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ER-α36和Akt在 PC12细胞缺糖应激反应中的关系

梁小峰, 方晨, 马依妮, 关心, 刘洋, 韩朝, 刘晶, 邹伟

辽宁师范大学，生命科学学院，大连 116021；辽宁师范大学，发展和教育心理学研究中心，大连 116029；大连医科大学附属第一医院，中英再生医学中心，大连 116011

摘要

ER-α36是一种分子量为36 kD的雌激素受体（ER）新亚型。已有文献报道，ER-α36主要通过介导膜雌激素信号通路参与多种细胞生理、病理等过程。本文利用RNA干扰技术敲低PC12(大鼠肾上腺嗜铬细胞瘤细胞)细胞的ER-α36基因，研究了ER-α36与Akt在神经细胞缺糖损伤中的关系。结果发现（1）随着缺糖损伤时间的增加，细胞的存活率逐渐降低，凋亡率升高，6 h后具有极显著性差异（p<0.01）；葡萄糖转运蛋白Glut-4表达水平显著降低（p<0.01）；（2）缺糖损伤3 h时，ER-α36明显减少（p<0.01），但随后逐渐增加；缺糖可引起抗凋亡蛋白Akt 磷酸化水平先升高后降低（p<0.05）；（3）敲低ER-α36后缺糖处理，细胞凋亡率高于野生型，具有极显著性差异（p<0.01）；进一步发现，敲低ER-α36的细胞缺糖导致的Akt激活减弱，Caspase-3表达增多。结果说明，在PC12细胞的缺糖中，Akt激活情况可能与ER-α36相关，两者共同参与缺糖损伤应激调控，机制有待进一步研究。

关键词： ER-α36; 缺糖损伤; Akt抗凋亡信号通路; PC12细胞

分类号：Q256

[Relationship between ER-α36 and Akt in PC12 cells exposed to glucose deprivation.] [Article in Chinese]

LIANG Xiao-Feng, FANG Chen, MA Yi-Ni, GUAN Xin, HAN Chao, LIU Yang, LIU Jing, ZOU Wei

College of Life Science, Liaoning Normal University, Dalian 116021, China；The Research Center of Developmental and Educational Psychology, Liaoning Normal University, Dalian 116029, China； Regenerative Medicine Centre, First Affiliated Hospital of Dalian Medical University, Dalian 116011, China.

Abstract

ER-α36 is a novel 36-kDa variant of ER-α. A large of evidence demonstrated that ER-α36 responded to membrane-initiated estrogen signaling pathways which were involved in the physiological and pathological process in many kinds of cells. In this study, knock-down of ER-α36 expression in pheochromocytoma (PC12) cells (named as PC12-36L cells) by using the shRNA method was used to evaluate the relationship between ER-α36 and Akt in neurons under glucose deprivation. The effect of ER-α36 on outgrowth of PC12 cells, as well as the neuroprotective effect of ER-α36 on injured PC12 cells exposed to glucose deprivation was observed by using MTT assay, Western blot and Annexin V/PI staining et al. The results showed that, (1) Glucose deprivation induced by MEM treatment for 6 h reduced survival rate and increased apoptotic rate in PC12 cells significantly compared to control group (P < 0.01); and it produced a decrease in the expression of Glut-4 protein (P < 0.01); (2) The expression level of ER-α36 was decreased significantly at 3 h of glucose deprivation, and then increased, while phosphorylation of Akt participated in the glucose deprivation was increased at first and then reduced; LY294002 (PI3K inhibitor) contributed to decreased expression of ER-α36, and suppressed the activation of Akt; (3) The rate of apoptosis was significantly increased in PC12-36L cells after glucose deprivation compared with that in wild type PC12 cells (P < 0.01). Furthermore, phosphorylation of Akt was decreased and Caspase-3 was increased by glucose deprivation in PC12-36L cells compared with those in wild type PC12 cells. The study reveals that phosphorylation of Akt is associated with ER-α36 in PC12 cells exposed to glucose deprivation, and both are involved in the regulation of stress responses.

Key words: ER-α36; glucose-deprivation; Akt signaling pathway; PC12 cell

收稿日期：2012-11-22　　录用日期：2013-04-22

通讯作者：邹伟　　E-mail: weizou60@126.com

引用本文：

梁小峰, 方晨, 马依妮, 关心, 刘洋, 韩朝, 刘晶, 邹伟. ER-α36和Akt在 PC12细胞缺糖应激反应中的关系[J]. 生理学报 2013; 65 (4): 381-388.

LIANG Xiao-Feng, FANG Chen, MA Yi-Ni, GUAN Xin, HAN Chao, LIU Yang, LIU Jing, ZOU Wei. [Relationship between ER-α36 and Akt in PC12 cells exposed to glucose deprivation.] [Article in Chinese]. Acta Physiol Sin 2013; 65 (4): 381-388 (in Chinese with English abstract).