葡萄籽原花青素对人骨肉瘤143B细胞的作用及机制研究  

The Effects of Grape Seed Proanthocyanidins on Human Osteosarcoma 143B Cell Line and Its Mechanisms

作　　者：谢佳卿 赵佳丽[1] 李爱芳[1] 谷月[1] 陈露 黄茂 彭棋 曾宗跃[1] 周兰[1] 

机构地区：[1]重庆医科大学检验医学院,临床检验诊断学教育部重点实验室,重庆400016

出　　处：《中国细胞生物学学报》2017年第12期1543-1549,共7页Chinese Journal of Cell Biology

摘　　要：该文研究葡萄籽原花青素(grape seed procyanidins,GSP)对人骨肉瘤143B细胞增殖、凋亡的影响及其机制。用不同剂量GSP处理骨肉瘤143B细胞,结晶紫染色和集落形成试验分别用于检测细胞增殖和集落形成能力的变化;流式细胞术检测细胞周期与凋亡,Western blot法检测周期和凋亡相关蛋白;RT-PCR和Western blot法检测Wnt/β-catenin信号通路相关分子表达的变化;采用腺病毒Adβ-catenin感染以过表达β-catenin,探讨GSP抑制143B细胞增殖的作用是否与Wnt/β-catenin信号通路相关。结果显示,GSP在20~60μg/m L剂量范围内呈剂量和时间依赖性抑制143B细胞增殖活力和降低细胞克隆形成能力;GSP引起G0/G1周期阻滞和周期相关蛋白cyclin D1下调,还使细胞凋亡率明显增高,并伴有活化的凋亡相关蛋白cleaved PARP(poly ADP-ribose polymerase)和cleaved caspase-8的水平增高;GSP降低143B细胞中β-catenin m RNA水平及核内与总β-catenin蛋白质水平,抑制GSK3β磷酸化并上调GSK3β蛋白质水平;过表达β-catenin可部分减弱GSP对该细胞增殖活性的抑制作用。结果表明,GSP抑制骨肉瘤143B细胞的增殖和促进其凋亡,其机制涉及Wnt/β-catenin信号通路的抑制。To investigate the effect of grape seed proanthocyanidins（GSP） on cell proliferation and apoptosis, and its relevant mechanisms in human osteosarcoma cells, human osteosarcoma 143 B cell line was treated with different concentration of GSP. The cell proliferation activity was detected by crystal violet staining and colony forming assay. The cell cycle and apoptosis of 143 B cells were determined by flow cytometry. Overexpressing β-catenin by infection of adenovirus Adβ-catenin was performed to investigate whether Wnt/β-catenin signaling pathway was involved in the GSP-induced proliferation suppression in 143 B cells. RT-PCR and Western blot assay were used to detect the m RNA level and protein level of the related proteins, respectively. The result of crystal violet staining showed that 20~60 μg/m L GSP inhibited the cell proliferation activity in a dose-and time-dependent manner. The colony forming assay showed that GSP treatment decreased the number of colony. GSP induced G0/G1 phase arrest by the flow cytometry analysis and decreased the cell cycle related protein cyclin D1. The apoptosis rate and the apoptosis related proteins cleaved PARP（poly ADP-ribose polymerase） and cleaved caspase-8 were higher than their controls. At the same time, GSP increased GSK3β protein and reduced β-catenin m RNA, p-GSK3β, total β-catenin protein, nuclear β-catenin protein, and cyclin D1 protein. β-catenin overexpression partially attenuated the inhibition effect of GSP on cell proliferation. Ourstudy demostrated that GSP could inhibit the proliferation and promote the apoptosis in human osteosarcoma 143 B cells, and its mechanism of the proliferation inhibition supposed to be related to inhibition of Wnt/β-catenin signaling pathway.

关 键 词：葡萄籽原花青素 骨肉瘤细胞 增殖 凋亡 WNT/Β-CATENIN信号通路 

分 类 号：R738[医药卫生—肿瘤]