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细胞周期蛋白 D1 基因沉默对肝癌细胞增殖、细胞周期和凋亡的影响。

Effects of cyclin D1 gene silencing on cell proliferation, cell cycle, and apoptosis of hepatocellular carcinoma cells.

机构信息

Hepatic Surgery Center, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, P.R. China.

Department of Clinical Immunology, School of Medical Laboratory, Tianjin Medical University, Tianjin, P.R. China.

出版信息

J Cell Biochem. 2018 Feb;119(2):2368-2380. doi: 10.1002/jcb.26400. Epub 2017 Oct 27.

Abstract

This study aims to investigate the effects of Cyclin D1 silencing on cell cycle, cell proliferation, and apoptosis of hepatocellular carcinoma cells (HCC). Cells were divided into the blank group, negative control group (HCC cells transfected with control shRNA), Cyclin D1 shRNA group (HCC cells transfected with Cyclin D1 shRNA), and the normal group (human normal liver L-02 cells). Expressions of Cyclin D1, Caspase-3, Bcl-2, and C-myc were detected by RT-qPCR and Western blotting. Cell proliferation was detected by Cell Counting Kit-8. Cell cycle and apoptosis were detected by flow cytometry. Tumor xenograft in nude mice was performed to detect in vivo tumorigenesis. HCC tissues and HCC cells exhibited elevated expression levels of Cyclin D1. Cyclin D1 expression levels was found to be correlated with tumor size and tumor staging. Compared with the normal group, the blank group showed enhanced cell proliferation, a reduction in the amount of cells in G0/G1 phase, increased number cells in S and G2/M phase, reduced apoptosis, elevated expressions of Cyclin D1, Bcl-2, and C-myc, decreased Caspase-3 activity and significant tumorigenicity. In comparison with the blank group, the Cyclin D1 shRNA group revealed weakened cell proliferation, reduced cells in S and G2/M phase, increased cells in G0/G1 phase, increased Annexin V positive cell ratio, decreased expression of Cyclin D1, Bcl-2, and C-myc, elevated Caspase-3 activity and inhibited tumorigenicity. In conclusion, Cyclin D1 gene silencing suppresses cell proliferation and inhibits cell apoptosis, which may be a new target approach in the treatment and management for HCC.

摘要

本研究旨在探讨沉默 Cyclin D1 对肝癌细胞(HCC)细胞周期、细胞增殖和凋亡的影响。将细胞分为空白组、阴性对照组(转染对照 shRNA 的 HCC 细胞)、Cyclin D1 shRNA 组(转染 Cyclin D1 shRNA 的 HCC 细胞)和正常组(人正常肝 L-02 细胞)。通过 RT-qPCR 和 Western blot 检测 Cyclin D1、Caspase-3、Bcl-2 和 C-myc 的表达。通过细胞计数试剂盒-8 检测细胞增殖。通过流式细胞术检测细胞周期和凋亡。通过裸鼠肿瘤异种移植检测体内肿瘤发生情况。HCC 组织和 HCC 细胞中 Cyclin D1 的表达水平升高。Cyclin D1 的表达水平与肿瘤大小和肿瘤分期有关。与正常组相比,空白组细胞增殖增强,G0/G1 期细胞减少,S 和 G2/M 期细胞增多,凋亡减少,Cyclin D1、Bcl-2 和 C-myc 表达升高,Caspase-3 活性降低,肿瘤生成能力增强。与空白组相比,Cyclin D1 shRNA 组细胞增殖减弱,S 和 G2/M 期细胞减少,G0/G1 期细胞增多,Annexin V 阳性细胞比例增加,Cyclin D1、Bcl-2 和 C-myc 表达降低,Caspase-3 活性升高,肿瘤生成能力受到抑制。综上所述,沉默 Cyclin D1 基因抑制细胞增殖并抑制细胞凋亡,可能成为 HCC 治疗和管理的新靶点。

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