菠萝蛋白酶对人胃肠道癌细胞系（MKN45、KATO-III、HT29-5F12 和 HT29-5M21）的细胞毒性作用。
Cytotoxic effects of bromelain in human gastrointestinal carcinoma cell lines (MKN45, KATO-III, HT29-5F12, and HT29-5M21).
机构信息
Department of Surgery, St George Hospital, University of New South Wales, Sydney, NSW, Australia.
出版信息
Onco Targets Ther. 2013 Apr 16;6:403-9. doi: 10.2147/OTT.S43072. Print 2013.
BACKGROUND
Bromelain is a pineapple stem extract with a variety of therapeutic benefits arising from interaction with a number of different biological processes. Several preclinical studies and anecdotal clinical observations have reported the anticancer properties of bromelain. In the present study, we investigated the cytotoxic effects of bromelain in four human cancer cell lines of gastrointestinal origin and the mechanisms involved.
METHODS
The gastric carcinoma cell lines (KATO-III and MKN45) and two chemoresistant subpopulations of the HT29 colon adenocarcinoma cell line (HT29-5M21 and HT29-5F12) were treated with a range of concentrations of bromelain, as well as with cisplatin as a positive control. The effect of bromelain on the growth and proliferation of cancer cells was determined using a sulforhodamine B assay after 72 hours of treatment. Expression of apoptosis-associated proteins in MKN45 cells treated with bromelain was analyzed by Western blotting.
RESULTS
Data from our sulforhodamine B assay showed that bromelain inhibited proliferation of HT29-5F12, HT29-5M21, MKN45, and KATO-III cells, with respective half maximal inhibitory concentration values of 29, 34, 94, and 142 μg/mL. Analyzing the expression of proapoptotic and antiapoptotic proteins in bromelain-treated MKN45 cells, we observed activation of the caspase system, cleavage of PARP and p53, overexpression of cytochrome C, attenuation of phospho-Akt and Bcl2, and removal of MUC1. Apart from the caspase-dependent apoptosis observed, emergence of cleaved p53 supports a direct, extranuclear apoptotic function of p53. Moreover, interrupted Akt signaling and attenuation of Bcl2 and MUC1 oncoproteins suggest impaired survival of cancer cells.
CONCLUSION
Our findings collectively indicate that bromelain exerts cytotoxic effects in a panel of human gastric and colon carcinoma cells. Our study of MKN45 cells implicated different mechanisms in bromelain-induced cell death. While promoting apoptosis with involvement of the caspase system and extranuclear p53, bromelain also appears to impair cancer cell survival by blocking the Akt pathway and attenuating Bcl2 and MUC1 oncoproteins.
背景
菠萝蛋白酶是一种从菠萝茎中提取的物质，具有多种治疗功效，源于与多种不同生物过程的相互作用。一些临床前研究和临床观察报告了菠萝蛋白酶的抗癌特性。在本研究中，我们研究了菠萝蛋白酶对四种胃肠道来源的人类癌细胞系的细胞毒性作用及其机制。
方法
用一系列浓度的菠萝蛋白酶处理胃腺癌细胞系（KATO-III 和 MKN45）和 HT29 结肠腺癌细胞系的两个化疗耐药亚群（HT29-5M21 和 HT29-5F12），并以顺铂作为阳性对照。用磺酰罗丹明 B 法在 72 小时治疗后测定菠萝蛋白酶对癌细胞生长和增殖的影响。用 Western blot 法分析菠萝蛋白酶处理的 MKN45 细胞中凋亡相关蛋白的表达。
结果
磺酰罗丹明 B 检测结果显示，菠萝蛋白酶抑制 HT29-5F12、HT29-5M21、MKN45 和 KATO-III 细胞的增殖，其半数最大抑制浓度分别为 29、34、94 和 142μg/ml。分析菠萝蛋白酶处理的 MKN45 细胞中促凋亡和抗凋亡蛋白的表达，我们观察到 caspase 系统的激活、PARP 和 p53 的切割、细胞色素 C 的过表达、磷酸化 Akt 和 Bcl2 的衰减以及 MUC1 的去除。除了观察到 caspase 依赖性凋亡外，裂解的 p53 的出现支持 p53 的直接、核外凋亡功能。此外，Akt 信号中断和 Bcl2 和 MUC1 癌蛋白的衰减提示癌细胞的存活受到损害。
结论
我们的研究结果表明，菠萝蛋白酶对一组人类胃和结肠癌细胞具有细胞毒性作用。我们对 MKN45 细胞的研究表明，在菠萝蛋白酶诱导的细胞死亡中涉及不同的机制。虽然通过 caspase 系统和核外 p53 促进凋亡，但菠萝蛋白酶似乎还通过阻断 Akt 通路和衰减 Bcl2 和 MUC1 癌蛋白来损害癌细胞的存活。
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