槲皮素诱导人结肠癌细胞HT-29凋亡的机制研究

Mechanism of Apoptosis Induced by Quercetin in Human Colon Cancer HT-29 Cells

关键词:  槲皮素  人结肠癌细胞  凋亡  信号通路

Key words:  quercetin  human colon cancer cells  apoptosis  signal pathway

摘要:

目的 探讨槲皮素在体外对人结肠癌HT-29细胞凋亡的影响，并对其诱导机制进行研究。方法 体外常规培养HT-29细胞，随机设定对照组和高、中、低剂量（32，16，8 μg·mL^-1）槲皮素组，药物干预24 h后，采用CCK-8试剂盒检测槲皮素对HT-29细胞增殖的影响，倒置显微镜观察细胞形态变化，蛋白免疫印迹法检测MKK3/6/p38信号通路蛋白表达，并同时检测凋亡相关蛋白Caspase 8、Caspase 3、Bax及Bcl-2的表达水平。结果 槲皮素干扰HT-29细胞24 h后，CCK-8实验发现槲皮素可明显抑制细胞的增殖；形态学观察发现细胞密度明显减小、细胞体积变小；免疫印迹实验发现槲皮素可诱导MKK3/6及p38的磷酸化，进而促进促凋亡蛋白Caspase8、Caspase3及Bax的表达，抑制了抗凋亡蛋白Bcl-2的表达。结论 槲皮素可诱导人结肠癌HT-29细胞凋亡，其作用机制可能与激活MKK3/6/p38信号通路有关。

Abstract:

OBJECTIVE To investigate the influence of quercetin on the apoptosis of human colon cancer HT-29 cells in vitro and study the induced mechanism. METHODS Human colon cancer HT-29 cells were cultured in vitro and divided into 4 groups, including control group, high-dose quercetin group (32 μg·mL^-1), medium-dose quercetin group (316 μg·mL^-1) and low-dose quercetin group (8 μg·mL^-1). After quercetin intervention for 24 h, CCK-8 kit was used to observe the effect of quercetin on HT-29 cells proliferation; alterations of HT-29 cells morphology were determined using inverted microscope; Western blot technology was used to detect the expression levels of MKK3/6, p38, p-MKK3/6, p-p38, Caspase 8, Caspase 3, Bax and Bcl-2. RESULTS These findings showed that quercetin could significantly inhibite the proliferation of HT-29 cells in vitro; HT-29 cells density significantly decreased and cells size significantly lessened after quercetin intervention for 24 h; moreover, the phosphorylation levels of MKK3/6 and p38 were significantly rised induced by quercetin, and pro-apoptotic protein (Caspase8, Caspase3, Bax) significantly up-regulated while anti-apoptotic protein (Bcl-2) significantly inhibited by quercetin. CONCLUSION Quercetin can induce the apoptosis of human colon cancer HT-29 cells and the mechanism may relate to the activation of MKK3/6/p38 signal pathway.