天津医药

天津医药 ›› 2019, Vol. 47 ›› Issue (4): 371-376.doi: 10.11958/20182240

• 细胞与分子生物学 • 上一篇    下一篇

胰岛素样生长因子-1对氯化钴诱导的PC12细胞凋亡的抑制作用

马珂 1，2，徐会友 1，2，赵飞 2，张健 1，2，江继鹏 1，2，代晨 1，2，王仁杰 2，陈旭义 2△   

  1. 1天津，中国人民武装警察部队后勤学院（邮编 300162）；2中国人民武装警察部队特色医学中心脑科中心，中国人民武装警察 部队脑创伤与神经疾病研究所，天津市神经创伤修复重点实验室
  • 收稿日期:2019-01-03 修回日期:2019-03-03 出版日期:2019-04-15 发布日期:2019-05-27
  • 通讯作者: 马珂 E-mail:mark1991_1@163.com
  • 作者简介:马珂（1991），男，硕士研究生在读，主要从事神经系统疾病诊疗及神经损伤修复的研究
  • 基金资助:
    国家重点研发计划;国家自然科学基金;天津市救援医学临床中心基金;天津市科技支撑重点项目;天津市自然科学基金项目

The Inhibitory effect of insulin-like growth factor 1 on PC12 cell apoptosis induced by CoCl2

MA Ke1,2, XU Hui-you1,2, ZHAO Fei2, ZHANG Jian1,2, JIANG Ji-peng1,2, DAI Chen1,2, WANG Ren-jie2, CHEN Xu-yi2△   

  1. 1 Logistics University of People’s Armed Police Forces, Tianjin 300162, China; 2 Department of Neurology, People’s Armed Police Forces Medical Center, Institution of Brain Trauma and Neurology Disease of People’s Armed Police Forces, Tianjin Key Laboratory of Neurotrauma Repair
  • Received:2019-01-03 Revised:2019-03-03 Published:2019-04-15 Online:2019-05-27
  • Contact: Ke MA E-mail:mark1991_1@163.com

PDF (PC)

3826

摘要： 目的 探讨胰岛素样生长因子-1（IGF-1）对氯化钴（CoCl2）诱导的肾上腺嗜铬细胞瘤 PC12细胞凋亡的作 用及机制。方法 取对数生长期的 PC12细胞，分别以 10、20、40、80 μmol/L的 CoCl2溶液和 100、200、400 μg/L的 IGF- 1溶液处理细胞，CCK-8法检测细胞活力，得出 CoCl2和 IGF-1最佳干预浓度。根据选定的实验条件，应用 CoCl2处理 PC12细胞建立 HIE细胞模型，实验分为对照组、CoCl2处理组、IGF-1+CoCl2处理组。分组处理 24 h后采用 CCK-8法 检测细胞存活率；TUNEL染色检测细胞凋亡情况；Western blot检测各组细胞 Bax、Bcl-2及 Caspase-3的蛋白的表达 水平。最后在上述实验的基础上，设 CoCl2处理组、CoCl2+IGF-1处理组、HIF-1α抑制剂 2-甲氧雌二醇（2-MeOE2）+ CoCl 2组和 CoCl2+2-MeOE2+IGF-1组，Western blot观察 IGF-1、2-MeOE2及两者共用对 PC12细胞 HIF-1α及 Bax的表 达水平的影响。结果 CCK-8检测结果显示，40 μmol/L CoCl2和 200 μg/L IGF-1为最佳干预浓度。利用以上浓度分 组干预细胞 24 h后，与 CoCl2组相比，IGF-1+CoCl2处理组细胞存活率明显提升，TUNEL阳性细胞数和细胞比例降低， 同时抗凋亡蛋白 Bcl-2表达上调，促凋亡蛋白 Bax、Caspase-3，HIF-1α表达下调，差异均有统计学意义（均P＜0.05）。 应用 2-MeOE2对细胞进行预处理后，CoCl2+2-MeOE2组与 2-MeOE2+IFG-1组 HIF-1α和 Bax蛋白表达差异无统计 学意义，但均较 CoCl2+IGF-1组明显下调（P＜0.01）。结论 IGF-1可抑制 CoCl2诱导的 PC12细胞凋亡，其保护作用 可能与 HIF-1α表达下调有关。

关键词: 胰岛素样生长因子Ⅰ, 缺氧缺血, 脑, 细胞凋亡, 缺氧诱导因子 1, α亚基, PC12细胞, 氯化钴

Abstract: Objective To explore the effect and mechanism of insulin-like growth factor 1 (IGF - 1) on cobalt chloride （CoCl2) induced apoptosis of pheochromocytoma (PC12) cells. Methods PC12 cells in logarithmic growth phase were treated with 10, 20, 40 and 80 μmol/L CoCl2 solution and 100, 200, 400 μg/L IGF-1 solution. CCK-8 assay was used to detect cell viability, and the optimal intervention concentrations of CoCl2 and IGF-1 were obtained. According to the selected experimental conditions, PC12 cells were treated with CoCl2 to establish HIE cell model. The experimental cells were divided into control group, CoCl2 treatment group and IGF-1 + CoCl2 treatment group. After 24 h of preincubation in each group, CCK-8 assay was used to detect cell survival rate, TUNEL staining was used to detect the cell apoptosis, and Western blot assay was used to detect the protein expression levels of Bax, Bcl-2 and Caspase-3. Finally, IGF-1 inhibitor 2- methoxyestradiol (2-MeOE2) group and 2-MeOE2+IGF-1 group were added on the basis of the above experiments. Western blot analysis was performed to observe the effects of IGF-1, 2-MeOE2 and both of them on HIF-1α and Bax expressions in PC12 cells. Results CCK-8 assay showed that the optimal concentration of CoCl2 was 40 μmol / L, and the optimal concentration of IGF-1 was 200 μg/L. After 24 h of intervention with the concentration of above groups, the cell survival rate was significantly improved in IGF-1 + CoCl2 group compared with the CoCl2 group, and the number proportion of TUNEL (+) cells was significantly lower in the IGF-1 + CoCl2 group than that of CoCl2 group. The expression of anti-apoptotic protein Bcl-2 was up-regulated in the IGF-1 + CoCl2 group, and the expressions of apoptotic protein Bax, caspase-3 and HIF-1α were significantly down-regulated in the CoCl2+ IGF-1 group compared with the CoCl2 group (P＜0.05). However, after the pretreatment with 2-MeOE2, there were no significant differences in HIF-1α and Bax expressions between CoCl2 + 2- MeOE2 group and 2-MeOE2 + IGF-1 group, but they were significantly down-regulated compared with CoCl2 + IGF-1 group (P＜0.01). Conclusion IGF-1 can inhibit the apoptosis of PC12 cells induced by CoCl2, and its protective effect is related to the down-regulation of HIF-1α expression.

Key words: insulin-like growth factor Ⅰ, hypoxia-ischemia, brain, apoptosis, hypoxia-inducible factor 1, alpha subunit, PC12 cells, CoCl2