[DADLE通过激活PKC途径抑制人肝癌HepG2细胞增殖并提高对顺二氯二氨铂的敏感性]

[DADLE suppresses the proliferation of human liver cancer HepG2 cells by activation of PKC pathway and elevates the sensitivity to cis-diammine dichloridoplatium].

作者信息

Tang Bo, DU Jian, Gao Zhen-ming, Liang Rui, Sun De-guang, Jin Xue-li, Wang Li-ming

机构信息

Department of General Surgery, the Second Affiliated Hospital of Dalian Medical University, China.

出版信息

Zhonghua Zhong Liu Za Zhi. 2012 Jun;34(6):425-9.

PMID:22967443

Abstract

OBJECTIVE

To investigate the effect of DADLE, a δ-opioid receptor agonist, on the proliferation of human liver cancer HepG2 cells and explore the mechanism involving PKC pathway.

METHODS

HepG2 cells were treated with DADLE at different doses (0.01, 0.1, 1.0 and 10 µmol/L). Cell viability was determined using methyl thiazolyl terazolium (MTT) assay. The expression of PKC mRNA and p-PKC protein were examined by RT-PCR and Western blot assay. After treated separately with DADLE plusing NAL or PMA, the cell cycle of HepG2 cells was analyzed by flow cytometer. MTT was used to detect their proliferation capacity and Western blot was used to examine the p-PKC expression. The growth inhibitory rate of HepG2 cells treated with DADLE and cis-diammine dichloridoplatinum (CDDP) was analyzed.

RESULTS

DADLE at different concentrations showed an inhibitory effect on the proliferation of HepG2 cells though inhibiting the expression of PKC mRNA and p-PKC protein. The results of flow cytometry showed that compared with the control group, the percentage of S + G(2)/M cells in DADLE-treated group was lowered by 3.94% (P < 0.01). Meanwhile, after treated with NAL and PMA, the percentage was elevated by 3.22% and 3.63%, respectively (P < 0.01). The MTT and Western blot assays showed that compared with the control group, the values of A570 and p-PKC protein levels in the HepG2 cells of DADLE-treated group were significantly decreased (P < 0.01). After treatment with NAL and PMA, the values of A570 and p-PKC protein levels were elevated significantly (P < 0.01). The growth inhibitory rate of DADLE + CDDP group was 79.9%, significantly lower than 25.2% and 43.2% of the DADLE and CDDP groups, respectively.

CONCLUSIONS

Activation of δ-opioid receptor by DADLE inhibits the apoptosis of human liver cancer HepG2 cells. The underlying mechanism may be correlated with PKC pathway. DADLE can enhance the chemosensitivity of HepG2 cells to CDDP.

摘要

目的

研究δ-阿片受体激动剂DADLE对人肝癌HepG2细胞增殖的影响，并探讨其涉及PKC途径的机制。

方法

用不同剂量（0.01、0.1、1.0和10 μmol/L）的DADLE处理HepG2细胞。采用甲基噻唑基四唑溴盐（MTT）法测定细胞活力。通过RT-PCR和蛋白质免疫印迹法检测PKC mRNA和p-PKC蛋白的表达。分别用DADLE加NAL或PMA处理后，用流式细胞仪分析HepG2细胞的细胞周期。用MTT检测其增殖能力，用蛋白质免疫印迹法检测p-PKC表达。分析DADLE和顺二氯二氨铂（CDDP）处理的HepG2细胞的生长抑制率。

结果

不同浓度的DADLE通过抑制PKC mRNA和p-PKC蛋白的表达，对HepG2细胞的增殖有抑制作用。流式细胞术结果显示，与对照组相比，DADLE处理组S+G(2)/M期细胞百分比降低了3.94%（P<0.01）。同时，用NAL和PMA处理后，该百分比分别升高了3.22%和3.63%（P<0.01）。MTT和蛋白质免疫印迹分析显示，与对照组相比，DADLE处理组HepG2细胞的A570值和p-PKC蛋白水平显著降低（P<0.01）。用NAL和PMA处理后，A570值和p-PKC蛋白水平显著升高（P<0.01）。DADLE+CDDP组的生长抑制率为79.9%，显著低于DADLE组和CDDP组的25.2%和43.2%。