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作 者:王炳淑 梁荣珍 戢楠楠 WANG Bingshu;LIANG Rongzhen;JI Nannan(Department of Pathology,the Second Affiliated Hospital of Hainan Medical College,Haikou 570311,Hainan,China;Department of Geriatrics,the Second Affiliated Hospital of Hainan Medical College,Haikou 570311,Hainan,China;Department of Radiotherapy,the Second Affiliated Hospital of Hainan Medical College,Haikou 570311,Hainan,China)
机构地区:[1]海南医学院第二附属医院病理科,海南海口570311 [2]海南医学院第二附属医院老年病科,海南海口570311 [3]海南医学院第二附属医院放疗科,海南海口570311
出 处:《中国肿瘤生物治疗杂志》2020年第2期135-141,共7页Chinese Journal of Cancer Biotherapy
基 金:海南省卫生计生委科研资助项目(No.19A200161)。
摘 要:目的:探讨珠子参多糖(panax japlcus var polysaccharide,PJPS)对胃癌MKN45细胞增殖和凋亡的影响及其调控机制。方法:选用人胃癌细胞系HGC27、MGC803、MKN45和胃黏膜上皮细胞株GES-1,分别将let-7a mimics、let-7a inhibitor转染进MKN45细胞;以100μg/ml PJPS处理胃癌细胞系并挑选后续实验细胞株为MKN45细胞;分别添加0、10、50、100、120μg/ml PJPS处理转染后各组MKN45细胞,用CCK-8法、流式细胞术分别检测MKN45细胞增殖活力和细胞凋亡率,用Western blotting检测MKN45细胞中细胞周期依赖性激酶6(cycle dependent kinase 6,CDK6)和凋亡相关蛋白的表达,用qPCR法检测调控胃癌细胞增殖的miRNAs表达水平。用双荧光素酶报告基因实验验证let-7a和CDK6的靶向关系。结果:与其他胃癌细胞比较,100μg/ml PJPS可显著抑制MKN45细胞的增殖活力(P<0.01);同时,100μg/ml PJPS处理后,可显著上调MKN45细胞中let-7a的表达(P<0.01)。双荧光素酶报告基因实验证实CDK6是let-7a的靶基因。进一步实验显示,PJPS通过上调let-7a靶向抑制CDK6的表达,从而抑制MKN45细胞的增殖并诱导其凋亡(均P<0.01)。结论:PJPS通过调控let-7a/CDK6分子轴抑制胃癌MKN45细胞的增殖并诱导其凋亡。Objective: To investigate the effect of panax japlcus var polysaccharide(PJPS) on the proliferation and apoptosis of gastric cancer MKN45 cells and its regulatory mechanism. Methods: Human gastric cancer cell lines(HGC27, MGC803, MKN45) and gastric mucosal epithelial cell line GES-1 were selected for this study. Let-7 a mimics and let-7 a inhibitor were transfected into MKN45 cells;Gastric cancer cell lines were treated with 100 μg/ml PJPS and MKN45 was selected as the subsequent experimental cell line. MKN45 cells were cultured with 0, 10, 50, 100 and 120 μg/ml PJPS, respectively. The proliferation and apoptosis rate of MKN45 cells were detected by CCK-8 and flow cytometry, respectively. Expressions of cell cycle dependent kinase 6(CDK6) and apoptosis-related proteins in MKN45 cells were detected by Western blotting, and the expression level of miRNAs regulating the proliferation of gastric cancer cells was detected by Real-time quantitative PCR(qPCR). The Dual luciferase reporter gene assay was used to validate the targeting relationship between let-7 a and CDK6. Results: Compared with other gastric cancer cells, 100 μg/ml PJPS significantly inhibited the proliferation of MKN45 cells(P<0.01). At the same time, 100 μg/ml PJPS significantly up-regulated the expression of let-7 a in MKN45 cells(P<0.01). The Dual luciferase reporter gene assay confirmed that CDK6 was the target gene of let-7 a. Furthermore, PJPS inhibited the expression of CDK6 by up-regulating let-7 a, thereby inhibiting the proliferation and inducing apoptosis of MKN45 cells(all P<0.01). Conclusion: PJPS inhibits proliferation and induces apoptosis of gastric cancer MKN45 cells by regulating the let-7 a/CDK6 axis.
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