Department of Hepatobiliary and Pancreatic Surgery and Retroperitoneal Tumor Surgery, the Affiliated Hospital of Qingdao University, Qingdao, People's Republic of China.
Department of Anesthesiology, National Cancer Center/National Clinical Research Center for Cancer/Cancer Hospital, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing, People's Republic of China.
Kaohsiung J Med Sci. 2022 Nov;38(11):1080-1092. doi: 10.1002/kjm2.12590. Epub 2022 Oct 6.
This study was designed to explore the effects of exosomal miR-421 secreted by cancer-associated fibroblasts (CAFs) on pancreatic cancer (PC) progression and the mechanisms involved. CAFs and exosomes (exos) were isolated and identified. PC cells were treated with CAF-derived exos (CAF-exos). Western blotting and quantitative polymerase chain reaction (qPCR) were used to measure miR-421, sirtuin-3 (SIRT3), and hypoxia duciblefactors-1 alpha (HIF-1α) levels. Cell counting kit-8 (CCK-8), wound-healing, and transwell migration assays were used to measure proliferation, migration, and invasion abilities of the cells. Dual-luciferase assay and RNA immunoprecipitation (RIP) experiment analyzed the relationship between miR-421 and SIRT3. Chromatin immunoprecipitation (f)-verified H3K9Ac enrichment in the HIF-1α promoter region. In vivo tumorigenesis experiments were performed to further explore the effects of exosomal miR-421 from CAFs on PC. CAFs and exos were successfully isolated. CAF-exo-treated PC cells highly expressed miR-421 and had increased cell proliferation, migration, and invasion abilities. Knocking down miR-421 increased the expression of SIRT3. SIRT3 is a target of miR-421, and inhibiting the expression of SIRT3 reversed the negative effects of miR-421 knockdown on PC cell. Knocking down miR-421 in CAF-exo inhibited the expression of HIF-1α in PC cells. Moreover, SIRT3-mediated HIF-1α expression by regulating H3K9Ac. HIF-1α overexpression reversed the inhibiting effects of SIRT3 overexpression on PC progression and counteracted the inhibiting effects of miR-421 knockdown on glycolysis. Moreover, in vivo tumorigenesis experiments showed that knocking down miR-421 attenuated CAF-exo induced tumor growth. Exosomal miR-421 from CAFs promoted PC progression by regulating the SIRT3/H3K9Ac/HIF-1α axis. This study provided insights into the molecular mechanism of PC.
这项研究旨在探索肿瘤相关成纤维细胞（CAFs）分泌的外泌体 miR-421 对胰腺癌（PC）进展的影响及其相关机制。分离并鉴定 CAFs 和外泌体（exos）。用 CAF 来源的外泌体（CAF-exos）处理 PC 细胞。Western blot 和定量聚合酶链反应（qPCR）用于测量 miR-421、沉默调节蛋白 3（SIRT3）和缺氧诱导因子-1α（HIF-1α）的水平。细胞计数试剂盒-8（CCK-8）、划痕愈合和 Transwell 迁移实验用于测量细胞的增殖、迁移和侵袭能力。双荧光素酶报告基因实验和 RNA 免疫沉淀（RIP）实验分析了 miR-421 和 SIRT3 之间的关系。染色质免疫沉淀（f）实验验证了 HIF-1α启动子区域 H3K9Ac 的富集。进行体内肿瘤发生实验进一步探讨 CAFs 来源的外泌体 miR-421 对 PC 的影响。成功分离 CAFs 和 exos。CAF-exo 处理的 PC 细胞高表达 miR-421，细胞增殖、迁移和侵袭能力增强。敲低 miR-421 增加了 SIRT3 的表达。SIRT3 是 miR-421 的靶标，抑制 SIRT3 的表达逆转了 miR-421 敲低对 PC 细胞的负向作用。在 CAF-exo 中敲低 miR-421 抑制了 PC 细胞中 HIF-1α 的表达。此外，SIRT3 通过调节 H3K9Ac 介导 HIF-1α 的表达。HIF-1α 的过表达逆转了 SIRT3 过表达对 PC 进展的抑制作用，并抵消了 miR-421 敲低对糖酵解的抑制作用。此外，体内肿瘤发生实验表明，敲低 miR-421 可减弱 CAF-exo 诱导的肿瘤生长。CAFs 来源的外泌体 miR-421 通过调节 SIRT3/H3K9Ac/HIF-1α 轴促进 PC 进展。本研究为 PC 的分子机制提供了新的见解。
