尼美舒利诱导人鼻咽癌CNE2细胞凋亡作用

Study on Nimesulide-induced Apoptosis of Human Nasopharyngeal Carcinoma Cells

目的观察尼美舒利(NIM)诱导人鼻咽癌CNE2细胞凋亡作用。方法 0,0.05,0.1,0.2,0.4,0.8 mmol·L-1NIM作用于体外培养的CNE2细胞株,光学显微镜观察细胞形态和数量变化,透射电子显微镜观察细胞形态和结构变化,噻唑蓝(MTT)法检测细胞悬液吸光度值,流式细胞术(FCM)检测细胞周期分布和凋亡率,蛋白免疫印迹反应(Western Blot)检测细胞环氧化酶2(COX-2)蛋白表达情况。结果不同浓度NIM作用3 d后,CNE2细胞皱缩,数量减少,与药物浓度呈正相关;透射电镜下,细胞形态结构随NIM浓度增加而出现不同程度破坏和死亡征象;相同作用时间下,NIM浓度越高,吸光度值越低;FCM结果显示,NIM各剂量组均在G0/G1峰前出现一个亚二倍体峰(凋亡峰),随着NIM浓度增高,G0/G1期细胞所占比例逐渐下降,G2/M期细胞比例升高,凋亡率逐渐增高;Western Blot结果显示,随着NIM浓度增加,COX-2蛋白表达量逐渐减少。上述结果均与NIM浓度相关,经统计学处理,与对照组和相邻前一浓度组比较,均差异有统计学意义(P<0.05)。结论 NIM能诱导人鼻咽癌CNE2细胞凋亡,且呈剂量依赖性,该作用与抑制COX-2有关。

Objective To observe apoptosis of nasopharyngeal carcinoma CNE2 cells induced by selective cyclooxygenase 2（COX-2） inhibitor nimesulide（NIM）.Methods The CNE2 cells cultured in vitro were treated with gradient concentrations of NIM（0,0.05,0.1,0.2,0.4 and 0.8 mmol·L-1）.Light microscope was used to observe morphology number of the cells.Transmission electron microscope was applied to study changes of cell morphology and structure.Absorbance and viability of the cells was detected by MTT assay.Cell cycle distribution and apoptosis was detected by flow cytometry（FCM）.COX-2 expression was analyzed by Western blot.Results After the CNE2 cells were treated with NIM for 3 days,cell shrinkage and reduction was observed under the light microscope,and morphological changes,structural damage and death of the cells appeared under the transmission electron microscope.MTT results showed that the higher the concentration of NIM was,the lower the absorbance of the cell suspension was detected under the same reaction time.FCM results showed that a hypodiploid peak（apoptotic peak） appeared before G0/G1 peak in all dose groups of NIM.With gradual increasing of the concentration of NIM,the cells in G0/G1 phase decreased,while the cells in G2/M phase increased,and apoptotic rate increased gradually.Western blot results showed that NIM decreased the protein expression of COX-2.These results were related with the concentration of NIM.Statistical analysis showed that the difference was significant（P〈0.05） as compared with the control group and the adjacent group.Conclusion Selective COX-2 inhibitor NIM can induce apoptosis of human nasopharyngeal carcinoma CNE2 cells,which is correlated with its inhibition on COX-2 in a dose-dependent manner.