吉林大学学报(医学版) ›› 2026, Vol. 52 ›› Issue (1): 152-161.doi: 10.13481/j.1671-587X.20260116

• 基础研究 • 上一篇    下一篇

溶血磷脂酸联合6-羟基多巴胺对SH-SY5Y细胞凋亡的诱导作用及其机制

李亚萍1,2,谭春艳3,赵璐洁1,2,赵嘉怡1,2,李婷2,杨晓4(),杨晓云1,2()   

  1. 1.山东第二医科大学基础医学院生物化学与分子生物学系,山东 潍坊 261053
    2.山东第二医科大学基础医学院智能诊疗与分子疾病协同创新重点实验室,山东 潍坊 261053
    3.山东第二医科大学基础医学院生理学系,山东 潍坊 261053
    4.山东第二医科大学护理学院护理学系,山东 潍坊 261053
  • 收稿日期:2025-01-16 接受日期:2025-03-31 出版日期:2026-01-28 发布日期:2026-02-24
  • 通讯作者: 杨晓,杨晓云 E-mail:yx0204@163.com;yangxiaoyun@sdsmu.edu.cn
  • 作者简介:李亚萍(1997-),女,内蒙古自治区乌兰察布市人,在读硕士研究生,主要从事溶血磷脂酸生化药理学方面的研究。
  • 基金资助:
    山东省卫健委医药卫生科技发展计划项目(2018WS063);山东省教育厅基金项目(J14LK15);山东省教育厅国家级大学生创新创业训练计划项目(202010438026);山东第二医科大学博士科研启动基金科研创新计划项目(2024BKQ048)

Inductive effect of lysophosphatidic acid combined with 6-hydroxydopamine on apoptosis of SH-SY5Y cells and its mechanism

Yaping LI1,2,Chunyan TAN3,Lujie ZHAO1,2,Jiayi ZHAO1,2,Ting LI2,Xiao YANG4(),Xiaoyun YANG1,2()   

  1. 1.Department of Biochemistry and Molecular Biology,Shandong Second Medical University,Weifang 261053,China
    2.Weifang Key Laboratory of Collaborative Innovation of Intelligent Diagnosis and Treatment and Molecular Diseases,School of Basic Medical Sciences,Shandong Second Medical University,Weifang 261053,China
    3.Department of Physiology,Shandong Second Medical University,Weifang 261053,China
    4.Department of Nursing,Shandong Second Medical University,Weifang 261053,China
  • Received:2025-01-16 Accepted:2025-03-31 Online:2026-01-28 Published:2026-02-24
  • Contact: Xiao YANG,Xiaoyun YANG E-mail:yx0204@163.com;yangxiaoyun@sdsmu.edu.cn

摘要:

目的 探讨溶血磷脂酸(LPA)联合6-羟基多巴胺(6-OHDA)对SH-SY5Y细胞凋亡的诱导作用,并阐明其相关作用机制。 方法 采用100 μmol·L-1 6-OHDA建立帕金森病(PD)细胞模型,选用SH-SY5Y细胞、PC12细胞和N2a细胞,分为对照组(仅用不完全培养基)、4 μmol·L-1 LPA组、100 μmol·L-1 6-OHDA组、4 μmol·L-1 LPA+100 μmol·L-1 6-OHDA组、10 μmol·L-1 LPA组和10 μmol·L-1 LPA+100 μmol·L-1 6-OHDA组,处理24 h。采用噻唑蓝(MTT)法检测各组SH-SY5Y细胞活性,异硫氰酸荧光素标记的膜联蛋白Ⅴ(Annexin Ⅴ-FITC)/碘化丙啶(PI)染色结合流式细胞术检测各组SH-SY5Y细胞凋亡率,Western blotting法检测3种细胞中含半胱氨酸的天冬氨酸蛋白水解酶3(Caspase-3)相关蛋白表达水平以及SH-SY5Y细胞中B细胞淋巴瘤-2(Bcl-2)、Bcl-2相关X蛋白(Bax)、p38丝裂原活化蛋白激酶(p38 MAPK)和磷酸化的p38 MAPK(p-p38 MAPK)蛋白以及LPA1受体蛋白表达水平。 结果 与100 μmol·L-1 6-OHDA组比较,4 μmol·L-1 LPA+100 μmol·L-1 6-OHDA组和10 μmol·L-1 LPA+100 μmol·L-1 6-OHDA组SH-SY5Y细胞数和细胞活性均明显降低(P<0.05或P<0.01),细胞凋亡率升高(P<0.05或P<0.01)。与100 μmol·L-1 6-OHDA组比较,4 μmol·L-1 LPA+100 μmol·L-1 6-OHDA组和10 μmol·L-1 LPA+100 μmol·L-1 6-OHDA组3种细胞中Caspase-3蛋白表达水平无明显变化,差异无统计学意义(P>0.05);4 μmol·L-1 LPA+100 μmol·L-1 6-OHDA组SH-SY5Y细胞中裂解的Caspase-3(Cleaved Caspase-3)蛋白表达水平明显升高(P<0.01);4 μmol·L-1 LPA+100 μmol·L-1 6-OHDA组和10 μmol·L-1 LPA+100 μmol·L-1 6-OHDA组PC12细胞中Cleaved Caspase-3蛋白表达水平明显升高(P<0.05或P<0.01);4 μmol·L-1 LPA+100 μmol·L-1 6-OHDA组N2a细胞中Cleaved Caspase-3蛋白表达水平升高(P<0.05)。与100 μmol·L-1 6-OHDA 组 比 较, 4 μmol·L-1 LPA+100 μmol·L-1 6-OHDA 和 10 μmol·L-1 LPA + 100 μmol·L-1 6-OHDA组SH-SY5Y细胞中Bax和Bcl-2蛋白表达水平无明显变化,差异无统计学意义(P>0.05),Bax/Bcl-2比值升高(P<0.05);10 μmol·L-1 LPA+100 μmol·L-1 6-OHDA组SH-SY5Y细胞中p38 MAPK蛋白表达水平升高(P<0.05),4 μmol·L-1 LPA+100 μmol·L-1 6-OHDA组SH-SY5Y细胞中p-p38 MAPK蛋白表达水平明显升高(P<0.01),p-p38 MAPK/p38 MAPK比值升高(P<0.05);10 μmol·L-1 LPA+100 μmol·L-1 6-OHDA组SH-SY5Y细胞中LPA1受体蛋白表达水平降低(P<0.05)。 结论 LPA与6-OHDA联合应用能明显诱导SH-SY5Y细胞凋亡,其作用机制可能与上调细胞中p38 MAPK和p-p38 MAPK蛋白表达水平有关。

