环氧合酶-2抑制剂对结肠癌细胞株的体外研究

目的 观察选择性环氧合酶-2（COX-2）抑制剂对COX-2高表达的结肠癌细胞株HT-29增殖和凋亡的影响，明确以COX2为靶点治疗结肠癌的作用途径以及与COX-2活性、表达水平的相关关系。方法 将选择性COX-2抑制剂NS-398作用于结肠癌细胞系HT29，运用MTT法检测细胞增殖状态。流式细胞仪观察NS-398对细胞凋亡的影响。进一步用逆转录聚合酶链式反应（RT-PCR）检测药物作用前后HT-29中COX-2mRNA表达。ELISA法测定前列腺素E2（PGE2）水平。Western blot检测药物作用前后细胞周期素D1、Bcl-2的表达。结果 结肠癌细胞系HT-29中COX-2 mRNA高表达，NS-398呈时间和剂量依赖性抑制HT-29细胞增殖，促进其凋亡。加入NS-398的HT-29细胞中COX-2mRNA表达水平无明显变化（P〉0．05），PGE2却显著下降（P〈0．01）。72h时空白组与NS-398（75μmol／L）处理组细胞周期素D1、Bcl-2表达水平比值分别为2．21和3．25（P〈0．01），两者表达水平随作用时间延长而下降。结论 选择性COX-2抑制剂NS-398不影响结肠癌细胞COX-2 mRNA表达水平，而与其活性相关（PGE2水平）．可能通过细胞周期素D1、Bcl-2影响结肠癌细胞系HT-29的增殖与凋亡，揭示了COX-2为靶点治疗结肠癌的分子机制。

The study of selective cyclooxygenase-2 inhibitor NS-398 on proliferation and apoptosis of human colon cancer cells

Objective To investigate the influence of NS-398, a selective cyclooxygenase(COX)-2 inhibitor, on the proliferation and apoptosis of colorectal cancer cell HT-29, and to explore its potential mechanism. Methods Cultured HT-29 cells were treated with NS-398. MTT assay and flow cytometry were used to measure the proliferation and apoptosis. RT-PCR analysis was performed to measure the level of COX-2 mRNA expression at different time points in HT-29 cells. The expression of prostaglandin (PG)E2,Cyclin D1 and Bcl-2 were measured by ELISA and Western blot, respectively.Results High expression of COX-2 mRNA was detected in colorectal cancer cell line HT-29 and no obvious change of COX-2 mRNA was found after the treatment of NS-398 . NS-398 inhibited the cells proliferation and induced apoptosis in a dose-and time-dependent manner, and resulted in a significant down-regulation of Cyclin D1, Bcl-2 and PGE2.Conclusions Our results show that NS-398 may inhibit the proliferation and induce apoptosis of colon cancer cell lines HT-29 through decreasing expression of Cyclin D1 and Bcl-2 . COX-2 activity rather than its mRNA expression was related to NS-398-mediated pathway on HT-29 cells. This may be a new interfering target of selective COX-2 inhibitor on colon cancer.