FcγRIIa介导的血小板激活及KLF6在血小板促肝癌生长和转移中的作用及机制研究

Clinical and experimental evidences support that platelet in the tumor microenvironment plays a crucial role in tumor growth and metastasis. But the mechanism of platelet interplay with hepatocellular carcinoma (HCC) is unclear. Our preliminary data implied that supernatant from activated platelets could reduce KLF6 expression in HepG2 cells and promote HepG2 cells proliferation. And platelets incubated with anti- FcγRIIa monoclonal antibody IV.3 abolished the effects. We speculate that platelet would promote the growth and metastasis of HCC through depressing expression of KLF6 by platelet granules releasing. We will explore the reciprocal effect between platelet and HCC cells with different metastasis capacity in vitro and in vivo. We will observe the effects of KLF-6 by its gene silence and overexpression. We also compare the phosphorylation of FcγRIIa between healthy people and patients with HCC. FcγRIIa transgene mice will be used to detect the effect of FcγRIIa in the reciprocal effect. All above would disclose the role of FcγRIIa and KLF6 in promoting HCC growth and metastasis as well as the molecular mechanism. And this would discover the new therapeutic targets for prevention and treatment of HCC.

实验研究和临床观察均提示血小板是介导肿瘤转移的重要因素，血小板与肝癌细胞相互作用促进转移的分子机理仍不清楚。我们前期研究发现肝癌HepG2细胞可激活血小板，而激活血小板的上清液降低HepG2细胞中转录因子KLF6的表达，促进HepG2细胞增殖；使用FcγRIIa特异性抗体IV.3预孵育血小板后，其促进HepG2细胞增殖及降低KLF6作用消失。申请者推测肝癌细胞通过FcγRIIa激活血小板，激活后的血小板释放颗粒物质促进肝癌细胞增殖转移，该作用与肝癌细胞中KLF6相关。本课题采用不同转移潜能的肝癌细胞株、FcγRIIa特异性抗体及KLF6基因沉默和过表达方法研究血小板与肝癌细胞的相互作用；利用FcγRIIa转基因小鼠和检测肝癌患者血小板FcγRIIa磷酸化水平研究血小板FcγRIIa的作用，以揭示FcγRIIa和KLF6在血小板对肝癌增殖转移中的作用及分子机制，为肝癌防治提供新的思路和靶点。

肝细胞肝癌即肝癌(hepatocellular carcinoma ，HCC)是临床上最常见的消化系统恶性肿瘤之一，其发病机制十分复杂，涉及细胞粘附、迁移及肿瘤血管生成等多个环节。血小板在肿瘤血管生成和肿瘤生长转移中起着至关重要的作用，血小板是由骨髓中成熟巨核细胞的胞质分割而成的，是人体血液循环中最小的无核细胞，早在19世纪60年代，Armand Trousseau指出隐匿性内脏肿瘤患者往往先出现不寻常的游走性血栓，人们开始意识到血小板与肿瘤密切相关，随后，大量临床证据表明，恶性肿瘤患者普遍存在血小板及其参数升高的情况，且多数与病情进展和恶化相关。血小板表面表达FcγRIIa，通过此受体可激活血小板。KLF6 (Krüppel-like factor 6)，一种广泛表达于正常组织的锌指转录因子，已被确认为多种肿瘤（包括肝癌）的抑癌基因。我们分别在细胞和整体动物水平，研究了激活血小板上清对不同肝癌细胞的作用，利用裸鼠荷瘤模型验证细胞研究结果，并研究了肝癌病人血小板FcγRIIa磷酸化水平及其下游信号分子，我们发现血小板激活后释放的活性物质如TGF-β，作用于肝癌细胞下调KLF6的表达从而促进肝癌生长，利用TGF-β阻断剂或miRNA敲减KLF6均抑制此现象发生。IV.3阻断FcγRIIa后，激活血小板对肝癌细胞的增殖作用减弱，其降低肝癌细胞KLF6表达的作用亦减弱。此研究结果提示有望利用阻断血小板FcγRIIa的小分子化合物或单抗药物抑制肝癌的发生发展，为肝癌防治寻找新的路径。