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HIF-1?-BNIP3-mediated mitophagy in tubular cells protects against renal ischemia/reperfusion injury.

ABSTRACT: In the present study, we hypothesized that hypoxia-inducible factor 1? (HIF-1?)-mediated mitophagy plays a protective role in ischemia/reperfusion (I/R)-induced acute kidney injury (AKI). Mitophagy was evaluated by measuring the changes of mitophagy flux, mitochondria DNA copy number, and the changes of mitophagy-related proteins including translocase of outer mitochondrial membrane 20 (TOMM20), cytochrome c oxidase IV (COX IV), microtubule-associated protein 1 light chain 3B (LC3B), and mitochondria adaptor nucleoporin p62 in HK2 cells, a human tubular cell line. Results show that HIF-1? knockout significantly attenuated hypoxia/reoxygenation (H/R)-induced mitophagy, aggravated H/R-induced apoptosis, and increased the production of reactive oxygen species (ROS). Similarly, H/R induced significantly increase in Bcl-2 19-kDa interacting protein 3 (BNIP3), a downstream regulator of HIF-1?. Notably, BNIP3 overexpression reversed the inhibitory effect of HIF-1? knockout on H/R-induced mitophagy, and prevented the enhancing effect of HIF-1? knockout on H/R-induced apoptosis and ROS production. For in vivo study, we established HIF-1?^flox/flox; cadherin-16-cre mice in which tubular HIF-1? was specifically knockout. It was found that tubular HIF-1? knockout significantly inhibited I/R-induced mitophagy, and aggravated I/R-induced tubular apoptosis and kidney damage. In contrast, adenovirus-mediated BNIP3 overexpression significantly reversed the decreased mitophagy, and prevented enhanced kidney damage in tubular HIF-1? knockout mice with I/R injury. In summary, our study demonstrated that HIF-1?-BNIP3-mediated mitophagy in tubular cells plays a protective role through inhibition of apoptosis and ROS production in acute kidney damage.

SUBMITTER: Fu ZJ

PROVIDER: S-EPMC7452120 | biostudies-literature | 2020 Sep

REPOSITORIES: biostudies-literature

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HIF-1α-BNIP3-mediated mitophagy in tubular cells protects against renal ischemia/reperfusion injury.

Fu Zong-Jie ZJ Wang Zhi-Yu ZY Xu Lian L Chen Xiao-Hui XH Li Xiang-Xiao XX Liao Wei-Tang WT Ma Hong-Kun HK Jiang Meng-Di MD Xu Ting-Ting TT Xu Jing J Shen Yan Y Song Bei B Gao Ping-Jin PJ Han Wei-Qing WQ Zhang Wen W

Redox biology 20200807

In the present study, we hypothesized that hypoxia-inducible factor 1α (HIF-1α)-mediated mitophagy plays a protective role in ischemia/reperfusion (I/R)-induced acute kidney injury (AKI). Mitophagy was evaluated by measuring the changes of mitophagy flux, mitochondria DNA copy number, and the changes of mitophagy-related proteins including translocase of outer mitochondrial membrane 20 (TOMM20), cytochrome c oxidase IV (COX IV), microtubule-associated protein 1 light chain 3B (LC3B), and mitocho ...[more]

PMID: 32829253

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Project description:BackgroundThe morbidity of nephrolithiasis is 2-3 times higher in males than in females, suggesting that androgen plays a key role in nephrolithiasis. The death of renal tubular epithelial cells (TECs) is an important pathophysiological process contributing to the development of nephrolithiasis. Therefore, the aim of this study is to investigate whether androgen directly induces TECs apoptosis and necrosis and its underlying mechanisms in kidney stone formation.Materials and methodsWe compared serum testosterone level between male and female healthy volunteers and kidney stone patients. The in vivo nephrolithiasis model was established using glyoxylic acid, and calcium deposits were detected by van Kossa staining. In the in vitro study using mouse TECs (TCMK-1 cells) and human TECs (HK-2 cells), apoptosis, necrosis, and the expression of BH3-only protein Bcl-2-like 19 kDa-interacting protein 3 (BNIP3) were examined incubated with different doses of testosterone using flow cytometry. Levels of apoptosis-related proteins transfected with the BNIP3 siRNA were examined by western blotting. The mitochondrial potential (ΔΨm) was detected by JC-1 staining and flow cytometry. We monitored BNIP3 expression in the testosterone-induced TECs injury model after treatment with hypoxia inducible factor 1α (HIF-1α) and/or hypoxia inducible factor 2α (HIF-2α) inhibitors to determine the upstream protein regulating BNIP3 expression. Additionally, ChIP and luciferase assays were performed to confirm the interaction between HIF-1α and BNIP3.ResultsBoth male and female patients have significantly higher testosterones compared with healthy volunteers. More calcium deposits in the medulla were detected in male mice compared to female and castrated male mice. Testosterone induced TECs apoptosis and necrosis and increased BNIP3 expression in a dose-dependent manner. Testosterone also increased Bax expression, decreased Bcl-2 expression and induced a loss of ΔΨm. This effect was reversed by BNIP3 knockdown. HIF-1α inhibition significantly decreased BNIP3 expression and protected TECs from testosterone-induced apoptosis and necrosis. HIF-2α inhibition, however, did not influence BNIP3 expression or TECs apoptosis or necrosis. Finally, HIF-1α interacted with the BNIP3 promoter region.ConclusionBased on these results, testosterone induced renal TECs death by activating the HIF-1α/BNIP3 pathway.

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HIF-1?-BNIP3-mediated mitophagy in tubular cells protects against renal ischemia/reperfusion injury.

Publications

HIF-1α-BNIP3-mediated mitophagy in tubular cells protects against renal ischemia/reperfusion injury.

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