摘要
【目的】观察缺氧大鼠心肌细胞和经红景天苷(salidroside,SAL)干预的大鼠心肌细胞蛋白表达差异,探讨SAL对缺氧心肌的保护作用机制。【方法】以H9c2大鼠心肌细胞为靶细胞,使用不同浓度的氯化钴(CoCl2)和SAL处理H9c2细胞,筛选适宜的CoCl2刺激浓度和最佳的SAL保护浓度。采用稳定同位素标记细胞结合质谱的方法筛选缺氧组、红景天苷预处理组和对照组的差异表达蛋白,运用生物信息学对数据进行分析。【结果】CoCl2对H9c2细胞的半数抑制浓度为500μmol/L。常氧状态下SAL对H9c2细胞无明显促进增殖作用。低氧状态下SAL使H9c2细胞活力升高,且100 nmol/L时效果最佳(P<0.05)。缺氧组较正常组,共鉴定到30个上调蛋白和62个下调蛋白;SAL预处理组较缺氧组,共鉴定到70个上调蛋白和36个下调蛋白。SAL保护缺氧H9c2细胞的作用机制主要集中于细胞糖代谢如乙酰辅酶A代谢、三羧酸循环、氧化呼吸等生理过程。【结论】SAL通过调节三羧酸循环在保护心肌细胞免受缺氧损伤中发挥关键作用,对琥珀酰辅酶A连接酶1(succinyl-CoA ligase-1,SUCLG1)、琥珀酰辅酶A连接酶2(succinyl-CoA ligase-2,SUCLG2)、苹果酸脱氢酶1(malate dehydrogenase-1,MDH1)、苹果酸脱氢酶2(malate dehydrogenase-2,MDH2)的调控作用尤为突出。
【Objective】To observe the difference of protein expression in myocardial cells of hypoxic rats and myocardial cells treated with salidroside(SAL),and explore the protective mechanism of SAL on hypoxic myocardium.【Methods】H9 c2 cells were stimulated with different doses of CoCl2 to select the appropriate CoCl2 stimulation concentration.Then appropriate CoCl2 concentration was used to stimulate the H9 c2 cells followed by the treatment with different concentrations of SAL for different periods of time.The optimal concentration of SAL was found.Stable isotope labeling with amino acids was utilized in cell culture(SILAC)coupled to mass spectrometry to screen the different expression proteins in different groups.Raw data were analyzed with DAVID online software.【Results】The IC50 value for CoCl2 was 500μmol/L in H9 c2 cells.SAL had no significant proliferation effect on H9 c2 cells under normoxic conditions.SAL increased the viability of H9 c2 cells under hypoxia conditions,and the optimal protective concentration of SAL was 100 nmol/L(P<0.05).A total of 30 up-regulated proteins and 62 down-regulated proteins were identified in the hypoxic group.Compared with the hypoxic group,70 up-regulated proteins and 36 down-regulated proteins were identified in the SAL pretreated group.The protective mechanism of SAL in protecting hypoxic H9 c2 cells was mainly focused on cellular glucose metabolism,such as acetyl-CoA metabolism,tricarboxylic acid cycle(TCA cycle),and oxidative respiration.【Conclusion】SAL plays a critical role in protecting cardiomyocytes against oxidative injury by restoring the TCA cycle.It has prominent effective on tricarboxylic acid cycle enzymes SUCLG1,SUCLG2,MDH1 and MDH2.
作者
陈茜
徐忠伟
马闻
CHEN Xi;XU Zhong-wei;MA Wen(Sichuan Corps Hospital of PAP,Leshan 614000,China)
出处
《武警后勤学院学报（医学版）》
CAS
2020年第4期8-15,2,共9页
Journal of Logistics University of PAP（Medical Sciences）
关键词
氯化钴
红景天苷
缺氧
稳定同位素标记细胞
定量蛋白组学
Cobalt chloride
Salidroside
Hypoxia
Stable isotope labeled cells
Quantitative proteomics
