目的：探讨miR-124通过调节Jagged1（JAG1）/Notch信号通路对肾细胞癌（RCC）细胞增殖、凋亡、迁移和侵袭的影响。方法：收集2018年6月至2021年10月在武汉市第三医院治疗的38例RCC患者的RCC组织和癌旁组织标本，并体外培养RCC细胞（Caki-2、A498、ACHN、786-O、OS-RC-2）和人正常肾细胞（293T），采用免疫组织化学法、qPCR和WB法检测miR-124和JAG1蛋白在RCC组织和细胞中的表达水平。选择miR-124表达与293T细胞差异最大的OS-RC-2细胞进行转染，按转染物不同分为Control组、NC mimic组、miR-124 mimic组、miR-124 mimic+pcDNA组和miR-124 mimic+pc-JAG1组。采用双荧光素酶报告基因实验验证miR-124与JAG1的关系；qPCR法检测miR-124、JAG1 mRNA表达；免疫组化法分析JAG1蛋白表达；WB法检测JAG1、凋亡相关蛋白（cleaved caspase-3、BAX 和 Bcl2）和 Notch 信号通路相关蛋白（NICD、HES1 和 HES5）的表达；MTT 法检测OS-RC-2细胞增殖；Transwell 检测 OS-RC-2 细胞迁移和侵袭；流式细胞术检测OS-RC-2细胞凋亡。结果：与癌旁组织比较，RCC组织中miR-124表达降低，JAG1 mRNA和蛋白表达均升高（均 P<0.01）；与 293T 细胞比较，Caki-2、A498、ACHN、786-O、OS-RC-2细胞中miR-124水平降低，JAG1 mRNA和蛋白表达均升高（均P<0.05）；miR-124直接负调控JAG1。与Control组和NC mimic组比较，miR-124 mimic组miR-124表达水平、细胞凋亡率以及cleaved caspase-3和BAX蛋白表达均升高，JAG1 mRNA和蛋白表达均降低，细胞活力（24、48、72 h）下降，迁移、侵袭细胞数减少，Bcl2及NICD、HES1、HES5蛋白表达降低（均P<0.05）；与miR-124 mimic+pcDNA 组和 miR-124 mimic 组相比，miR-124 mimic+pc-JAG1 组 miR-124 表达水平、细胞凋亡率以及 cleaved caspase-3和BAX蛋白表达均降低，JAG1 mRNA和蛋白表达均升高，细胞活力（24、48、72 h）增加，迁移、侵袭细胞数增多，Bcl2及NICD、HES1、HES5蛋白表达均增加（均P<0.05）。结论：miR-124通过下调JAG1抑制Notch信号通路，降低RCC OS-RC-2细胞的增殖、迁移和侵袭能力，促进细胞凋亡。

Objective: To investigate the effects of miR-124 on the proliferation, apoptosis, migration and invasion of renal cell carcinoma (RCC) cells by regulating the Jagged1 (JAG1)/Notch signaling pathway. Methods: The RCC tissues and paracancerous tissues of 38 RCC patients treated in Wuhan Third Hospital from June 2018 to October 2021 were collected, and RCC cells (Caki-2,A498, ACHN, 786-O, OS-RC-2) and human normal kidney cells (293T) were cultured in vitro. The expression levels of miR-124 and JAG1 proteins in RCC tissues and cells were detected by immunohistochemistry, qPCR and WB assay. OS-RC-2 cells, which had the greatest difference in expression of miR-124 with that of 293T cells, were selected for transfection and divided into control group, NC mimic group, miR-124 mimic group, miR-124 mimic+pcDNA group and miR-124 mimic+pc-JAG1 group according to the difference in transfectants. The relationship between miR-124 and JAG1 was verified by dual-luciferase reporter gene experiments; the expressions of miR-124 and JAG1 mRNA were detected by qPCR; the protein expression of JAG1 was analyzed by immunohistochemistry method; WB assay was performed to measure the expressions of JAG1, apoptosis-related proteins (cleaved caspase-3, BAX, Bcl2) and Notch signaling pathway-related proteins (NICD, HES1 and HES5); MTT method was performed to measure the proliferation of OS-RC-2 cells; Transwell assay was performed to measure the migration and invasion of OS-RC-2 cells;and flow cytometry was performed to measure the apoptosis of OS-RC-2 cells. Results:Compared with paracancerous tissues, the expression of miR-124 in RCC tissues decreased, and the expressions of JAG1 mRNA and protein increased (all P<0.01); compared with 293T cells, the levels of miR-124 decreased while the expressions of JAG1 mRNA and protein increased in Caki-2, A498, ACHN,786-O and OS-RC-2 cells (all P<0.05); miR-124 directly negatively regulated JAG1. Compared with the Control group and the NC mimic group, the expression level of miR-124, the apoptosis rate and the expressions of cleaved caspase-3 and BAX proteins in the miR-124 mimic group increased; the expressions of JAG1 mRNA and protein decreased; the cell viability (24, 48, 72 h) decreased; the numbers of migrating and invasive cells decreased; the expressions of Bcl2 and NICD, HES1, HES5 proteins decreased (all P<0.05).Compared with the miR-124 mimic+pcDNA group and the miR-124 mimic group, the expression level of miR-124, the apoptosis rate and the expressions of cleaved caspase-3 and BAX proteins in the miR-124 mimic+pc-JAG1 group decreased; the expressions of JAG1 mRNA and protein increased; the cell viability (24, 48, 72 h) increased; the numbers of migrating and invasive cells increased and the expressions of Bcl2 and NICD, HES1, HES5 proteins increased (all P<0.05). Conclusion: miR-124 reduced the proliferation, migrationand invasion abilities of RCC OS-RC-2 cells and promoted apoptosis by downregulating JAG1 and inhibiting the Notch signaling pathway.

武汉市医学科研项目资助（No. WX21B04）