基于p62/Keap1/Nrf2介导自噬-铁死亡crosstalk探讨益母草碱对肾缺血再灌注损伤的保护作用及机制研究

肾小管上皮细胞死亡在肾缺血再灌注损伤过程中处于中心地位。该损伤过程引起了肾小管上皮细胞的自噬与铁死亡，且二者相互调节、交叉串话，共同决定肾小管上皮细胞的结局。新近研究和我们前期实验结果提示，p62依赖性选择性自噬在该应激反应途径中产生Nrf2扩增的反馈环，p62/Keap1/Nrf2可能是肾缺血再灌注损伤中自噬-铁死亡crosstalk的关键机制；活血利水中药益母草的有效成分益母草碱能有效改善肾脏病理损伤，保护糖氧剥夺/复糖复氧模型下的肾小管上皮细胞，减轻脂质过氧化，亦能抑制自噬，但其机制尚不明确。本项目拟结合体内外实验，从动物、细胞、分子三个层面，通过慢病毒感染、电镜、激光共聚焦、免疫共沉淀等方法，阐明p62/Keap1/Nrf2介导自噬-铁死亡crosstalk的具体分子机制，并以此为切入点解析益母草碱保护肾缺血再灌注损伤的途径和作用靶点，为临床防治肾缺血再灌注损伤提供新思路和新方法。

Renal tubular epithelial cell death plays a central role in the pathophysiological process of renal ischemia reperfusion injury. The process of injury causes autophagy and ferroptosis of renal tubular epithelial cells, and the two interact and crosstalk with each other, which together determine the outcome of cell death or survival. Recent findings and our previous research indicate that, p62-dependent selective autophagy creates a feedback loop for Nrf2 amplification in stress response pathways by promoting Keap1 degradation, and p62/Keap1/Nrf2 may probably serve as the key signaling pathway mediating the crosstalk between autophagy and ferroptosis. Our preliminary study has also demonstrated that, leonurine, a major active component in Motherwort, exerts renal protective roles by alleviating histological damages, attenuating oxidative stress and inhibiting autophagy, as well as protects renal tubular epithelial cells under glucose oxygen deprivation/recovery (OGD/R) stress. However, the underlying mechanisms involved have not been elucidated and deserve to be addressed. Therefore, in this project, by in vivo and in vitro experiments, we try to clarify the mechanism of p62/Keap1/Nrf2 in regulating autophagy-ferroptosis crosstalk and pharmacological pathway and targets of leonurine. We will use a series of experimental methods including lentivirus infection, electron microscopy, confocal laser scanning and immunocoprecipitation, to explore the mechanism at animal, cell and molecular levels. Our research will provide new methods and ideas for the treatment of renal ischemia reperfusion injury in clinic.