CDX2对SW480和HCT-116细胞增殖、凋亡、自噬及COX-2蛋白表达的影响

Effects of CDX2 on cell growth,apoptosis and autophagy of SW480 and HCT-116 cells and on COX-2 expression in vitro

目的:探讨CDX2对人结肠癌细胞株SW480和HCT-116细胞增殖、凋亡、自噬和对COX-2蛋白表达的影响以及作用机制.方法:应用MTT法检测细胞增殖抑制率;流式细胞仪(FCM)分析其细胞周期和凋亡;光镜下HE和PAS染色观察CDX2处理48 h后细胞形态学改变;透射电镜(TEM)观察细胞凋亡和自噬的超微结构变化;免疫组化(IHC)检测CDX2处理后细胞中COX-2蛋白的表达.结果:①在5～40mg/L的剂量范围内,CDX2可抑制癌细胞的生长,其抑制作用呈时间和剂量依赖性;②FCM结果表明不同剂量(2.5,5,10,20,40 mg/L)的CDX2作用48 h后,SW480和HCT-116细胞周期发生改变,细胞G0/G1期相对延长,S期和G2期缩短,凋亡率增高,与对照组相比具有统计学意义(P<0.05);③光镜发现HE和PAS染色的细胞爬片上CDX2能够诱导肿瘤细胞向成熟方向分化,特别是对分化程度低的HCT-116细胞株.TEM检测到细胞凋亡和自噬的变化,两种形态学改变可交叉重叠出现;④IHC显示COX-2在SW480和HCT-116细胞中均有表达,经10 mg/L CDX2作用48 h,细胞内COX-2蛋白表达率较空白对照组下降,具有统计学意义(P<0.05).结论:CDX2能够降低细胞增殖率,改变其生长周期,诱导细胞分化成熟.通过下调COX-2的表达,CDX2抑制SW480和HCT-116细胞增殖并促进其凋亡,诱导凋亡和/或自噬可能是其抗肿瘤的主要机制之一.

AIM： To investigate the effect and mechanism of CDX2 on the growth, apoptosis and autophagy of human colon cancer cell lines SW480 and HCT-116 and on the expression of COX-2 proteins in vitro. METHODS： Cell cycle analysis and apoptosis rates were evaluated by flow cytometery （FCM） and cell proliferative inhibition rates were measured by MTT assay. After treatment of CDX2 for 48 h, the morphological features of cells including apoptosis and autophagy were observed using transmission electron microscopy （TEM）. The expressions of COX-2 proteins were detected by immunocytochemistry. RESULTS： Within the dose range from 5 to 40 mg/L, CDX2 inhibited the proliferation of SW480 and HCT-116 cells in a timeand dosedependent manner. Cell cycle redistributed after the cells were treated with CDX2 at different doses （2.5, 5, 10, 20 and 40 mg/L） for 48 h and the rate of apoptosis increased. Phases of G0/G1 were prolonged and S and G2 were significantly shortened compared with those in control group （ P 〈 0.05 ）. Light microscopy observations of anchorage-dependent cells grown on cover slips and stained with H ＆ E and PAS showed that CDX2 induced some cells into maturation, especially HCT-116 cells which were low differentiated cells. Transmission electron microscopy revealed apoptosis and autophagy, with the two co-existing in a single cell. IHC showed that both SW480 and HCT-116 cells expressed COX-2 proteins. After the cells were treated with CDX2 for 48 h, the expression rates of COX-2 were significantly attenuated compared with that in control group （P 〈 0. 05 ）. CONCLUSION： CDX2 inhibits cell growth, alters cell cycle and motivates cells to be well-differentiated by down-regulating the expression of COX-2 in SW480 and HCT-116 cell lines and thus inducing apoptosis and/or autophagy. Apoptosis and/or autophagy may be the main mechanism by which CDX2 serves as a tumorsuppressor.