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HIF stabilization inhibits renal epithelial cell migration and is associated with cytoskeletal alterations.

ABSTRACT: Acute kidney injury (AKI) is a common and potentially lethal complication in the hospitalized patients, with hypoxic injury being as a major cause. The loss of renal tubular epithelial cells (TEC), one of the AKI hallmarks, is potentially followed by tubular regeneration process orchestrated by the remaining uninjured TECs that undergo proliferation and migration. In this study, we used human primary TEC to investigate the initiation of tubular cell migration and associated cytoskeletal alterations in response to pharmacological HIF stabilization which resembles the pathophysiology of hypoxia. Tubular cells have been shown to migrate as cohorts in a wound healing assay. Importantly, cells of distal tubular origin moved faster than those of proximal origin. HIF stabilization impaired TEC migration, which was confirmed by live single cell tracking. HIF stabilization significantly reduced tubular cell migration velocity and promoted cell spreading. In contrast to the control conditions, HIF stabilization induced actin filaments rearrangement and cell adhesion molecules including paxillin and focal adhesion kinase. Condensed bundling of keratin fibers was also observed, while the expression of different types of keratins, phosphorylation of keratin 18, and the microtubule structure were not altered. In summary, HIF stabilization reduced the ability of renal tubular cells to migrate and led to cytoskeleton reorganization. Our data suggested an important involvement of HIF stabilization during the epithelial migration underlying the mechanism of renal regeneration in response to AKI.

SUBMITTER: Muller S

PROVIDER: S-EPMC6015081 | biostudies-other | 2018 Jun

REPOSITORIES: biostudies-other

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HIF stabilization inhibits renal epithelial cell migration and is associated with cytoskeletal alterations.

Müller Simon S Djudjaj Sonja S Lange Janina J Iacovescu Mihail M Goppelt-Struebe Margarete M Boor Peter P

Scientific reports 20180622 1

Acute kidney injury (AKI) is a common and potentially lethal complication in the hospitalized patients, with hypoxic injury being as a major cause. The loss of renal tubular epithelial cells (TEC), one of the AKI hallmarks, is potentially followed by tubular regeneration process orchestrated by the remaining uninjured TECs that undergo proliferation and migration. In this study, we used human primary TEC to investigate the initiation of tubular cell migration and associated cytoskeletal alterati ...[more]

PMID: 29934555

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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 stabilization inhibits renal epithelial cell migration and is associated with cytoskeletal alterations.

Publications

HIF stabilization inhibits renal epithelial cell migration and is associated with cytoskeletal alterations.

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