骨肉瘤来源Exosome通过miR-27a-3p胞间转移调控肺部炎症微环境及促进骨肉瘤肺转移的作用机制研究

Osteosarcoma pulmonary metastasis greatly influence the prognosis of patients with osteosarcoma, and the mechanism remains undefined with no effective early diagnosis index has been established by far. Exosomes released by tumor cells are membranous bodies containing protein and RNA, but the role of exosomes in the mechanisms of osteosarcoma metastasis remains largely unknown. Our previous studies found that, circulating exosomes in osteosarcoma patients’ peripheral blood is rich in miR-27a-3p, and exosomal miR-27a-3p can promote the seceration of inflammatary factors in pulmonary cells. Therefore, our current study aims to explore the exosome-mediated microRNA transfer mechanism between cells, and the regulatory role of these transferred microRNAs on their target genes. Further studies include the in vitro analysis of the functions of exosome enriching in or deleting miR-27a-3p and the in vivo analysis to the osteosarcoma pulmonary metastasis after transferring exosomes into rat tumor models to reveal the role of circulating exosome and miR-27a-3p in osteosarcoma, and the foundation of non-invasive diagnosis system. Our plan may add new knowledge to the mechanism of osteosarcoma pulmonary metastasis and find some new strategy for osteosarcoma diagnosis and therapy.

骨肉瘤肺部转移严重影响患者预后，目前缺乏充分的机制研究及有效的早期诊断指标，尤其是循环肿瘤Exosome在骨肉瘤肺部转移中的意义不明。本课题组前期研究发现，骨肉瘤患者外周血循环exosomes中富含miR-27a-3p，且exosomes能够促进miR-27a-3p胞间转移并抑制肺上皮细胞表达炎症抑制因子TNFAIP3，导致肺上皮细胞过度分泌炎症因子。这一作用可能对肺部炎症微环境营造和骨肉瘤转移细胞定植具有重要意义。因此，本项目拟首先验证exosomal miR-27a-3p与骨肉瘤的动态相关性；随后体外研究Exosome介导miR-27a-3p胞间转移的机制及其对肺上皮细胞的调控作用；在体研究miR-27a-3p相关exosomes对骨肉瘤肺转移灶的影响，最后在临床样本中分析exosomal miR-27a-3p对骨肉瘤的诊断价值，从而为骨肉瘤肺转移机制和无创诊断研究提供新的参考。

肺转移是骨肉瘤（OS）发生发展中常见的转移现象，但其分子机制仍未完全清楚。外泌体在肿瘤转移中起着重要作用，尤其是OS的肺转移。然而，外泌体来源的长链非编码RNA（lncRNA）在OS肺转移中的机制尚不清楚。在这里，我们报道了OS外泌体来源的linc00881促进OS的肺转移，并诱导正常的肺成纤维细胞转化为肿瘤相关成纤维细胞（CAFs）。我们发现OS细胞分泌外泌体中的linc00881可以通过ceRNA机制与miR-29c-3p结合，调节HFL-1细胞中基质金属蛋白酶2（MMP2）表达，活化肺成纤维细胞中NF-κB信号通路。最后，我们发现了linc00881 / miR-29c-3p / MMP2信号轴调控OS细胞分泌IL-1β、IL-6和IL-8等促炎细胞因子。这些结果表明，OS来源的外泌体可以介导OS细胞与肺成纤维细胞之间的串扰，并影响OS的肺转移。我们通过对外泌体进行转录组测序，选取高表达microRNA进行研究。研究发现miR-125b-5p在OS中低表达，通过结合RAB3D抑制OS迁移和增殖。同时RAB3D对OS权益和增殖具有促进作用。 另外，我们研究发现miR-27a-3p作用于肺成纤维细胞内靶基因TNFAIP3，活化下游NF-κB通路，进而促进OS肺转移，并维持OS肺转移微环境的炎症状态。本研究为骨肉瘤肺转移的治疗提供了潜在的靶点。