To investigate the mechanism of renal ischemia-reperfusion injury (IRI) via the regulation of N6-methyl-adenosine (m6A) and relevant genes, IRI was induced in Sprague Dawley rats and the urine and serum creatinine levels and tissue structure changes were observed. m6A and METTL3 protein levels were assessed via dot blotting and western blotting, respectively. The hypoxia/reoxygenation (H/R) cell model was constructed using NRK-52E cells, and METTL3 protein levels were assessed. METTL3 was inhibited to observe its impact on NRK-52E cell apoptosis and m6A expression in H/R processes. Methylated RNA immunoprecipitation (MeRIP) sequencing was conducted, followed by MeRIP-qRT-PCR and qRT-PCR validation. Our results indicated that urine and serum creatinine levels increased and that renal injury and cell apoptosis were both observed in IRI model. In additon, m6A expression increased in the IRI model, and METTL3 protein levels significantly increased in the IRI and H/R models. When METTL3 was inhibited, the m6A levels were accordingly decreased and cell apoptosis was suppressed in the H/R in vitro model. Based on MeRIP sequencing, tfap2a, cyp1b1, and foxd1 were significantly differentially expressed, as was m6A, which is involved in the negative regulation of cell proliferation and kidney development. We confirmed that foxd1 mRNA and its methylation levels contributed to IRI and H/R.

中文翻译：

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METTL3通过调节Foxd1甲基化而导致肾脏缺血再灌注损伤。

为了通过调节N6-甲基腺苷（m6A）和相关基因来研究肾脏缺血再灌注损伤（IRI）的机制，在Sprague Dawley大鼠中诱导了IRI，观察了尿液和血清肌酐水平以及组织结构的变化。m6A和METTL3蛋白水平分别通过斑点印迹和蛋白质印迹进行评估。使用NRK-52E细胞构建缺氧/复氧（H / R）细胞模型，并评估METTL3蛋白水平。抑制METTL3观察其对N / 52过程中NRK-52E细胞凋亡和m6A表达的影响。进行甲基化的RNA免疫沉淀（MeRIP）测序，然后进行MeRIP-qRT-PCR和qRT-PCR验证。我们的结果表明，IRI模型中观察到尿液和血清肌酐水平升高，并且观察到肾损伤和细胞凋亡。另外，在IRI模型中m6A表达增加，并且在IRI和H / R模型中METTL3蛋白水平显着增加。当METTL3被抑制时，在H / R体外模型中，m6A水平相应降低，细胞凋亡受到抑制。根据MeRIP测序，tfap2a，cyp1b1和foxd1显着差异表达，而m6A也参与细胞增殖和肾脏发育的负调控。我们证实foxd1 mRNA及其甲基化水平有助于IRI和H / R。与foxd1和m6A明显差异表达，而m6A参与细胞增殖和肾脏发育的负调控。我们证实foxd1 mRNA及其甲基化水平有助于IRI和H / R。与foxd1和m6A明显差异表达，而m6A参与细胞增殖和肾脏发育的负调控。我们证实foxd1 mRNA及其甲基化水平有助于IRI和H / R。