关键词: 溶血磷脂酸, 6-羟基多巴胺, SH-SY5Y细胞, 细胞凋亡, p38丝裂原活化蛋白激酶

Abstract:

Objective To discuss the inductive effect of lysophosphatidic acid (LPA) combined with 6-hydroxydopamine (6-OHDA) on the apoptosis of SH-SY5Y cells, and to clarify its related mechanism. Methods The Parkinson’s disease (PD) cell model was established by using 100 μmol·L-1 6-OHDA; the SH-SY5Y cells, PC12 cells and N2a cells were selected and divided into control group (incomplete medium only), 4 μmol·L-1 LPA group, 100 μmol·L-1 6-OHDA group, 4 μmol·L-1 LPA+100 μmol·L-1 6-OHDA group, 10 μmol·L-1 LPA group, and 10 μmol·L-1 LPA+100 μmol·L-1 6-OHDA group, and treated for 24 h. Methylthiazolydiphenyl-tetrazolium bromide(MTT) method was used to detect the activities of the SH-SY5Y cells in various groups; annexin Ⅴ fluorescein isothiocyanate(Annexin Ⅴ-FITC)/propidium iodide(PI) staining and flow cytometry were used to detect the apoptotic rates of the SH-SY5Y cells in various groups; Western blotting method was used to detect the expression levels of Caspase-3 related proteins in three kinds of cells and the expression levels of B-cell lymphoma-2 (Bcl-2), Bcl-2-associated X protein (Bax), p38 mitogen-activated protein kinase (p38 MAPK), phosphorylated p38 MAPK (p-p38 MAPK) proteins and LPA1 receptor protein in the SH-SY5Y cells in various groups. Results The MTT assay results showed that compared with 100 μmol·L?1 6-OHDA group, the numbers and activities of the SH-SY5Y cells in 4 μmol·L-1 LPA+100 μmol·L-1 6-OHDA group and 10 μmol·L-1 LPA+100 μmol·L-1 6-OHDA group were significantly decreased (P<0.05 or P<0.01). The flow cytometry results showed that compared with 100 μmol·L-1 6-OHDA group, the apoptotic rates of the SH-SY5Y cells in 4 μmol·L-1 LPA+100 μmol·L-1 6-OHDA group and 10 μmol·L-1 LPA+100 μmol·L-1 6-OHDA group were increased (P<0.05 or P<0.01). The Western blotting results showed that compared with 100 μmol·L-1 6-OHDA group, the expression levels of Caspase-3 protein in three kinds of cells in 4 μmol·L-1 LPA+100 μmol·L-1 6-OHDA group and 10 μmol·L-1 LPA+100 μmol·L-1 6-OHDA group had no significant changes, and the differences were not statistically significant (P>0.05); the expression levels of Cleaved Caspase-3 protein in the SH-SY5Y cells in 4 μmol·L-1 LPA+100 μmol·L-1 6-OHDA group were significantly increased (P<0.01); the expression levels of Cleaved Caspase-3 protein in the PC12 cells in 4 μmol·L-1 LPA+100 μmol·L-1 6-OHDA group and 10 μmol·L-1 LPA+100 μmol·L-1 6-OHDA group were significantly increased (P<0.05 or P<0.01); the expression level of Cleaved Caspase-3 protein in the N2a cells in 4 μmol·L?1 LPA+100 μmol·L?1 6-OHDA group was increased (P<0.05). The Western blotting results showed that compared with 100 μmol·L-1 6-OHDA group, the expression levels of Bax and Bcl-2 proteins in the SH-SY5Y cells in 4 μmol·L-1 LPA+100 μmol·L?1 6-OHDA group and 10 μmol·L?1 LPA+100 μmol·L?1 6-OHDA group had no significant changes, and the differences were not statistically significant (P>0.05), but the Bax/Bcl-2 ratio was increased (P<0.05); the expression level of p38 MAPK protein in the SH-SY5Y cells in 10 μmol·L-1 LPA+100 μmol·L-1 6-OHDA group was increased (P<0.05); the expression level of p-p38 MAPK protein in the SH-SY5Y cells in 4 μmol·L-1 LPA+100 μmol·L-1 6-OHDA group was significantly increased (P<0.01), and the p-p38 MAPK/p38 MAPK ratio was increased (P<0.05); the expression level of LPA1 receptor protein in the SH-SY5Y cells in 10 μmol·L-1 LPA+100 μmol·L-1 6-OHDA group was decreased (P<0.05). Conclusion The combined application of LPA and 6-OHDA can significantly induce the apoptosis of SH-SY5Y cells, and its mechanism may be related to the up-regulated expression levels of p38 MAPK and p-p38 MAPK proteins in the cells.

Key words: Lysophosphatidic acid, 6-hydroxydopamine, SH-SY5Y cell, Apoptosis, p38 mitogen-activated protein kinase

中图分类号: 

  • Q545