Dai Guangyao, Yao Xiaoguang, Zhang Yubin, Gu Jianbin, Geng Yunfeng, Xue Fei, Zhang Jingcheng
First Hospital of Shijiazhuang City, Department of General Surgery, Shijiazhuang, China.
Hebei University of Chinese Medicine, Hebei Key Laboratory of Integrative Medicine on Liver-kidney Patterns, Shijiazhuang, China; Hebei University of Chinese Medicine, College of Integrative Medicine, Shijiazhuang, China.
Bull Cancer. 2018 Apr;105(4):336-349. doi: 10.1016/j.bulcan.2017.12.009. Epub 2018 Feb 26.
Cancer-associated fibroblasts (CAFs) contribute to the proliferation of colorectal cancer(CRC) cells. However, the mechanism by which CAFs develop in the tumor microenvironment remains unknown. Exosomes may be involved in activating CAFs.
Using a miRNA expression profiling array, we determined the miRNA expression profile of secretory exosomes in CRC cells and then identified potential miRNAs with significant differential expression compared to normal cells via enrichment analysis. Predicted targets of candidate miRNAs were then assessed via bioinformatics analysis. Realtime qPCR, western blot, and cell cycle analyses were performed to evaluate the role of candidate exosomal miRNAs. Luciferase reporter assays were applied to confirm whether candidate exosomal miRNAs control target pathway expression. A CRC xenograft mouse model was constructed to evaluate tumor growth in vivo.
Exosomes from CRC cells contained significantly higher levels of miR-10b than did exosomes from normal colorectal epithelial cells. Moreover, exosomes containing miR-10b were transferred to fibroblasts. Bioinformatics analysis identified PIK3CA, as a potential target of miR-10b. Luciferase reporter assays confirmed that miR-10b directly inhibited PIK3CA expression. Co-culturing fibroblasts with exosomes containing miR-10b significantly suppressed PIK3CA expression and decreased PI3K/Akt/mTOR pathway activity. Finally, exosomes containing miR-10b reduced fibroblast proliferation but promoted expression of TGF-β and SM α-actin, suggesting that exosomal miR-10b may activate fibroblasts to become CAFs that express myofibroblast markers. These activated fibroblasts were able to promote CRC growth in vitro and in vivo.
CRC-derived exosomes actively promote disease progression by modulating surrounding stromal cells, which subsequently acquire features of CAFs.
癌症相关成纤维细胞（CAFs）有助于结直肠癌（CRC）细胞的增殖。然而，CAFs在肿瘤微环境中发育的机制仍不清楚。外泌体可能参与激活CAFs。
使用miRNA表达谱芯片，我们确定了CRC细胞中分泌性外泌体的miRNA表达谱，然后通过富集分析鉴定了与正常细胞相比具有显著差异表达的潜在miRNA。随后通过生物信息学分析评估候选miRNA的预测靶点。进行实时定量PCR、蛋白质印迹和细胞周期分析以评估候选外泌体miRNA的作用。应用荧光素酶报告基因测定法来确认候选外泌体miRNA是否控制靶途径表达。构建CRC异种移植小鼠模型以评估体内肿瘤生长。
CRC细胞来源的外泌体中miR-10b的水平明显高于正常结直肠上皮细胞来源的外泌体。此外，含有miR-10b的外泌体被转移到成纤维细胞中。生物信息学分析确定PIK3CA是miR-10b的潜在靶点。荧光素酶报告基因测定法证实miR-10b直接抑制PIK3CA表达。将成纤维细胞与含有miR-10b的外泌体共培养可显著抑制PIK3CA表达并降低PI3K/Akt/mTOR途径活性。最后，含有miR-10b的外泌体减少了成纤维细胞增殖，但促进了TGF-β和SMα-肌动蛋白的表达，这表明外泌体miR-10b可能激活成纤维细胞成为表达肌成纤维细胞标志物的CAFs。这些活化的成纤维细胞能够在体外和体内促进CRC生长。
CRC来源的外泌体通过调节周围基质细胞来积极促进疾病进展，随后这些基质细胞获得CAFs的特征。
