吉林大学学报(医学版) ›› 2023, Vol. 49 ›› Issue (1): 1-7.doi: 10.13481/j.1671-587X.20230101

• 基础研究 •    下一篇

黄芪甲苷对脂多糖诱导的 MC3T3-E1细胞氧化损伤的保护作用及其机制

邵帅1,鲁美丽2,王洪新2,高秀秋1()   

  1. 1.锦州医科大学附属第二医院牙周科,辽宁 锦州 121000
    2.锦州医科大学药理教研室,辽宁 锦州 121000
  • 收稿日期:2022-05-11 出版日期:2023-01-28 发布日期:2023-02-03
  • 通讯作者: 高秀秋 E-mail:jygaoxiuqiu1988@163.com
  • 作者简介:邵 帅(1994-),男,黑龙江省齐齐哈尔人,在读硕士研究生,主要从事牙周免疫学方面的研究。
  • 基金资助:
    国家自然科学基金项目(81973553);辽宁省辽西地区口腔微创临床治疗中心创新平台项目(201820102)

Protective effect of astragaloside Ⅳ on oxidative injury of MC3T3-E1 cells induced by liposolysaccharide and its mechanism

Shuai SHAO1,Meili LU2,Hongxin WANG2,Xiuqiu GAO1()   

  1. 1.Department of Periodontics,Second Affiliated Hospital,Jinzhou Medical University,Jinzhou 121000,China
    2.Department of Pharmacology,Jinzhou Medical University,Jinzhou 121000,China
  • Received:2022-05-11 Online:2023-01-28 Published:2023-02-03
  • Contact: Xiuqiu GAO E-mail:jygaoxiuqiu1988@163.com

摘要:

目的 探讨黄芪甲苷(AS-Ⅳ)对脂多糖(LPS)诱导的成骨细胞前体细胞MC3T3-E1氧化损伤的保护作用,阐明其相关机制。 方法 将对数生长期MC3T3-E1细胞分为对照组、LPS组和不同浓度(10、25、50及100 mg·L-1) AS-Ⅳ组,采用CCK-8法检测各组细胞活性。将对数生长期MC3T3-E1细胞分为对照组、LPS组、AS-Ⅳ组和AS-Ⅳ+LY294002[磷脂酰肌醇3-激酶/蛋白激酶B(PI3K/Akt)信号通路抑制剂]组,采用流式细胞术检测各组细胞中活性氧(ROS)水平,采用相关试剂盒检测各组细胞中丙二醛(MDA)和谷胱甘肽(GSH)水平及超氧化物歧化酶(SOD)活性,采用实时荧光定量 PCR(RT-qPCR)法检测各组细胞中血红素加氧酶1(HO-1)mRNA表达水平,采用Western blotting法检测各组细胞中HO-1、PI3K、磷酸化Akt(p-Akt)、核因E2相关因子2(Nrf2)、B细胞淋巴瘤2(Bcl-2)和Bcl-2相关X蛋白(Bax)蛋白表达水平。 结果 CCK-8法检测,与对照组比较,LPS组细胞活性明显降低(P<0.01);与LPS组比较,50和100 mg?L-1AS-Ⅳ组细胞活性明显升高(P<0.01)。与对照组比较,LPS组细胞中ROS和MDA水平明显升高(P<0.01),GSH水平和SOD活性明显降低(P<0.01);与LPS组比较,AS-Ⅳ组细胞中ROS和MDA水平明显降低(P<0.01),GSH水平和SOD活性明显升高(P<0.01);与AS-Ⅳ组比较,AS-Ⅳ+LY294002组细胞中ROS和MDA水平明显升高(P<0.01),GSH水平和SOD活性明显降低(P<0.01)。与对照组比较,LPS组细胞中HO-1 mRNA表达水平和HO-1、p-Akt、Nrf2及Bcl-2蛋白表达水平明显降低(P<0.01),Bax蛋白表达水平明显升高(P<0.01);与LPS组比较,AS-Ⅳ组细胞中HO-1 mRNA表达水平和HO-1、p-Akt、Nrf2及Bcl-2蛋白表达水平明显升高(P<0.01),Bax蛋白表达水平明显降低(P<0.01);与AS-Ⅳ组比较,AS-Ⅳ+LY294002组细胞中和HO-1 mRNA表达水平和HO-1、p-Akt、Nrf2及Bcl-2蛋白表达水平明显降低(P<0.01),Bax蛋白表达水平明显升高(P<0.01)。 结论 AS-Ⅳ可以通过介导PI3K/Akt/Nrf2信号通路对LPS诱导的成骨细胞氧化损伤起到保护作用。

