6- 姜辣素对黑色素瘤细胞增殖和凋亡的影响及机制研究

Inhibitory effect of 6-gingerol on proliferation and apoptosis of melanoma cells via regulating endoplasmic reticulum stress

摘要:

目的??探究 6- 姜辣素对黑色素瘤细胞增殖和凋亡的影响及其分子机制。方法??使用不同浓度 6- 姜 辣素处理黑色素瘤细胞，采用 MTT 法检测黑色素瘤增殖能力的变化 ； 流式细胞仪检测 6- 姜辣素对黑色素瘤细 胞凋亡的影响 ； 应用 Western?blotting 检测黑色素瘤细胞半胱天冬氨酸蛋白酶 3（Caspase-3）/ 活化型半胱天冬 氨酸蛋白酶 3（Cleaved-Caspase-3） 、多聚腺苷二磷酸核糖聚合酶（PARP）/ 活化型多聚腺苷二磷酸核糖聚合 酶（Cleaved-PARP） 、免疫球蛋白结合蛋白（BIP） 、肌醇激酶 1（IRE1）和 c-Jun 氨基末端激酶（JNK）蛋白表 达水平变化 ； 敲减 IRE1 阻断内质网应激信号通路，检测 6- 姜辣素对黑色素瘤细胞增殖和凋亡的影响。结果? 6- 姜辣素抑制黑色素瘤细胞增殖呈浓度依赖性，对 M14 和 A375 细胞的半数抑制浓度（IC 50 ）分别为 46.070 和 33.152?μmol/L； 6- 姜辣素处理后， 黑色素瘤细胞 Cleaved-Caspase-3 和 Cleaved-PARP 蛋白表达上调（ P <0.05） ， 细胞凋亡率升高（ P <0.05） ； 6- 姜辣素激活内质网应激，BIP、IRE1 和 JNK 蛋白表达上调（ P <0.05） ； 6- 姜辣 素处理 IRE1 基因敲减细胞， 与对照组相比对细胞增殖抑制减弱、 凋亡蛋白Cleaved-Caspase-3表达下调 （ P <0.05） 。 结论??6- 姜辣素可通过激活内质网应激， 抑制黑色素瘤细胞增殖并促进其凋亡， 从而发挥抗肿瘤功能。

Abstract:

Objective To explore the inhibitory effect of 6-gingerol on the proliferation and apoptosis of melanoma cells and its molecular mechanism. Methods Melanoma cells were treated with different concentrations of 6-gingerol. Cellular proliferation and apoptosis rate were identified by MTT and flow cytometry, respectively. The protein levels of Caspase-3/Cleaved-Caspase-3, PARP/Cleaved-PARP, BiP, IRE1 and JNK in melanoma cell line were measured by Western blotting. Endoplasmic reticulum stress signaling pathway was blocked by knockdown of IRE1 for further mechanism study. Result The 6-gingerol inhibited the proliferation of melanoma cells in a concentration- and time-dependent manner while apoptosis rate was enhanced (P < 0.05). IC 50 of 6-gingerol was 46.070 μmol and 33.152 μmol/L for M14 and A375 cell line, respectively. Treatment with 6-gingerol induced increased expression of Cleaved-Caspase-3 and cleaved-PARP (P < 0.05). Endoplasmic reticulum stress was activated by 6-gingerol with increase of BiP, IRE1 and JNK. IRE1 knockdown reversed 6-gingerol-induced upregulation of proliferation, cleaved-Caspase-3 (P < 0.05). Conclusion The 6-gingerol inhibits proliferation of melanoma cell by activating the endoplasmic reticulum stress.