)检测TNFRⅡ+196位点的多态性。结果 EV71感染患儿中HLA-A33表型阳性率(24.39%)与健康儿童组阳性率(11.82%)的差异有统计学意义(χ2=6.099,P<0.05)。EV71中枢感染患儿TNF-α-308位点A等位基因频率明显高于正常对照组(χ2=6.367,P<0.05),感染组TNF-α-308GG基因型分布频率显著低于正常对照儿童(χ2=5.393,P<0.05);而轻症组与重症组相比,其差异无统计学意义(P>0.05)。TNFRⅡ+196位点T等位基因频率在EV71感染组与对照组、轻症组与重症组之间的差异均无统计学意义(P>0.05)。HLA-A33(+)患儿中,EV71感染组TNFRⅡ+196TG基因型频率明显高于正常对照组(χ2=3.866,P<0.05),而在HLA-A33(-)儿童中,其差异无统计学意义。结论 HLA-A33表型、TNF-α-308位点基因多态性与EV71中枢感染有关,TNF-α-308GG基因型可能为儿童不易感染EV71的保护基因。TNFRⅡ+196位点基因多态性与EV71感染无关;TNFRⅡ+196TG基因型在一定程度上升高了HLA-A33(+)患儿EV71中枢感染的发病概率。

Objective To investigate the genetic susceptibility factors of enterovirus71 （EV71） infection by studying the interactions among HLA-A33 phenotype and TNF-α/TNFR lI by SNP and the information about the effects of different genes on risk of encephalitis of EV71 infection. Methods Reverse transcription polymerase chain reaction（RT-PCR） was used to detect the specific primers of EV71 in the faeces of patients with EVT1 infection. Genomic DNA of white blood cell were extracted,then PCR-SSP were used to determine phenotype of HLA-A33 and genotype for the A/G polymorphism at-308 position of TNF-α gene and PCR-RFLP was used to genotype for the T/ G polymorphism at ＋ 196 position of TNFR II gene. Results Compared with that of the normal children, there were obvious statistical significance in EV71 infection patients of the rate of HLA-A33 phenotype （ x^2 = 6. 099, P 〈 0.05）. In EV71 infection groups, the genotype frequency of TNF-et-308GG was significantly lower than that in normal controls（x^2 =5. 393 ,P 〈0. 05）, and the TNF-α-308A allele was more common than in normal group（ X^2 = 6. 367, P 〈 0. 05）, although there were no obvious statistical significance between mild and critical case groups. There were no obvious statistical significance in the frequencies of TNFR II ＋ 196T allele between the patients and control group （P 〉0. 05）. In EV71 group,TG genotype of TNFR II ＋ 196 was more common than in control group based on HLA-A33 phenotype（ ＋ ） （ x^2 = 3. 866, P 〈 0. 05 ）. Conclusions The polymorphism of HLA-A33, TNF-α-308 single nueleotide may correlate with EV71 infection ,maybe TNF-α-308GG is the protective genotype which protect children keep away from EV71. Although there is no obvious statistical significance in EV71 infection patients by SNP, the TNFR 11 ＋ 196（TG）may increase the risk of EV71 infection based on HLA-A33 phenotype（ ＋ ）. [Key words] Enterovirus A,human; HLA antigens; Tumor necrosis factors; Receptors,tumor necrosis factor; Polymorphism,single nucleotide