Abstract：
N6-methyladenosine (m6A) and MELLT3 assume a role in the development of acute kidney injury (AKI). However, their mechanism in AKI remains under-explored. On this basis, this study explored the mechanism of MELLT3 in mitochondrial damage and ferroptosis of kidney tubular epithelial cells after AKI. HK-2 cells were induced by lipopolysaccharide (LPS) to simulate AKI, followed by gain and loss of function of genes, detection of mitochondrial damage and ferroptosis indicators, and analysis of gene interactions. An AKI mouse model was developed using the cecal ligation and puncture (CLP) method to investigate the effect of METTL3 knockdown on kidney injury. MDM2 and LMNB1 were upregulated and p53 was downregulated in LPS-treated HK-2 cells. Mechanistically, the E3 ubiquitin ligase MDM2 increased p53 ubiquitination to activate LMNB1. METTL3 knockdown decreased m6A methylation of MDM2, thus diminishing YTHDF1-mediated MDM2 mRNA stability and translation in LPS-treated HK-2 cells. Knockdown of LMNB1, MDM2, or METTL3 reduced NO, MDA, iron ion, and ROS levels as well as mitochondrial damage and raised SOD, GSH, XCT, GPX4, FPN1, and TFR1 levels in LPS-treated HK-2 cells. The in vivo results showed that METTL3 knockdown reduced renal injury and ferroptosis in CLP mice. METTL3 knockdown prevents mitochondrial damage and ferroptosis of kidney tubular epithelial cells after AKI via the MDM2-p53-LMNB1 axis.
摘要：
N6-甲基腺苷（m6A）和MELLT3在急性肾损伤（AKI）的发展中起作用。然而,它们在AKI中的机制仍未被探索。在此基础上,本研究探讨MELLT3在AKI后肾小管上皮细胞线粒体损伤和铁凋亡中的作用机制。用脂多糖(LPS)诱导HK-2细胞模拟AKI,其次是基因功能的获得和丧失，线粒体损伤和铁死亡指标的检测，和基因相互作用的分析。使用盲肠结扎和穿孔(CLP)方法开发AKI小鼠模型以研究METTL3敲低对肾损伤的影响。在LPS处理的HK-2细胞中，MDM2和LMNB1上调，p53下调。机械上,E3泛素连接酶MDM2增加p53泛素化以激活LMNB1。METTL3敲低降低了MDM2的m6A甲基化，从而降低了LPS处理的HK-2细胞中YTHDF1介导的MDM2mRNA稳定性和翻译。敲除LMNB1、MDM2或METTL3可降低NO，MDA,铁离子，和ROS水平以及线粒体损伤和升高的SOD，GSH,XCT,LPS处理的HK-2细胞中的GPX4、FPN1和TFR1水平。体内结果显示METTL3敲低可降低CLP小鼠的肾损伤和铁凋亡。METTL3敲低通过MDM2-p53-LMNB1轴防止AKI后肾小管上皮细胞的线粒体损伤和铁凋亡。
