红景天苷上调EFNA1激活PI3K/AKT/Nrf2信号通路减轻顺铂引起的急性肾损伤

Salidroside attenuates cisplatin-induced acute kidney injury by activating PI3K/AKT/Nrf2 signaling pathway via upregulating EFNA1

摘要:

目的探究红景天苷(salidroside, SAL)在顺铂肾毒性中的保护机制。 方法体内外模型中确定SAL对顺铂诱导的急性肾损伤的保护作用，并通过Western印迹法及血清检测肾损伤标志物及凋亡相关蛋白的表达。流式细胞术检测细胞活性氧(reactive oxygen species, ROS)水平，透射电镜观察线粒体损伤。转录组测序分析差异表达基因，结合GO/KEGG富集筛选相关通路。利用siRNA敲低靶基因验证SAL通过上调EFNA1发挥保护中作用。检测PI3K和AKT的磷酸化水平及Nrf2核定位情况，评估抗氧化酶(SOD、GSH-px、CAT)活性变化。 结果SAL可以降低顺铂处理的HK2细胞及小鼠的肾损伤标志物水平，促进抗氧化酶的水平并抑制ROS，减少线粒体损伤和细胞凋亡。转录组测序结合WB结果显示SAL通过上调EFNA1促进PI3K和AKT磷酸化，介导下游蛋白Nrf2核转位，上调SOD、CAT、GSH-px等抗氧化酶水平，抑制细胞凋亡减轻顺铂诱导的急性肾损伤。 结论SAL通过上调EFNA1激活PI3K/AKT/Nrf2通路，抑制氧化应激，减轻顺铂诱导的急性肾损伤。

Abstract:

ObjectiveTo investigate the protective mechanism of salidroside(SAL) in cisplatin nephrotoxicity. MethodsThe protective effect of SAL against cisplatin-induced acute kidney injury was determined by in vitro and in vivo model, and the expression of renal injury markers and apoptosis-related proteins were detected by Western blotting assay and serum examination. Flow cytometry was used to detect apoptosis and reactive oxygen species(ROS), and transmission electron microscopy was used to observe mitochondrial damage. Transcriptome sequencing was used to analyze differentially expressed genes, and relevant pathways were screened by combining GO/KEGG enrichment; the siRNA was used to knock down the target gene to verify the protective role of SAL by upregulating EFNA1. In addition, the phosphorylation levels of phosphoinositide 3-kinase(PI3K) and protein kinase B(AKT) and the nuclear localization of Nrf2 were detected, and changes in the activities of antioxidant enzymes [superoxide dismutase(SOD), glutathione peroxidase(GSH-px), catalase(CAT)] were assessed. ResultsSAL reduced the levels of renal injury markers, promoted the levels of antioxidant enzymes and inhibited ROS, reduced mitochondrial damage and apoptosis in cisplatin-treated HK2 cells and mice. Transcriptome sequencing combined with WB results showed that SAL promoted the phosphorylation of PI3K and AKT through up-regulation of EFNA1, mediated the nuclear translocation of the downstream protein Nrf2, and ultimately elevated the levels of antioxidant enzymes, such as SOD, CAT, and GSH-px, and inhibited apoptosis to alleviate cisplatin-induced acute kidney injury. ConclusionSAL inhibits oxidative stress and attenuates cisplatin-induced acute kidney injury by up-regulating EFNA1 to activate the PI3K/AKT/Nrf2 pathway.