小鼠β-防御素2对结肠癌细胞增殖和凋亡的影响

Effects of mouse β-defensin 2 on proliferation and apoptosis of colon cancer cells

目的 探究小鼠β-防御素(mBD)2对结肠癌细胞SW480细胞增殖、凋亡作用及可能作用机制,为结肠癌的临床治疗提供新思路。方法 体外培养SW480细胞,转染mBD2重组质粒分为空白组(Control组)、阴性转染组(NC组)、mBD2转染组,CCK8法检测细胞增殖情况;平板细胞克隆实验检测细胞克隆形成数目情况;Hoechst33342染色法观察细胞形态变化;流式细胞仪检测细胞凋亡及细胞周期情况;Western印迹法检测凋亡有关蛋白Bax、活化的含半胱氨酸的天冬氨酸蛋白水解酶(caspase)3、Bcl-2蛋白表达情况。结果 随着细胞培养时间延长,mBD2转染组细胞增殖抑制率逐渐升高,在同一时间点,与Control、NC组相比,mBD2转染组细胞增殖抑制率显著升高( P <0.05)。与Control、NC组相比,mBD2转染组细胞克隆形成数目均降低,mBD2转染组细胞凋亡率显著升高( P <0.05)。与Control、NC组相比,mBD2转染组细胞G1期DNA量均显著升高( P < 0.05)。3组G2期、S期DNA量无明显变化( P >0.05)。与Control、NC组相比,mBD2转染组细胞中Bax、活化caspase3蛋白表达均显著升高,Bcl-2蛋白表达显著降低( P <0.05)。结论 mBD2可能通过使细胞阻滞于G1期,上调凋亡蛋白表达,进而抑制SW480细胞增殖,促进其凋亡,发挥抗肿瘤作用。

Objective To explore the proliferation and apoptosis effects of mouse β-defensin (mBD) 2 on the colon cancer cell SW480 cells and its possible mechanism. Methods SW480 cells were cultured in vitro. mBD2 recombinant plasmids were divided into blank group (control group), negative transfection group (NC group) and mBD2 transfection group. CCK8 assay was used to detect cell proliferation. Colony formation was detected by the panel cell cloning experiment. Morphological changes of cells were observed by Hoechst33342 staining. Cell apoptosis and cell cycle distribution were detected by flow cytometry. Western blot was used to detect the expressions of apoptosis related protein Bax, cleaved-caspase3 and Bcl-2 protein. Results With the prolongation of cell culture time, the inhibition rate of mBD2 transfection group increased gradually, at the same time point, compared with control and NC groups, the cell inhibition rate of mBD2 transfection group increased ( P <0.05). Compared with control and NC groups, the number of cell colony decreased in mBD2 transfection group, the apoptosis rate of mBD2 transfection group was increased ( P <0.05). Compared with control and NC groups, the G1 phase DNA level of mBD2 transfection group increased ( P <0.05). There was no significant change in G2 and S phase DNA levels among three groups ( P <0.05). Compared with control and NC groups, the expressions of Bax and cleaved-caspase3 protein in mBD2 transfection group increased, the expression of Bcl-2 protein was decreased ( P <0.05). Conclusions MBD2 might inhibit the proliferation of SW480 cells and promote its apoptosis by blocking cell in G1 phase and up-regulating expression of apoptosis protein and play the antitumor effect.