关键词: 黄芪甲苷, 脂多糖, 成骨细胞, 氧化损伤

Abstract:

Objective To investigate the protective effect of astragaloside Ⅳ(AS-Ⅳ) on the lipopolysaccharide(LPS)-induced oxidative injury of osteoblasts precursor cells MC3T3-E1, and to clarify its related mechanism. Methods The MC3T3-E1 cells in logarithmic growth phase were divided into control group, LPS group and different concentrations (10,25,50 and 100 mg·L-1) of AS-Ⅳ groups. CCK-8 method was used to detect the cell activities in various groups.The MC3T3-E1 cells in logarithmic growth phase were divided into control group, LPS group, AS-Ⅳ group, AS-Ⅳ+LY294002 [phosphatidylinositol-3-kinase(PI3K)/protein kinase B(Akt)signaling pathway inhibitor]group. Flow cytometry was used to detect the levels of reactive oxygen species (ROS) in the cells in various groups. The levels of malondialdehyde (MDA) and glutathione(GSH) and the activities of superoxide dismutase(SOD) in the cells in various groups were determined with the related kits. The expression levels of heme oxygenase (HO-1) mRNA in the cells in various groups were detected by real-time fluorescence quantitative PCR(RT-qPCR) method. Western blotting method was used to detect the expression levels of PI3K,phosphorylated Akt(p-Akt),and nuclear factor-E2 related factor 2(Nrf2),B-cell lymphoma-2 (Bcl-2), and Bcl-2 associated X protein(Bax) in the cells in various groups. Results The CCK-8 method results showed that compared with control group, the cell activity in LPS group was significantly decreased (P<0.01); compared with LPS group, the activities of cells in 50 and 100 mg·L-1 AS-Ⅳ groups were significantly increased (P<0.01). Compared with control group, the ROS and MDA levels in the cells in LPS group were significantly increased (P<0.01), while the SOD activity and GSH level were decreased (P<0.01); compared with LPS group, the ROS and MDA levels in the cells in AS-Ⅳ group were significantly decreased (P<0.01), the SOD activity and GSH level were significantly increased (P<0.01); compared with AS-Ⅳ group, the ROS and MDA levels in the cells in AS-Ⅳ+LY294002 group were significantly increased (P<0.01), the SOD activity and GSH level were significantly decreased (P<0.01). Compared with control group, the expression level of HO-1 mRNA and the expression levels of p-Akt, Nrf2, HO-1, and Bcl-2 proteins in the cells in LPS group were significantly decreased(P<0.01), while the expression level of Bax protein was significantly increased (P<0.01); compared with LPS group, the expression level of HO-1 mRNA and the expression levels of p-Akt, Nrf2, HO-1,and Bcl-2 proteins in the cells in AS-Ⅳ group were signicantly increased (P<0.01), while the expression level of Bax protein was significantly decreased (P<0.01);compared with AS-Ⅳ group, the expression level of HO-1 mRNA and the expression levels of p-Akt,Nrf2 HO-1 and Bcl-2 proteins in the cells in AS-Ⅳ+ LY294002 group were significantly decreased (P<0.01),and the expression level of Bax protein was significantly increased (P<0.01). Conclusion AS-Ⅳ can play protective effect on the LPS-induced osteoblast injury by mediating PI3K/Akt/Nrf2 signaling pathway.

Key words: Astragaloside Ⅳ, Lipopolysaccharide, Osteoblast, Oxidative injury

中图分类号: 

  • R285.5