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中国临床研究英文版:2022,35(11):1516-1522
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circBIRC6靶向miR-449b-5p调控胃癌SUN-1细胞的增殖、细胞周期及凋亡
(武汉科技大学附属孝感医院消化内科,湖北 孝感 432000)
circBIRC6 in regulation of the proliferation, cell cycle and apoptosis of gastric cancer SUN-1 cells by targeting miR-449b-5p
(Department of Gastroenterology, Xiaogan Hospital Affiliated to Wuhan University of Science and Technology, Xiaogan, Hubei 432000, China)
摘要
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Received:March 14, 2022   Published Online:November 20, 2022
中文摘要: 目的 探讨circBIRC6对胃癌细胞增殖、细胞周期及凋亡的影响及可能的作用机制。方法收集2020年1月至5月武汉科技大学附属孝感医院收治的53例胃癌患者的癌组织及其癌旁组织标本,RT-qPCR法检测胃癌组织、癌旁组织与人胃黏膜细胞GES1、人胃癌细胞SNU-1、AGS、HS-746T中circBIRC6、miR-449b-5p的表达量;以SNU-1细胞为研究对象,随机分组为si-NC组、si-circBIRC6组、miR-NC组、miR-449b-5p组、si-circBIRC6+anti-miR-NC组、si-circBIRC6+anti-miR-449b-5p组;CCK-8法、流式细胞仪分别检测细胞增殖、细胞周期及凋亡率;双荧光素酶报告实验检测miR-449b-5p过表达对野生型载体WT-circBIRC6荧光素酶活性的影响;western blot检测cleaved caspase-3蛋白表达量。结果 与癌旁组织比较,胃癌组织中circBIRC6的表达量升高(P<0.05),miR-449b-5p的表达量降低(P<0.05);与GES1细胞比较,SNU-1、AGS、HS-746T细胞中circBIRC6的表达量升高(P<0.05),miR-449b-5p的表达量降低(P<0.05)。与si-NC组比较,si-circBIRC6组细胞活力和S期细胞比例降低(P<0.05),G0/G1期细胞比例、细胞凋亡率和cleaved caspase-3蛋白水平升高(P<0.05);与miR-NC组比较,miR-449b-5p组细胞活力和S期细胞比例降低(P<0.05),G0/G1期细胞比例、细胞凋亡率和cleaved caspase-3蛋白水平升高(P<0.05)。miR-449b-5p过表达可降低WT-circBIRC6的荧光素酶活性(P<0.05)。与si-circBIRC6+anti-miR-NC组比较,si-circBIRC6+anti-miR-449b-5p组细胞活力和S期细胞比例升高(P<0.05),G0/G1期细胞比例、细胞凋亡率和cleaved caspase-3蛋白水平降低(P<0.05)。结论 干扰circBIRC6表达可通过上调miR-449b-5p表达而抑制胃癌细胞增殖、细胞周期进程及诱导细胞凋亡。
中文关键词: 胃癌  circBIRC6  miR-449b-5p  细胞增殖  凋亡  细胞周期
Abstract:Objective To explore the effects of circBIRC6 on the proliferation, cell cycle and apoptosis of gastric cancer cells and its possible mechanism. Methods The specimens of cancer tissues and para-cancerous tissue were collected in 53 patients with gastric cancer treated in Xiaogan Central Hospital Affiliated to Wuhan University of Science and Technology from January 2020 to May 2020. RT-qPCR was used to detect the expression of circBIRC6 and miR-449b-5p in gastric cancer tissues, para-cancerous tissues, human gastric mucosal epithelial cells GES1, and human gastric cancer cells lines (SNU-1, AGS and HS-746T). SNU-1 cells were taken as the research objects and were divided into si-NC group, si-circBIRC6 group, miR-NC group, miR-449b-5p group, si-circBIRC6+anti-miR-NC group and si-circBIRC6+anti-miR-449b-5p group. Proliferation, cell cycle and apoptosis of SNU-1 cells were detected by cell counting kit-8 (CCK-8) and flow cytometry assays, respectively. The dual luciferase reporter experiment was conducted to detect the effect of miR-449b-5p overexpression on luciferase activity of wild-type vector WT-circBIRC6. Western blot was used to detect the expression of cleaved caspase-3 protein. Results Compared with para-cancerous tissues, the expression of circBIRC6 in gastric cancer tissues was increased significantly(P<0.05), while the expression of miR-449b-5p was decreased (P<0.05). Compared with GES1 cells, the expressions of circBIRC6 in SNU-1, AGS, and HS-746T cells significantly increased (P<0.05), and the expression of miR-449b-5p decreased (P<0.05). Compared with those in si-NC group, the cell viability and the proportion of S-phase cells were decreased (P<0.05), while the proportion of cells in the G0/G1 phase, the rate of apoptosis and the protein level of cleaved caspase-3 were increased in si-circBIRC6 group (P<0.05). Compared with those in miR-NC group, the cell viability and the S-phase cell ratio decreased (P<0.05), and the G0/G1 phase cell ratio, cell apoptosis rate and the protein level of cleaved caspase-3 significantly increased in miR-449b-5p group (P<0.05). Overexpression of miR-449b-5p significantly reduced the luciferase activity of WT-circBIRC6 (P<0.05). Compared with those in si-circBIRC6+anti-miR-NC group, the cell viability and the S-phase cell ratio were increased (P<0.05), while G0/G1 phase cell ratio, cell apoptosis rate and protein level of cleaved caspase-3 were decreased in si-circBIRC6+anti-miR-449b-5p group (P<0.05).Conclusions Interfering with the expression of circBIRC6 could inhibit the proliferation and cell cycle progression and induce apoptosis of gastric cancer cells by up-regulating the expression of miR-449b-5p.
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