G-MDSC通过分泌包含miR-224的外泌体促进肾细胞癌发生肺转移

In recent years, increasing efforts were made on studying the non-immunological role of G-MDSC in tumor metastasis. However, in metastatic renal cell carcinoma (mRCC), the role of the largely expanded G-MDSC remains incompletely understood. In our previous study, we detected significantly higher expression levels of IL-6, GM-CSF, G-CSF, CXCL1, CXCL2 and CXCL5 in the more aggressive cell line Renca-L4, compared to Renca. Renca-L4 also induced higher accumulation of G-MDSC in the lungs of the renal tumor-bearing mice even earlier than obvious metastasis was detected. Moreover, these G-MDSCs directly enhanced proliferation of Renca when cocultured with them in vitro. In aim to explore the mechanism of G-MDSC in RCC metastasis, we briefly analyzed the miRNA expression profile in exosomes secreted by lung G-MDSC and interestingly, detected a high level of miR-224, which had been reported to correlate with metastasis. We thus hypothesized that G-MDSC may promote RCC metastasis via secreting miR-224. In this study, tumor orthotopic transplantation model and samples of RCC patients were used to ensure the critical effect and detailed molecular biological process of G-MDSC in RCC metastasis. We expect to further elucidate the mechanism in G-MDSC-mediated metastasis and provide novel evidence and thoughts to the clinical therapy targeting G-MDSC and secreted miRNAs in mRCC.

近年来，G-MDSC通过非免疫学机制促进肿瘤转移的作用越来越受到人们的重视，但在肾细胞癌（RCC）中，G-MDSC影响转移的具体作用和机制尚不清楚。我们前期发现，相较于Renca，高侵袭性的Renca-L4高表达IL-6、GM-CSF、G-CSF、CXCL1、CXCL2和CXCL5，可以在早期诱导大量G-MDSC聚集在肾原位癌小鼠的肺部，这些G-MDSC能在体外直接促进肾癌细胞增殖。我们初步筛选发现，肺组织浸润的G-MDSC分泌的exosome中富含促癌的miR-224。我们推测G-MDSC可能通过分泌miR-224直接促进肾癌细胞在肺部定植。本研究拟利用小鼠肾原位癌模型及RCC患者血样，确定G-MDSC及其分泌的miRNA在肾癌肺转移中的重要作用。本研究有望进一步阐明G-MDSC促进肿瘤转移的作用机制，为针对G-MDSC及其分泌的miRNA的mRCC临床治疗提供新的理论依据和思路。

肾癌是泌尿系统中恶性程度较高的肿瘤，其发病率和死亡率在全球呈上升趋势。MDSC作为一种具有免疫抑制功能的间质细胞，广泛存在于肾癌肿瘤微环境。目前关于MDSC在肾癌转移中的作用尚无明确报道。我们经尾静脉迭代植瘤构建的肾癌高肺转移细胞系Renca-L4具有更高的增殖能力、转移能力和干性，显现出更高的恶性程度。使用Renca-L4构建小鼠原位肾癌后，其肺部形成更多转移灶。Renca-L4在小鼠植瘤早期可引起更多MDSC（尤其是G-MDSC）聚集在小鼠肺组织，减少小鼠外周血和脾脏CD8+T细胞比例。Renca-L4高表达MDSC诱导或活化相关细胞因子和趋化因子，可诱导产生更多MDSC，上调其精氨酸酶1的表达和内质网应激水平，增强其免疫抑制功能。使用Ly-6G中和抗体减少G-MDSC的比例后，小鼠肺转移受到抑制；而G-MDSC回输则加剧小鼠肺转移。这些结果提示G-MDSC在肾癌肺转移中发挥重要作用。通过收集和鉴定G-MDSC外泌体，并与Renca细胞共培养，我们发现来自Renca-L4荷瘤小鼠肺组织的G-MDSC外泌体明显抑制小鼠肾癌细胞凋亡，并富集mmu-miR-690。过表达miR-690后，Renca细胞的凋亡明显受到抑制。进一步研究发现，G-MDSC分泌的miR-690可能是通过直接结合Bcl2l13 mRNA的3’UTR，在转录后水平抑制Bcl2l13的表达，从而抑制肾癌细胞凋亡，促进其在肺部定植，促进肾癌肺转移。本研究初步阐释G-MDSC在肿瘤初期聚集在小鼠肺组织（转移前灶），通过分泌包含原癌miRNA的外泌体促进肾癌细胞在此定植，从而促进肾癌肺转移的作用和分子机制，在一定程度上填补了MDSC外泌体相关研究的理论空白。