氯喹对三阴乳腺癌MDA-MB-231细胞增殖的影响及其机制

目的观察氯喹对三阴乳腺癌MDA - MB - 231细胞增殖的影响,并探讨其机制。方法用氯喹20,40和80 μmol·L^-1分别处理MDA-MB-231细胞24和48 h,采用CCK - 8法和细胞克隆形成实验检测细胞增殖,流式细胞术检测细胞周期和凋亡,Western印迹法检测细胞周期相关蛋白即细胞周期蛋白D3、细胞周期蛋白依赖性激酶2(CDK2)和CDK4,细胞凋亡相关蛋白即活化胱天蛋白酶3和活化聚腺苷二磷酸-核糖聚合酶(PARP)及自噬标志蛋白即自噬微管相关蛋白轻链3B(LC3B)和自噬底物SQSTM1表达水平。结果氯喹20,40和80 μmol·L^-1与MDA - MB-231细胞作用24和48 h后,均能有效抑制细胞增殖(P<0.01)。与细胞对照组相比,氯喹20和40 μmol·L^-1处理24 h,G0/G1期细胞百分比显著增高(P<0.01);80 μmol·L^-1组G2/M期细胞百分比升高(P<0.01),且细胞凋亡率增加(P<0.01)。处理48 h,与细胞对照组相比,氯喹用药3组细胞凋亡率均显著升高(P<0.05)。氯喹3组处理24 h,与细胞对照组相比,细胞周期蛋白D3、CDK2和CDK4表达水平降低(P<0.01),活化胱天蛋白酶3和活化PARP表达增强(P<0.01),LC3B和SQSTM1表达水平显著升高(P<0.01)。结论氯喹可通过抑制MDA-MB-231细胞自噬、阻滞细胞周期进程并促进细胞凋亡而抑制其增殖。

Effect of chloroquine on proliferation of triple negative breast cancer MDA-MB-231 cells and its underlying mechanism

OBjECTIVE To explore the effect of chloroquine(CQ) on the proliferation of triple negative breast cancer MDA-MB-231 cells and its underlying mechanism. METHODS MDA-MB-231 cells were treated with CQ 20, 40 and 80 μmol·L^-1 for 24 and 48 h, respectively. CCK-8 and colony formation assays were used to detect the proliferation of MDA-MB-231 cells. Flow cytometry was employed to analyze cell cycle and apoptosis. Western blotting was used to detect the expressions of cell cycle proteins, apoptosis-related proteins and autophagy-related markers. RESULTS CQ 20, 40 and 80 μmol·L^-1 could effectively inhibit the proliferation of MDA-MB-231 cells both at 24 and 48 h. Compared with the cell control group, the percentage of G0/G1 cells significantly increased when exposed to CQ 20 and 40 μmol·L^-1 for 24 h (P<0.01), the percentage of G2/Mcells increased (P<0.01) and the apoptosis was relatively up-regulated (P<0.01) after treatment with CQ 80 μmol·L^-1 for 24 h. The apoptosis was significantly elevated after exposure to CQ 20, 40 and 80 μmol·L^-1 for 48 h in a concentration-dependent manner (P<0.05). After treatment with CQ, the expression of cell cycle proteins, such as cyclin-dependent kinase-2 (CDK2), CDK4 and cyclin D3, was decreased (P<0.01), the expression of apoptosis-related proteins cleaved-caspase 3 and cleaved- poly-ADP-ribose polymerase was both elevated (P<0.01), and the expression of autophagy-related markers microtubule-associated protein1 light chain 3 and SQSTM1 was both increased (P<0.01). CONCLUSION CQ can effectively suppress the proliferation of TNBC MDA-MB-231 cells by inhibiting autophagy, inducing cell cycle arrest and promoting apoptosis.