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- 2016
绿原酸对视网膜母细胞瘤细胞株HXO-RB44的生长抑制作用
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Abstract:
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。
ABSTRACT: 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 with 80μmol/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
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