丁苯酞通过混合谱系激酶3信号通路对1-甲基-4-苯基-吡啶离子诱导的SH-SY5Y细胞增殖和凋亡的影响

doi:10.3969/j.issn.1006-9771.2017.11.009

《中国康复理论与实践》 ›› 2017, Vol. 23 ›› Issue (11): 1284-1289.doi: 10.3969/j.issn.1006-9771.2017.11.009

丁苯酞通过混合谱系激酶3信号通路对1-甲基-4-苯基-吡啶离子诱导的SH-SY5Y细胞增殖和凋亡的影响

郭子梦¹, 吴庆文¹, 陈秀秀¹, 关亚丽², 李鹏飞², 王妍², 程月发²

1.华北理工大学护理与康复学院,河北唐山市 063210;
2.华北理工大学冀唐学院,河北唐山市 063210。

收稿日期:2017-06-12 出版日期:2017-11-25 发布日期:2017-12-10
作者简介:郭子梦(1990-),女,汉族,河北新乐市人,硕士研究生,主要研究方向：神经康复。通讯作者：吴庆文、程月发。E-mail: wxywqw@163.com (吴庆文);arthurcy@163.com (程月发)。
基金资助:
1; 华北理工大学研究生创新项目(No.2017S37); 2; 华北理工大学博士科研启动基金项目(No.35647699); 3; 河北省高等学校科学研究青年基金项目(No; QN201722)

Effects of DL-3-n-Butylphthalide on Proliferation and Apoptosis of 1-Methyl-4-Phenylpyridinium-induced SH-SY5Y Cells via Mixed Lineage Kinase 3 Signaling Pathway

GUO Zi-meng¹, WU Qing-wen¹, CHEN Xiu-xiu¹, GUAN Ya-li², LI Peng-fei², WANG Yan², CHENG Yue-fa²

1. College of Nursing and Rehabilitation, North China University of Science and Technology, Tangshan, Hebei 063210, China;
2. Jitang College of North China University of Science and Technology, Tangshan, Hebei 063210, China

Received:2017-06-12 Published:2017-11-25 Online:2017-12-10
Contact: WU Qing-wen, CHENG Yue-fa. E-mail: wxywqw@163.com (WU Qing-wen); arthurcy@163.com (CHENG Yue-fa)

摘要/Abstract

摘要： 目的探讨丁苯酞对1-甲基-4-苯基-吡啶离子(MPP⁺)诱导的SH-SY5Y细胞混合谱系激酶3 (MLK3)通路的影响,及其对细胞增殖与凋亡的作用机制。方法对数生长期SH-SY5Y细胞分为对照组、MPP⁺组、丁苯酞组和URMC-099组,对照组正常培养,MPP⁺组加1 mmol/L MPP⁺培养24 h,丁苯酞组予10 µmol/L丁苯酞预处理3 h后,加入MPP⁺培养24 h,URMC-099组予200 nmol/L MLK3通路特异性抑制剂URMC-099预处理3 h后,加入MPP⁺培养24 h。倒置相差显微镜观察细胞形态,噻唑蓝比色法检测细胞活性,Annexin-V/PI双染流式细胞术检测细胞凋亡率,Hoechst33342荧光染色法观察凋亡细胞,Western blotting检测MLK3磷酸化蛋白(p-MLK3)、c-Jun氨基末端激酶磷酸化蛋白(p-JNK)和细胞外调节蛋白激酶磷酸化蛋白(p-ERK1/2)的表达。结果 MPP⁺组细胞存活率低于对照组(P<0.05),丁苯酞组和URMC-099组细胞存活率高于MPP⁺组(P<0.05);MPP⁺组细胞凋亡率高于对照组(P<0.05),丁苯酞组和URMC-099组细胞凋亡率低于MPP⁺组(P<0.05);与对照组相比,MPP⁺组p-MLK3、p-JNK蛋白表达量增加(P<0.05),p-ERK1/2蛋白表达量降低(P<0.05);与MPP⁺组相比,丁苯酞组和URMC-099组p-MLK3、p-JNK蛋白表达量降低(P<0.05),p-ERK1/2蛋白表达量升高(P<0.05)。结论丁苯酞可减少MPP⁺诱导的SH-SY5Y细胞凋亡,促进细胞增殖,其机制可能是通过抑制MLK3通路,调节下游p-JNK、p-ERK1/2蛋白表达。

