微囊泡运载miRNA介导胃癌细胞与成纤维细胞之间双向调控机制研究

Tumors can regulate and transform the tumor microenviroment actively, malignant transformation of microenviroment is very important for tumor development. It is a new potential way of controlling the microenviroment by regulating the stroma with microRNAs carried by microvesical from gastric cancer cells. It remains to be elucidated the mechanism of regulating fibroblast by microRNAs in microvesical from gastric cancer cells. In our previous studies, 5 specific microRNAs secreted by gastric cancer had been identified, and the prognosis of patients with high level of miR-27a was poor. miR-27a enriched in microvesical secreted by gastric cancer cell significantly increased the migration capability of fibroblast and changed the genes profile. Bioinformatics analysis indicated CSRP2 was the target gene of miR-27a, and preliminary experiment results revealed that miR-27a could down regulate the protein expression of CSRP2. In this proposal, we will further verify whether gastric cancer cells can transform normal fibroblast into cancer associated fibroblast by miR-27a in microvesical, and whether the latter can regulate the gastric cancer cells reversely. We also will explore the bilateral regulation mechanism and effects between the gastric cancer cells and fibroblast. Our study will not only be important to reveal a novel microenviromental regulation mechanism of gastric cancer, but also provide a new direction and a target for gastric cancer targeting therapy.

肿瘤对其微环境存在主动调控和转化，恶性转化的微环境对肿瘤的发展具有重要作用。胃癌通过分泌微囊泡（MV）运输miRNAs作用于间质是其对微环境调控的一种新途径，胃癌通过MV运输miRNAs对间质核心成分成纤维细胞的机制尚无报道。课题组已发现胃癌分泌5个特征性miRNAs，临床病例分析发现miR-27a高表达者预后不良。胃癌细胞通过MV富集miR-27a并引起成纤维细胞迁移能力上升和基因特征变化。生物信息学提示CSRP2是miR-27a的靶基因，已初步验证miR-27a可下调成纤维细胞CSRP2基因蛋白表达。本课题拟进一步验证胃癌细胞能否通过MV运输miR-27a使成纤维细胞向癌相关成纤维细胞转化并反向调节胃癌生物学特征，探讨胃癌细胞和成纤维细胞之间的相互作用的分子机制和效应。本课题的实施不仅对揭示胃癌新的微环境调控机制有重要意义，更为胃癌靶向治疗提供了新的方向和靶点。

胃癌（GC）的恶性生物学行为不仅取决于癌细胞本身，而且还受微环境的调控。成纤维细胞在肿瘤微环境中占很大比例，并促进疾病的发展。目前，越来越多的证据表明外泌体可以作为细胞间转运系统来传递其内含物，特别是微RNA（miRNA）。首先，我们通过qRT-PCR检测到从胃癌细胞中分离出的外泌体中miR-27a的高表达水平。建立了MiR-27a过表达的体外和体内模型，以研究通过Western blot观察到的癌相关成纤维细胞的转化，以及使用CCK8和Transwell方法对胃癌细胞的恶性行为。此外，使用CCK8和Transwell方法，通过下调成纤维细胞中CSRP2的表达来评估胃癌的恶性程度。在这项研究中，我们发现来自GC细胞的外泌体中miR-27a的水平很高。发现miR-27a具有致癌基因的功能，不仅可以诱导成纤维细胞重编程为癌相关的成纤维细胞（CAF），而且还可以促进体内外癌细胞的增殖，运动和转移。相反，miR-27a过度表达的CAF可以多效性地增加GC细胞的恶性行为。首次，我们揭示了CSRP2是miR-27a的下游靶标。 CSRP2的下调可能会增加GC细胞的增殖和运动能力。因此，该报告表明，源自GC细胞的外泌体中的miR-27a对微环境具有至关重要的影响，并且可以用作治疗GC的潜在治疗靶标。