肝细胞癌相关miRNAs遗传变异与疾病易感性及其功能的研究

The pathogenesis of hepatocellular carcinoma (HCC) is regarded as the result of the interaction between environmental and genetic factors, however, the underlying mechanism is still unclear. MicroRNAs (miRNAs) play a crucial role in hepatocellular carcinogenesis through negative regulation of their target genes. In our preliminary work, the expression profiles of miRNAs were analyzed using a GeneChip miRNA array on HCC, and the six most differentially expressed miRNAs (hsa-miR-182-5p，hsa-miR-224， hsa-miR-3651，hsa-miR-1301-3p，hsa-miR-4793-3p and hsa-miR-106b-3p）were screened out, and validated by performing a qPCR assay. The results implied that these miRNAs may participate in HCC development. Hence, we speculated that genetic variants in these miRNAs can have profound effects on miRNA expression and function, which would affect regulation on the target genes of miRNAs, and thereby contribute to HCC. 40 SNPs located in the eight miRNA genes then were predicted and screened by using bioinformatic approaches. Based on the results mentioned above, this case-control study on the molecular epidemiology will be conducted to investigate the associations between the 40 SNPs and the susceptibility to HCC and to identify the interaction effects of gene-gene and gene-environment for HCC risk. Furthermore, molecular biology techniques will be conducted to examine functional relevance of the SNPs associated with HCC susceptibility, and potential biomarkers of susceptibility to HCC will be identified. This study will be helpful to elucidate molecular mechanisms by which miRNAs are involved in HCC development and provide evidence for screening high risk population, personal prevention, early detection and treatment of HCC.

肝癌是遗传和环境因素相互作用的结果，但确切机制尚不明确。MiRNAs通过对靶基因表达的负性调控，在肝细胞癌（HCC）发生发展中起关键作用。本课题组前期通过miRNA芯片表达谱研究，经qPCR实验验证，筛选出HCC组织差异表达最显著的6个miRNAs，它们可能与HCC发生发展有关。这些miRNAs的遗传变异可能通过影响miRNA表达水平和功能而影响对靶基因的调控从而与HCC易感性相关。我们用生物信息学方法筛选出了40个上述miRNAs基因SNPs。本项目基于前期研究结果，运用病例对照研究结合实验室检测的方法，分析上述SNPs与HCC易感性的关联，探讨基因-基因、基因-环境交互作用在HCC发病中的作用；采用分子生物学方法探讨SNPs的生物学功能，并筛选出潜在易感性标志物。本研究为揭示miRNAs在HCC发生中的作用及其机制提供科学依据，为HCC易感人群筛查及个体化预防、早期诊断和治疗等提供参考

肝细胞癌（HCC）是环境因素和遗传因素相互作用的结果，但确切机制尚不明确。miRNAs通过对靶基因表达的负性调控，在HCC发生发展中起关键作用。在前期工作中，我们发现并验证了6个在HCC中表达显著上调的miRNAs（miR-182-5p，miR-224，miR-3651，miR-1301-3p，miR-4793-3p和miR-106b-3p）。应用生物信息学方法筛选出了36个上述miRNAs相关SNPs。我们采用病例-对照研究方法，以选取的863例确诊的HCC新发病例和908例非肿瘤患者为研究对象，对候选SNPs进行基因分型，综合应用单因素、多因素logistic回归分析统计学方法，评价这些SNPs及其与HBV感染、饮酒、吸烟等环境暴露因素交互作用与HCC易感性的关系。结果发现，1. miR-182 rs4541843与HCC发病风险相关，在共显性模型中，与携带GG基因型的个体相比，携带AG基因型的个体罹患肝癌的风险增加。在显性模型中，携带AG/AA基因型的个体罹患肝癌的风险增加。分层分析结果表明，A等位基因是广西地区年龄>49岁，男性，无饮酒史等人群HCC的危险等位基因。2. miR-106b rs2261360与HCC易感性相关，在共显性模型中，与携带GG基因型的个体相比，携带TG基因型的个体罹患肝癌的风险降低。在显性模型中，携带TG/TT基因型的个体罹患肝癌的风险降低。但未发现miR-106b rs1527423位点与HCC发病风险之间存在统计学关联。分层分析结果表明，T等位基因是广西地区年龄≤49岁、男性、无吸烟史、无饮酒史及HBV感染阳性等人群的保护因素，且rs2261360多态性与HBV感染存在交互作用。考虑到miR-106b与其靶基因ATF5rs1527423多态性在HCC发病风险中可能存在潜在联合作用，我们将rs2261360 GG基因型、rs1527423 GA/GG基因型和rs8647 GG基因型定义为假设危险基因型，联合作用分析结果表明，联合危险基因型在病例组和对照组中的分布差异具有统计学意义；与不携带危险基因型的个体相比，携带1-3个危险基因型的个体发生HCC的风险显著增加，尤其是在男性，无吸烟史，无饮酒史及HBV感染阳性的人群中。该研究结果提示，miRNAs及其与靶基因SNP之间的联合作用以及基因与环境的交互作用对HCC的遗传易感性有影响。