核受体LXR促进microRNA-206表达的机制及在抗HCC中的作用

Liver X receptor(LXR) and microRNA 206(miR-206) were the important anti-tumor molecules,and the levels were decreased in hepatocellular carcinoma (HCC). However, it is unclear whether LXR plays its anti-tumor role via up-regulating miR-206. Our preliminary results for the first time showed that LXR and miR-206 levels were decreased in HCC patient samples and HCC cell lines. Furthermore, we foud that LXR could significantly induce miR-206 expression and enhance its transcriptional activity. Bioinformatic analysis showed that there were potential binding sites for LXR in human miR-206 5′flanking region. In the present study, we will identify the exact binding site of LXR in the 5′flanking region of miR-206 in order to confirm that miR-206 is a novel target gene of LXR, which will reveal the mechanism of LXR upregulates miR-206 expression. Subsequently, we will investigate the significance of LXR-mediated miR-206 upregulation in the suppression of HCC in vitro. Moreover, we will evaluate the importance of miR-206 upregulation in LXR-mediated anti-HCC effect in vivo. The anticipated results of the present study may reveal that the LXR/miR-206 pathway is a novel anti-HCC mechanism of LXR, and may provide a new sight for exploring novel therapeutic strategy to prevention and treatment of HCC by targeting LXR/miR-206 pathway.

肝X受体（LXR）和非编码小RNA 206（mir-206）均是重要的抗肿瘤分子，且在肝细胞癌（HCC）中表达低于正常，但LXR能否通过调节mir-206表达而抗肿瘤，目前尚不清楚。鉴于此，本项目在发现临床HCC标本和HCC细胞中，mir-206和LXR表达水平降低；在人和小鼠HCC细胞中，活化LXR后能明显促进mir-206表达；生物信息学预测mir-206的5＇调控区有LXR可能结合位点的基础上，拟借助报告基因检测、EMSA、ChIP等实验，明确mir-206是LXR的一新靶分子，并阐明LXR促进mir-206表达的分子机制；经流式细胞检测、细胞凋亡检测等体外实验以及RNAi等体内实验，阐明LXR促进mir-206表达的机制及在抗HCC中的作用，从而揭示"LXR-mir-206通路"是LXR抗HCC的一新机制，为深入研究以"LXR-mir-206"为靶标的抗HCC策略提供新的科学依据。

肝癌（HCC）由于其发病率高，转移性复发和死亡率高，HCC已成为威胁人体健康的重要因素。微小RNA（miRNA）是一类内源性19-22nt非编码RNA，通过促进靶mRNA的降解或抑制靶mRNA转录后水平进行调控基因的表达。miRNA可以作为肿瘤抑制基因来抑制细胞增殖，侵袭，迁移和细胞凋亡，包括肝细胞癌等。研究结果表明：在临床HCC患者癌组织标本及肝癌细胞系中LXR和miR-206均为低表达。活化LXR后miR-206表达显著增加， 而miR-206 的靶基因CDK6及CyclinD1表达蛋白和mRNA表达明显下降，并抑制了细胞的增值；在荷瘤鼠模型及组织标本中，当活化LXR后，miR-206表达显著增加，CyclinD1表达下降，肿瘤体积减小。这一研究结果说明LXR-mir-206通路”是LXR抗HCC的一新机制，为深入研究以“LXR-mir-206”为靶标的抗HCC策略提供新的科学依据。与此同时，我们还探讨长链非编码 RNA H19对 miR-124-3p 表达的影响及在肝癌细胞增殖中的作用，结果表明：lncRNA H19 和 miR-124-3p在肝癌细胞系及肝癌组织中均低表达；过表达 lncRNA H19 后 miR-124-3p 表达显著增加，CCND1 蛋白和 mRNA 表达明显下降，细胞增殖受到抑制；过表达 lncRNA H19 同时干扰miR-124-3p 后，细胞增殖抑制作用减弱，这一结果说明了长链非编码 RNA H19 通过促进 miR-124-3p 表达，进而抑制其靶基因 cyclinD1 表达而抑制肝癌细胞增殖。另外。我们对CDK9是否参与miR-206介导的人肝细胞癌增殖和凋亡的抑制作用进行了研究，结果表明：在肝癌细胞中CDK9高表达，而miR-206低表达。miR-206通过与CDK9的3’UTR结合而下调其表达，进而抑制肝癌细胞的增殖，阻滞细胞周期并诱导细胞凋亡。这一结果说明在抗HCC策略中，miR-206可阻断CDK9相关信号通路，这为深入研究以“mir-206-CDK9”为靶标的抗HCC策略提供新的治疗方案。