乌贼墨多肽诱导人前列腺癌DU-145细胞凋亡的机制研究

Mechanism of Sepia Ink Polypeptide-induced Apoptosis in DU-145 Prostate Cancer Cells

摘要:

本文探讨了乌贼墨多肽（SHP）诱导DU-145细胞凋亡机制。采用CCK-8法检测SHP对DU-145细胞增殖的影响；采用HE染色和AO/EB荧光染色观察DU-145细胞的形态学的变化；采用流式细胞术检测细胞早期凋亡率；并通过Western Blotting检测细胞中p53、Bcl-2、Bax、Caspase-3、VEGF的蛋白表达变化。结果表明，SHP对DU-145细胞的增殖具有明显的抑制作用且呈现剂量和时间依赖性；SHP作用后的DU-145细胞出现凋亡的形态学特征；流式细胞术结果显示，随着SHP浓度和作用时间的增加，DU-145细胞的早期凋亡率从12.25%增加到34.20%；Western Blotting结果显示，当SHP作用24 h后，VEGF、Bcl-2蛋白表达量降低，p53、Bax、Caspase-3蛋白表达量增加。综上可知，SHP能够诱导DU-145细胞凋亡，其机制有可能是通过激活抑癌基因p53，下调Bcl-2/Bax比例；下调VEGF，激活凋亡蛋白酶Caspase-3，诱发凋亡级联反应来实现的。

Abstract:

In this study, the mechanism of apoptosis induced by sepia ink polypeptide (SHP) in DU-145 prostate cancer cells was explored. The effect of SHP on the proliferation of DU-145 cells was examined by the Cell Counting Kit-8 (CCK-8) assay. Typical morphological changes in DU-145 cells were observed with hematoxylin and eosin (HE) and acridine orange/ethidium bromide (AO/EB) staining. The early-stage apoptosis rate was measure using flow cytometry (FCM), and the changes in the expression of apoptosis-related genes (p53, Bcl-2, Bax, Caspase-3, and VEGF) were evaluated via western blotting. The results showed that SHP significantly inhibited the proliferation of DU-145 cells in a time-and dose-dependent manner. DU-145 cells developed morphological features of apoptosis after treated with SHP.FCM studies revealed that the early-stage apoptosis rate of DU-145 cells increased from 12.25% to 34.20% with increasing SHP concentration and duration of treatment. Western blotting results showed that after 24 h treatment with SHP, the expression of anti-apoptotic proteins Bcl-2 and VEGF decreased, while the expression of p53, Bax, and Caspase-3 increased. Collectively, these results suggest that SHP induced apoptosis in DU-145 cells. The mechanism might involve the decrease in Bcl-2/Bax expression ratio by activation of the tumor suppressor gene p53. Moreover, VEGF expression was down regulated, and apoptotic protease Caspase-3 was activated, thus triggering the apoptosis cascade reaction.