沉默JAG1基因对人乳腺癌MDA-MB-231细胞增殖和凋亡的影响

Effects of JAG1 gene silencing on proliferation and apoptosis of human breast cancer MDA-MB-231 cells

目的:探究沉默Jagged 1(JAG1)基因对人乳腺癌MDA-MB-231细胞增殖和凋亡的影响及其分子生物学机制。方法:用已构建的pRS-JAG1重组质粒转染人乳腺癌MDA-MB-231细胞,采用Western blotting方法检测重组质粒对JAG1蛋白表达的影响;MTT比色法测定沉默JAG1对细胞生长的抑制情况;流式细胞术检测细胞周期和细胞凋亡;蛋白印记分析细胞周期蛋白1(cyclin D1)、p21CIP1/WAF1、p27KIP1、p-Rb、Bcl-2、Bax、Bcl-xL和cleaved caspase-3蛋白水平的变化。结果:Western blotting结果证实重组质粒可在72h内有效抑制JAG1蛋白表达;人乳腺癌MDA-MB-231细胞JAG1被沉默后,细胞的生长速度明显减慢,细胞明显阻滞于G0/G1期,细胞凋亡率显著升高(P<0.05),cyclin D1、p-Rb、Bcl-2和Bcl-xL蛋白水平被下调(P<0.05),而p21CIP1/WAF1、p27KIP1、Bax和cleaved caspase-3的蛋白水平显著升高(P<0.05)。结论:沉默JAG1可有效抑制人乳腺癌MDA-MB-231细胞增殖,并诱导其凋亡。本研究为以JAG1为分子靶点的三阴乳腺癌治疗提供实验依据。

AIM ： To investigate the effects of Jagged 1 （JAG1） gene silencing on the proliferation and apopto- sis of human breast cancer MDA-MB-231 cells. METHODS： The specific recombinant vector pRS-JAG1 was transfected into MDA-MB-231 cells with lipofectamine. The protein expression of JAG1 was observed by Western blotting after trans- fection. MTT assay was used to detect the effect of JAG1 gene silencing on the growth of the cells. The apoptosis and cell cycle were analyzed by flow cytometry. The protein levels of cyclin D1, p21^CIPI/WAFI, p27^KIPI, p-Rb, Bcl-2, Bax, Bcl-xL and cleaved caspase-3 were determined by Western blotting. RESULTS： Compared with control group, the expression lev- el of JAGI was reduced by pRS-JAG1 transfection for 72 h （P 〈 0.05 ）. The growth of MDA-MB-231 cells in shJAG1 group was significantly inhibited （P 〈 0.05 ）. The percentages of G0/G1-phase cells and early apoptotic rate were obviously higher in shJAG1 group than those in control group （P 〈 0.05 ）. The shRNA-mediated JAG1 silencing decreased the pro- tein levels of cyclin D1, p-Rb, Bcl-2 and Bax, and increased the protein levels of p21^CIPI/WAFI, p27^KIPI, Bax and cleaved caspase-3 （ P 〈 0.05）. CONCLUSION ： JAG1 silencing effectively inhibits the proliferation and induces the apoptosis of human breast cancer cells, suggesting that JAG1 might serve as a therapeutic target for triple-negative breast cancer.