Division of Nephrology, Shaoxing Peoples' Hospital, Shaoxing, China.
Department of Clinical Laboratory, University of Fukui Hospital, 23-3 Matsuoka-shimoaizuki, Eiheiji-cho, Yoshida, Fukui, 910-1193, Japan.
Clin Exp Nephrol. 2021 Jun;25(6):598-607. doi: 10.1007/s10157-021-02039-2. Epub 2021 Mar 1.
Cisplatin-induced injury of renal proximal tubular cells results basically from increased apoptosis via mitochondrial damage, and is mitigated by appropriate enhancement of autophagy. Peroxisome proliferator-activated receptor-delta (PPAR-δ) reportedly protects against not only mitochondrial damages but also enhances autophagy. Thus, PPAR-δ may protect against cisplatin-induced kidney injury.
We examined the protective effects of PPAR-δ activation on cisplatin-induced cellular injury and their detailed mechanisms in a murine renal proximal tubular (mProx) cell line using GW0742, an authentic PPAR-δ activator. Cisplatin-induced cell damages were evaluated by TUNEL assay and immunoblot analyses for p53, 14-3-3, Bax, Bcl2, cytochrome C, and activated caspases. Autophagy status was examined by immunoblot analyses for p62 and LC3.
GW0742 suppressed cisplatin-induced apoptosis of mProx cells by reducing the activation of caspase-3 via attenuating the phosphorylation of p53 and 14-3-3, mitochondrial Bax accumulation, cytochrome C release from mitochondria to the cytosol and ensuing cytosolic caspase-9 activation. In contrast, GW0742 did not diminish cisplatin-enhanced activation of caspases-8 or -12 as extrinsic or endothelium reticulum apoptotic pathways, respectively. The inhibitory effect of GW0742 on cisplatin-induced caspase-3 activation was significantly diminished by silencing of the PPAR-δ gene expression. GW0742 itself had no influence on starvation-stimulated or cisplatin-induced autophagy in mProx cells, suggesting that the protective effects were not mediated by autophagy modification.
Our results indicate that GW0742 may serve as a candidate agent to mitigate cisplatin nephrotoxicity via inhibiting the mitochondrial apoptotic pathway considerably depending on PPAR-δ, without modulating autophagy.
顺铂诱导的肾近端小管细胞损伤主要通过线粒体损伤导致细胞凋亡增加，而适当增强自噬可以减轻这种损伤。过氧化物酶体增殖物激活受体-δ（PPAR-δ）不仅可以保护线粒体免受损伤，还可以增强自噬。因此，PPAR-δ 可能对顺铂诱导的肾损伤具有保护作用。
我们使用过氧化物酶体增殖物激活受体-δ 的激活剂 GW0742，在鼠肾近端小管（mProx）细胞系中检测 PPAR-δ 激活对顺铂诱导的细胞损伤的保护作用及其详细机制。通过 TUNEL 检测和免疫印迹分析 p53、14-3-3、Bax、Bcl2、细胞色素 C 和活化的半胱天冬酶评估顺铂诱导的细胞损伤。通过免疫印迹分析 p62 和 LC3 评估自噬状态。
GW0742 通过减少 p53 和 14-3-3 的磷酸化、抑制线粒体 Bax 积累、抑制细胞色素 C 从线粒体释放到细胞质以及随后抑制细胞质中 caspase-9 的激活，抑制 caspase-3 的活化，从而抑制顺铂诱导的 mProx 细胞凋亡。相反，GW0742 并未减少顺铂增强的 caspase-8 或 -12 的激活，分别作为外在或内质网凋亡途径。GW0742 对顺铂诱导的 caspase-3 活化的抑制作用在沉默 PPAR-δ 基因表达后显著减弱。GW0742 本身对饥饿刺激或顺铂诱导的 mProx 细胞自噬没有影响，表明保护作用不是通过自噬修饰介导的。
我们的研究结果表明，GW0742 可能通过抑制线粒体凋亡途径，在很大程度上依赖于 PPAR-δ，而不调节自噬，成为减轻顺铂肾毒性的候选药物。
