原理:急性肾损伤(AKI)具有相当高的死亡率和发病率,再加上缺乏有效的治疗选择。AKI通常转变为慢性肾病(CKD)并最终导致终末期肾衰竭。在缺血再灌注损伤(IRI)小鼠的肾脏中,干扰素刺激的基因15(ISG15)水平上调,顺铂,或单侧输尿管梗阻(UUO),然而,其在AKI发展和随后的AKI至CKD转变中的作用尚不清楚.方法:用双侧或单侧IRI攻击的Isg15敲除(Isg15KO)小鼠,顺铂,或UUO用于研究其在AKI中的作用。我们建立了ISG15过表达或敲除的细胞模型,并对其进行缺氧-复氧,顺铂,或转化生长因子-β1(TGF-β1)刺激。从AKI模型获得的肾脏RNA-seq数据来自公共数据库和我们的研究,用于检测ISG15及其相关基因的表达谱。此外,分析了来自人肾脏同种异体移植活检和小鼠IRI模型的已发表的单细胞RNA-seq数据,以研究ISG15和I型TGF-β受体(TGFβR1)的表达模式。西方印迹,qPCR,免疫共沉淀,我们进行了免疫组织化学染色试验以验证我们的发现.结果:Isg15KO小鼠IRI-,顺铂-,或UUO诱导的AKI和随后的AKI到CKD转变。在缺氧-复氧中,顺铂或TGF-β1处理的HK-2细胞,敲除ISG15减少刺激诱导的细胞纤维化,而具有修饰能力的ISG15的过表达加剧了细胞纤维化。免疫沉淀实验表明,ISG15促进TGFβR1的ISG化,并抑制其泛素化。此外,TGFβR1基因敲除阻断ISG15对HK-2细胞的纤维化加重作用,而TGFβR1的过度表达在IRI诱导的肾损伤期间消除了ISG15敲除的肾保护作用。结论:ISG15通过促进TGFβR1的ISG化,在AKI的发生和随后的AKI至CKD的转化中起重要作用。
Rationale: Acute kidney injury (AKI) has substantial rates of mortality and morbidity, coupled with an absence of efficacious treatment options. AKI commonly transits into chronic kidney disease (CKD) and ultimately culminates in end-stage renal failure. The interferon-stimulated gene 15 (ISG15) level was upregulated in the kidneys of mice injured by ischemia-reperfusion injury (IRI), cisplatin, or unilateral ureteral obstruction (UUO), however, its role in AKI development and subsequent AKI-to-CKD transition remains unknown. Methods:
Isg15 knockout (
Isg15 KO) mice challenged with bilateral or unilateral IRI, cisplatin, or UUO were used to investigate its role in AKI. We established cellular models with overexpression or knockout of ISG15 and subjected them to hypoxia-reoxygenation, cisplatin, or transforming growth factor- β1 (TGF-β1) stimulation. Renal RNA-seq data obtained from AKI models sourced from public databases and our studies, were utilized to examine the expression profiles of
ISG15 and its associated genes. Additionally, published single cell RNA-seq data from human kidney allograft biopsies and mouse IRI model were analyzed to investigate the expression patterns of ISG15 and the type I TGF-β receptor (TGFβR1). Western blotting, qPCR, co-immunoprecipitation, and immunohistochemical staining assays were performed to validate our findings. Results: Alleviated pathological injury and renal function were observed in
Isg15 KO mice with IRI-, cisplatin-, or UUO-induced AKI and the following AKI-to-CKD transition. In hypoxia-reoxygenation, cisplatin or TGF-β1 treated HK-2 cells, knockout ISG15 reduced stimulus-induced cell fibrosis, while overexpression of
ISG15 with modification capacity exacerbated cell fibrosis. Immunoprecipitation assays demonstrated that ISG15 promoted ISGylation of TGFβR1, and inhibited its ubiquitination. Moreover, knockout of TGFβR1 blocked ISG15\'s fibrosis-exacerbating effect in HK-2 cells, while overexpression of TGFβR1 abolished the renal protective effect of
ISG15 knockout during IRI-induced kidney injury. Conclusions: ISG15 plays an important role in the development of AKI and subsequent AKI-to-CKD transition by promoting TGFβR1 ISGylation.