摘要
目的探讨绿原酸(CHA)对体外培养人视网膜母细胞瘤(Rb)细胞株HXO-RB44的作用及机制。方法应用CCK-8体外抑制实验,筛选CHA对体外培养传6代的人Rb细胞株HXO-RB44的最佳抑癌浓度。Western blot检测人Rb细胞株HXO-RB44中抑癌基因Rb1和P16及细胞周期蛋白(Cyclin D2)和细胞周期蛋白依赖激酶(CDK 4)的表达变化;ELISA检测培养基中乳酸脱氢酶(LDH)水平。结果 CCK-8体外抑制实验结果显示,CHA对人Rb细胞株HXO-RB44的最佳抑癌浓度为80μmol/L。经80μmol/L CHA处理5d的HXO-RB44细胞株中,抑癌基因Rb1和P16均有少量表达,而在阳性对照组(HT1080细胞)和未处理的HXO-RB44细胞几乎没有表达。CHA处理5d的HXO-RB44细胞株的Cyclin D2和CDK4的表达明显低于阳性对照组(HT1080细胞)和未处理的HXO-RB44细胞(P<0.01)。CHA处理5d的HXO-RB44的培养基中LDH的含量为(226.75±42.74)U/L,明显低于正常培养的细胞培养基(425.84±31.67)U/L(P<0.01)。结论 80μmol/L CHA可能通过增加HXO-RB44抑癌基因的表达,降低细胞周期蛋白的表达及LDH水平,而抑制HXO-RB44细胞株的增殖。
Objective To explore the function and mechanism of chlorogenic acid（CHA）on human retinoblastoma cell line HXO-RB44 cultured in vitro.Methods The optic concentration of CHA which inhibited the 6th generation subcultured HXO-RB44 was determined by CCK8 inhibition experiment in vitro.The expressions of anti-oncogene Rb1 and P16,Cyclin D2 and CDK4 were determined by Western blot.The concentration of lactate dehydrogenase（LDH）was detected by enzyme-linked-immunosorbent serologic assay（ELISA）.Results The optimal concentration of CHA for inhibiting HXO-RB44 cell line was 80μmol/L determined by CCK8 inhibition experiment.The anti-oncogenes Rb1 and P16 were expressed in a low quantity after HXO-RB44 was treated with80μmol/L of CHA for 5 days.However,there was hardly Rb1 or P16 expression in the positive group（HT1080）and the normal cultured HXO-RB44（chlorogenic acid concentration of 0μmol/L）determined by Western blot methods.The expressions of Cyclin D2 and cyclin dependence kinase 4（CDK4）were lower after HXO-RB44 received 80μmol/L of CHA for 5 days than those in the HT1080 cell line and the normal culture HXO-RB44（P〈0.01）.The concentration of LDH in medium of HXO-RB44,treated with 80μmol/L of CHA for 5 days was lower than that of the normal cultured HXO-RB44[（226.75±42.74）U/L vs.（425.84±31.67）U/L,P〈0.01].Conclusion CHA of 80μmol/L may inhibit the proliferation of HXO-RB44 cell line via increasing the anti-oncogene expression and decreasing the expressions of Cyclin D2 and CDK4 as well as the concentration of LDH in medium.
出处
《西安交通大学学报（医学版）》
CAS
CSCD
北大核心
2016年第5期734-738,共5页
Journal of Xi’an Jiaotong University（Medical Sciences）
基金
辽宁省自然科学基金资助项目(No.2013022066
2013022048)
辽宁省教育厅创新团队项目(LT2012016)和一般项目(L2013335)~~
