姜黄素活化肝细胞Nrf2抗氧化系统效应及缓解胰岛素抵抗作用机制研究

Studies on curcumin-induced activation of the Nrf2 antioxidant system in hepatocytes and the underlying mechanisms on attenuating insulin resistance

目的探讨姜黄素对HepG2细胞Nrf2信号功能的调控作用与其改善胰岛素抵抗的效应关系,并进一步研究高脂饮食诱导小鼠胰岛素抵抗时姜黄素干预对肝脏Nrf2系统功能及糖异生作用的影响。方法在人肝癌细胞HepG2上分别用葡萄糖氧化酶(GO)或长期胰岛素处理,分别制成氧化应激和高胰岛素血症胰岛素抵抗模型;用蛋白印迹方法(WB)检测姜黄素对Nrf2抗氧化系统的作用。另外,对细胞进行短时胰岛素刺激,观察胰岛素信号(PKBSer473磷酸化水平)变化。为研究姜黄素的整体动物药物作用,在高脂诱导的胰岛素抵抗小鼠模型用姜黄素进行干预(雄性C57BL/6J小鼠,给予高脂饲料加上3%的姜黄素),在第25周进行丙酮酸耐量实验,并于第27周取肝脏组织检测Nrf2系统功能变化。结果姜黄素可明显激活HepG2细胞Nrf2系统,并对抗GO氧化应激所致的PKB磷酸化损害。在高脂饲喂小鼠胰岛素抵抗模型,姜黄素可改善丙酮酸耐量,提示其增强胰岛素作用(抑制肝脏糖异生),从而缓解肝脏糖代谢异常;姜黄素还明显激活高脂肥胖小鼠肝脏受抑制的Nrf2信号功能。结论姜黄素通过增强内源性Nrf2系统功能对抗氧化应激,是其缓解肝细胞胰岛素抵抗的重要药理机理。

Objective To examine the regulatory effect of curcumin on the nuclear factor erythroid-derived 2-like 2 （Nrf2） system in hepatocytes and to further explore the effect of curcumin on the intervention of hepatic Nrf2 function and its impact on gluconeogeuesis in the high fat diet-induced insulin resistance mouse model. Methods The models of oxidative stress and hyperinsulinemia-induced insulin resistance were established by treating HepG2 cells with glucose oxidase （GO） or long-term insulin treatment, respectively. The effect of curcumin on Nrf2 anti-oxidative function was detected by utilizing Western blotting method. In addition, a short term stimulation with insulin was performed to investigate the alterations of insulin signaling （PKB- Ser473 phosphorylation levels）. To study the in vivo pharmacological effects of curcumin, high fat diet （HFD）-induced insulin resistance male C57BL/6J mice were treated with 3% curcumin in diet, and pyruvate tolerance test was carried out at the 25~ week following the intervention. After 27 weeks, the mice were euthanized and the livers were isolated for WB analysis of the functional changes of the Nrf＇2 system. Results Curcumin could significantly activate the Nrf2 system in HepG2 cells. Curcumin also reduced PKB Ser473 phosphorylation caused by GO and oxidative stress. In HFD-induced insulin resistant mice, curcumin could improve pymvate tolerance, suggesting that curcumin suppressed hepatic gluconeogenesis and attenuated the abnormality in hepatic glucose metabolism. Curcumin also markedly activated the repressed Nrf2 system in the livers of HFD- induced obese mice. Conclusion Curcumin could enhance the function of endogenous Nrf2 system to against oxidative stress and attenuate the insulin resistance.