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Kidney Week

Abstract: SA-PO590

NADPH Oxidase 4 Mediates TGF-β1/Smad Signaling Pathway Induced AKI in Hypoxia

Session Information

Category: Acute Kidney Injury

  • 103 AKI: Mechanisms


  • Yoon, Se-Hee, Konyang University Hospital, Daejeon, Korea (the Republic of)
  • Cho, Jang-Hee, Kyungpook National University Hospital, Daegu, Korea (the Republic of)
  • Hwang, Won Min, Konyang University Hospital, Daejeon, Korea (the Republic of)
  • Yun, Sung-Ro, Konyang University Hospital, Daejeon, Korea (the Republic of)

Hypoxia is an important cause of acute kidney injury (AKI) in various conditions because kidneys are one of the most susceptible organ to hypoxia. Reactive oxygen species (ROS) plays an important role in hypoxia induced AKI. Among various sources of ROS, nicotinamide adenine dinucleotide 3-phosphate (NADPH) oxidase (Nox) is the major intracellular non- mitochondrial ROS source. Because among seven Nox families, Nox4 is the most abundant in the human kidney, changes of Nox4 expression in hypoxia are predicted to affect the progression of AKI by altering the intracellular ROS level in the kidney.
Based on these observations, we investigated the role of Nox4 and the benefits of Nox4 inhibition in hypoxia induced AKI.


The hypoxic injury induced in HK-2 cells by hypoxia chamber and CoCl2. Reverse transcription polymerase chain reaction for Nox4 and TGF-β1 was performed. Western blotting for Nox4 and Smad pathway were done. ROS production was detected using a DHE stain and Amplex red assay. HK-2 cells were transfected with siNox4 and pretreated with GKT137831(most specific Nox1/4 inhibitor). ELISA has been used to measure TGF-β1 levels. The effect of treatment with TGF-β1 type 1 tyrosine kinase inhibitor SB431542 on Nox4 expression was observed.


Expression of Nox4 in cultured human renal proximal tubular epithelial cells (HK-2) was significantly increased by hypoxic stimulation. TGF-β1 was endogenously secreted by hypoxic HK-2 cells. SB4315432 (a TGF-β1 receptor I inhibitor) significantly inhibited Nox4 expression in HK-2 cells through the Smad-dependent cell signaling pathway. Silencing of Nox4 using Nox4 siRNA and pharmacologic inhibition with GKT137831 reduced the production of ROS and attenuated the apoptotic pathway. In addition, knockdown of Nox4 increased cell survival in hypoxic HK-2 cells and pretreatment with GKT137831 reproduce these results.


This study demonstrates that hypoxia induces HK-2 cell apoptosis through a signaling pathway involving TGF-β1 via Smad pathway induction of Nox4-dependent ROS generation. Therapies targeting Nox4 may be effective against hypoxia-induced AKI.