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Bilirubin activates transcription of HIF-1? in human proximal tubular cells cultured in the physiologic oxygen content.

ABSTRACT: The expression of hypoxia-inducible factor (HIF) is influenced by reactive oxygen species (ROS). Effect of bilirubin on HIF-1 expression in proximal tubular cells was investigated under physiological oxygen concentration, which is relative hypoxic condition mimicking oxygen content in the medulla of renal tissue. The human kidney (HK2) cells were cultured in 5% oxygen with or without bilirubin. HIF-1? protein expression was increased by bilirubin treatment at 0.01-0.2 mg/dL concentration. The messenger RNA expression of HIF-1? was increased by 1.69±0.05 folds in the cells cultured with 0.1 mg/dL bilirubin, compared to the control cells. The inhibitors of PI3K/mTOR, PI3K/AKT, and ERK 1/2 pathways did not attenuate increased HIF-1? expression by bilirubin. HIF-1? expression decreased by 10 µM exogenous hydrogen peroxide (H2O2); scavenger of ROS with or without bilirubin in the HK2 cells increased HIF-1? concentration more than that in the cells without bilirubin. Exogenous H2O2 decreased the phosphorylation of P70S6 kinase, which was completely reversed by bilirubin treatment. Knockdown of NOX4 gene by small interfering RNA (siRNA) increased HIF-1? mRNA expression. In coonclusion, bilirubin enhances HIF-1? transcription as well as the up-regulation of HIF-1? protein translation through the attenuation of ROS and subunits of NADPH oxidase.

SUBMITTER: Kim SG

PROVIDER: S-EPMC4194285 | biostudies-literature | 2014 Sep

REPOSITORIES: biostudies-literature

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Bilirubin activates transcription of HIF-1α in human proximal tubular cells cultured in the physiologic oxygen content.

Kim Sung Gyun SG Ahn Shin-Young SY Lee Eun Seong ES Kim Sejoong S Na Ki Young KY Chae Dong-Wan DW Chin Ho Jun HJ

Journal of Korean medical science 20140930

The expression of hypoxia-inducible factor (HIF) is influenced by reactive oxygen species (ROS). Effect of bilirubin on HIF-1 expression in proximal tubular cells was investigated under physiological oxygen concentration, which is relative hypoxic condition mimicking oxygen content in the medulla of renal tissue. The human kidney (HK2) cells were cultured in 5% oxygen with or without bilirubin. HIF-1α protein expression was increased by bilirubin treatment at 0.01-0.2 mg/dL concentration. The me ...[more]

PMID: 25317019

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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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Bilirubin activates transcription of HIF-1? in human proximal tubular cells cultured in the physiologic oxygen content.

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Bilirubin activates transcription of HIF-1α in human proximal tubular cells cultured in the physiologic oxygen content.

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