中文摘要：

目的研究东亚钳蝎氯离子通道毒素（BmK CT）对胶质瘤U251细胞增殖的抑制作用及其机制。方法制备融合蛋白GST-BmK CT,体外检测GST-BmK CT对胶质瘤U251细胞增殖及细胞周期的影响。以GST-BmK CT处理组为实验组,以GST处理组为阴性对照,PBS处理组为空白对照。MTT法分析各处理组U251细胞的增殖速度;流式细胞术分析U251细胞周期变化;蛋白免疫印迹法检测MAPKAPK-2的蛋白的磷酸化及其上游蛋白p38的磷酸化。预先使用p38α特异性抑制剂SB203580处理细胞,检测p38α活化对细胞周期阻滞的影响。结果 MTT结果表明,融合蛋白GST-BmK CT体外处理U251细胞48 h和96 h,增殖抑制率分别为（25±1.9）%和（33±2.3）%,与GST处理组相比差异有统计学意义（P〈0.001）。流式细胞检测结果表明,处理U251细胞96 h,GST-BmK CT组比空白对照组S期和G2/M期比例增多,差异有统计学意义（P〈0.005）,GST阴性对照组相比空白组细胞周期各时相差异均无统计学意义（P〉0.05）。蛋白免疫印迹结果显示GST-BmK CT能激活p38α丝裂原活化蛋白激酶（p38αMAPK）,而用p38α特异性抑制剂SB203580能反转GST-BmK引起的细胞周期阻滞。结果进一步表明,BmK CT通过活化p38α激活下游丝裂原活化蛋白激酶激活的蛋白激酶-2（MAPKAPK-2,MK2）从而引起细胞S期和G2/M期的阻滞。结论 GST-BmK CT是通过p38αMAPK信号通路激活MK2来引起U251细胞周期在S期和G2/M期的阻滞,从而抑制细胞增殖。

英文摘要：

Objective To explore the inhibitive effect of BmK CT on proliferation of glioma U251 cells and its possible mechanism.Methods U251 cells were treated with GST-BmK CT in vitro to evaluate the effects on proliferation and cell cycle. U251 cells were treated with GST-BmK CT（ experiment group）,GST（ GST group） and PBS（ PBS group）,respectively. MTT was used to analyze the proliferation of U251 cells,and flow cytometry was used to analyze the cell cycle of U251 cells. Western blot was used to detect the expression of related proteins. Effect of pretreatment with SB203580 on cell cycle was determined. Results MTT results showed that the inhibition rates were（25 ±1. 9）% and（33 ±2. 3）% in U251 cells after treated with GST-BmK CT in vitro for 48 h and 96 h,which were significantly different from those of GST group（ P 〈 0. 001）. Flow cytometry analysis found that the cells in S phase and G2/M phase were increased in GSTBmK CT group after treatment for 96 h compared with PBS group（ P 〈 0. 005）,but there was no significant difference between GST group and PBS group during the cell cycle phase（ P 〉 0. 05）. Western blot showed that GST-BmK CT activated p38α-mitogen-activated protein kinase（ p38αMAPK）,and p38α inhibitor SB203580 significantly reversed cell cycle arrest triggered by GST-BmK CT. MAPKAPK-2 was phosphorylated to induce S and G2/M phase arrest. Conclusion GST-BmK CT may cause U251 cell cycle arrest in the S phase and G2/M phase by activating MK2 through the p38αMAPK signal pathway to inhibit the proliferation of U251 cells.