摘要
目的:探讨肝素结合分子中期因子(midkine,MK)对肝癌细胞Hep3B抵抗失巢凋亡的影响。方法:采用悬浮培养法建立人肝癌来源细胞系Hep3B失巢凋亡模型,以不同质量浓度(10、50、100 ng/ml)MK或PBS(对照组)处理失巢培养的肝癌细胞Hep3B,采用流式细胞术检测Hep3B细胞的凋亡,采用Western blotting检测凋亡相关蛋白Bcl-2和caspase-3的表达。结果:随着悬浮培养时间的延长,肝癌细胞Hep3B失巢凋亡率逐渐升高,培养72 h后悬浮培养的Hep3B细胞凋亡率显著高于贴壁培养的Hep3B细胞凋亡率[(38.76±4.23)%vs(6.76±1.43)%,P<0.01]。不同质量浓度MK处理24 h后,悬浮培养Hep3B细胞的凋亡率均明显低于对照组,且与MK的浓度呈负相关关系(r=0.951,P=0.049);同时,MK处理后Hep3B细胞内抗凋亡蛋白Bcl-2表达明显增加,而促凋亡蛋白caspase-3则明显下降。结论:MK可能通过上调Bcl-2蛋白表达和下调caspase-3蛋白的表达来提高肝癌细胞Hep3B在失巢状态下抵抗凋亡的能力。
Objective：To explore the effect of midkine （MK）, a heparin-binding cytokine, on the anoikis resistance in hepatocellular carcinoma Hep3B cells. Methods： Human hepatoma-derived cell line Hep3B was kept in suspension to induce an anoikis model and treated with different concentrations of MK （0, 10, 50 and 100 ng/ml） or PBS （control group）. The apoptosis of Hep3B cells was detected by flow cytometry and the expressions of apoptosis-related proteins Bcl-2 and caspase-3 were determined by Western blotting. Results：As incubation time was prolonged, the apoptotic rate of hepatocellular carcinoma Hep3B cells cultured in suspension was gradually increased. After incubation for 72 h, the apoptotic rate of Hep3B cells cultured in suspension was significantly higher than that in the adherent-cultured cells （／[38.76±423／]% vs ／[6.76±1.43／]%，P〈0.01）. After exposure of different concentrations of MK for 24 h, the apoptotic rate of the suspension-cultured Hep3B cells was significantly lower than that of the control group and negatively correlated with the concentration of MK （r=0.951, P=0.049）. Simultaneously, the expression of intracellular anti-apoptotic protein Bcl-2 was significantly increased, whereas the expression of pro-apoptotic protein caspase-3 was significantly decreased. Conclusion： MK confers enhanced anoikis resistance in hepatocellular carcinoma Hep3B cells cultured in suspension, which may be associated with the up-regulation of Bcl-2 protein and down-regulation of caspase-3 protein.
出处
《中国肿瘤生物治疗杂志》
CAS
CSCD
北大核心
2013年第5期565-568,共4页
Chinese Journal of Cancer Biotherapy
基金
国家自然科学基金资助项目(No.81272669
No.81272668)
国家传染病重大专项课题资助项目(No.2012ZX10002012-10)~~
