南方医科大学学报 ›› 2022, Vol. 42 ›› Issue (9): 1351-1358.doi: 10.12122/j.issn.1673-4254.2022.09.11

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非小细胞肺癌细胞外泌体源性FZD10促进体外血管生成

吴小凤,詹日明,程大钊,陈 黎,王天雨,唐旭东   

  1. 广东医科大学生物化学与分子生物学研究所,抗肿瘤活性物质研发协同创新中心,广东 湛江 524023;广东医科大学附属医院,广东 湛江 524001
  • 出版日期:2022-09-20 发布日期:2022-09-28

Exosomal FZD10 derived from non-small cell lung cancer cells promotes angiogenesis of human umbilical venous endothelial cells in vitro

WU Xiaofeng, ZHAN Riming, CHENG Dazhao, CHEN Li, WANG Tianyu, TANG Xudong   

  1. Institute of Biochemistry and Molecular Biology, Collaborative Innovation Center for Antitumor Active Substance Research and Development, Guangdong Medical University, Zhanjiang 524023, China; Affiliated Hospital of Guangdong Medical University, Zhanjiang 524001, China
  • Online:2022-09-20 Published:2022-09-28

摘要: 目的 探讨非小细胞肺癌(NSCLC)细胞外泌体源性FZD10在血管生成中的作用及其机制。方法 采用超速离心法分离外泌体,并利用Western blot和RT-qPCR技术分析NSCLC细胞(95D和H1299)、正常人支气管上皮细胞(BEAS-2B)及其外泌体中FZD10的表达;通过转染FZD10-siRNA敲低FZD10表达,用FZD10未敲低和敲低的NSCLC细胞外泌体分别处理HUVEC细胞,利用体外血管生成实验观察其成管能力,采用ELISA和RT-qPCR技术分析血管生成相关因子VEGFA和Ang-1的表达;进一步利用Western blot分析外泌体源性FZD10对信号通路PI3K、Erk1/2和YAP/TAZ激活的影响。结果 同BEAS-2B细胞及其外泌体相比较,FZD10在95D和H1299细胞及其外泌体中高表达(P<0.01);95D和H1299细胞来源的外泌体可促进HUVEC的微管形成及VEGFA、Ang-1的蛋白分泌、mRNA的表达(P<0.01),但在95D和H1299细胞敲低FZD10后这些效果受到抑制。FZD10的敲低可抑制PI3K、Erk1/2信号通路的激活,但对YAP/TAZ信号通路的影响不显著。结论 NSCLC细胞外泌体源性FZD10可促进体外血管生成,其机制可能与PI3K、Erk1/2信号通路的激活有关。

关键词: 外泌体;FZD10;血管生成;非小细胞肺癌

Abstract: Objective To investigate the effect of exosomal FZD10 derived from non-small cell lung cancer (NSCLC) cells on angiogenesis of human umbilical venous endothelial cells (HUVECs) and explore the possible mechanism. Methods We analyzed the expression of FZD10 in two NSCLC cell lines (95D and H1299 cells), normal human bronchial epithelial cells (BEAS-2B cells) and their exosomes isolated by ultracentrifugation. Cultured HUVECs were treated with the exosomes derived from NSCLC cells or NSCLC cells transfected with FZD10-siRNA, and the changes in tube formation ability of the cells were analyzed using an in vitro angiogenesis assay. ELISA was performed to determine the concentration of VEGFA and Ang-1 in the conditioned media of HUVECs, and RT-qPCR was used to analyze the mRNA levels of VEGFA and Ang-1 in the HUVECs. The effects of exosomal FZD10 on the activation of PI3K, Erk1/2 and YAP/TAZ signaling pathways were evaluated using Western blotting. Results Compared with BEAS-2B cells and their exosomes, 95D and H1299 cells and their exosomes all expressed high levels of FZD10 (P<0.01). The exosomes derived from 95D and H1299 cells significantly enhanced tube formation ability and increased the expressions of VEGFA and Ang-1 protein and mRNA in HUVECs (P<0.01), but FZD10 knockdown in 95D and H1299 cells obviously inhibited these effects of the exosomes. Exosomal FZD10 knockdown suppressed the activation of PI3K and Erk1/2 signaling pathways, but had no obvious effect on the activation of YAP/TAZ signaling pathway. Conclusion Exosomal FZD10 derived from NSCLC cells promotes HUVEC angiogenesis in vitro, the mechanism of which may involve the activation of PI3K and Erk1/2 signaling pathways.

Key words: exosomes; FZD10; angiogenesis; non-small cell lung cancer