关键词: 帕金森病, 丁苯酞, 混合谱系激酶3, 凋亡, SH-SY5Y细胞

Abstract: Objective To investigate the effects of DL-3-n-Butylphthalide (NBP) on proliferation and apoptosis of 1-methyl-4-phenylpyridinium (MPP⁺)-induced SH-SY5Y cells, and mechanisms via mixed lineage kinase 3 (MLK3) signaling pathway.Methods The SH-SY5Y cells were divided into control group, MPP⁺ group, NBP group and URMC-099 group, that cultured normally, with 1 mmol/L MPP⁺ for 24 hours, with 10 µmol/L NBP for 3 hours and then with MPP⁺ for 24 hours, and with 200 nmol/L MLK3 inhibitor URMC-099 for 3 hours and then with MPP⁺ for 24 hours, respectively. The morphology of SH-SY5Y cells was observed under inverted phase contrast microscope and the survival rate was measured with 3-(4,5-Cimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide assays. The apoptosis was quantified under flow cytometry with Annexin V/PI fluorescence staining, and the nuclear morphology was observed with Hoechst 33342 staining. The expression of phosphorylated protein of MLK3 (p-MLK3), c-Jun N-terminal kinase (p-JNK), extra cellular regulated protein kinases (p-ERK1/2) were detected with Western blotting.Results Compared with the control group, the survival rate reduced and apoptosis increased in MPP⁺ group (P<0.05), with the increase of p-MLK3 and p-JNK and decrease of p-ERK1/2 d (P<0.05). Compared with MPP⁺ group, the survival rate increased and apoptosis reduced in both NBP and URMC-099 groups (P<0.05), with the decrease of p-MLK3 and p-JNK and increase of p-ERK1/2 (P<0.05).Conclusion NBP can decrease the apoptosis and promote the proliferation of SH-SY5Y cells induced by MPP⁺, which may be associated with inhibiting MLK3 signaling pathway, and regulating the downstream p-JNK and p-ERK1/2.

Key words: Parkinson's disease, DL-3-n-butylphthalide, mixed-lineage kinase 3, apoptosis, SH-SY5Y cells

中图分类号:

R742.5

郭子梦, 吴庆文, 陈秀秀, 关亚丽, 李鹏飞, 王妍, 程月发. 丁苯酞通过混合谱系激酶3信号通路对1-甲基-4-苯基-吡啶离子诱导的SH-SY5Y细胞增殖和凋亡的影响[J]. 《中国康复理论与实践》, 2017, 23(11): 1284-1289.

GUO Zi-meng, WU Qing-wen, CHEN Xiu-xiu, GUAN Ya-li, LI Peng-fei, WANG Yan, CHENG Yue-fa. Effects of DL-3-n-Butylphthalide on Proliferation and Apoptosis of 1-Methyl-4-Phenylpyridinium-induced SH-SY5Y Cells via Mixed Lineage Kinase 3 Signaling Pathway[J]. 《Chinese Journal of Rehabilitation Theory and Practice》, 2017, 23(11): 1284-1289.

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丁苯酞通过混合谱系激酶3信号通路对1-甲基-4-苯基-吡啶离子诱导的SH-SY5Y细胞增殖和凋亡的影响

Effects of DL-3-n-Butylphthalide on Proliferation and Apoptosis of 1-Methyl-4-Phenylpyridinium-induced SH-SY5Y Cells via Mixed Lineage Kinase 3 Signaling Pathway

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