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Abstract: TH-PO472

Ferroptosis Is Activated by TGF-β/Smad3-Driven Renal Fibrosis Both In Vivo and In Vitro

Session Information

  • CKD: Mechanisms - I
    November 07, 2019 | Location: Exhibit Hall, Walter E. Washington Convention Center
    Abstract Time: 10:00 AM - 12:00 PM

Category: CKD (Non-Dialysis)

  • 2103 CKD (Non-Dialysis): Mechanisms

Authors

  • Li, Guisen, Sichuan Provincial People's Hospital, Chengdu, China
  • Zhong, Xiang, Department of Nephrology, Sichuan Academy of Medical Sciences and Sichuan Provincial People's Hospital, Chengdu, China
  • Yang, Xue, Sichuan Academy of Medical Science and Sichuan Provincial People's Hospital, Chengdu, China
  • Wang, Huan, Sichuan Academy of Sciences & Sichuan Provincial People's Hospital, Chengdu, China
  • Li, Yi, Sichuan Academy of Sciences & Sichuan Provincial People's Hospital, School of Medicine, University of Electronic Science and Technology of China, Chengdu, China
  • Wang, Li, Sichuan Academy of Sciences & Sichuan Provincial People's Hospital, School of Medicine, University of Electronic Science and Technology of China, Chengdu, China
Background

Ferroptosis is a novel non-programmed cell death driven by iron-based lipid peroxidation and its role in renal fibrosis is unknown. In this study, we hypothesized that ferroptosis is activated by TGF-β/Smad3-drivend renal fibrosis both in vivo and in vitro.

Methods

The morphology of ferroptosis and expression of ferroptosis indexes such as glutathione peroxidase 4(GPX4) and solute carrier family 7 member 11(SLC7A11) were detected both in TGF-β-stimulated rat tubular epithelial cells(rTEC) and unilateral ureteral obstruction(UUO) mouse model by electron microscopy(EM), IHC, western blot(WB) and quantitative real-time polymerase chain reaction(qRT-PCR). In addition, the ferroptosis was induced or inhibited by ferroptosis specifically agonist Erastin and inhibitor Ferrostatin in rTEC. Renal fibrosis markers of fibronectin(FN), collagen I(Col I) and α-smooth muscle actin(α-SMA) were detected by WB and qRT-PCR. Moreover, GXP4 was observed in SIS3 specifically inhibited Smad3 rTEC and Smad3 conditional knock out (KO) UUO mice by WB or IHC. The co-expression of P-Smad3 and GPX4 were detected by confocal and CO-IP in TGF-β-stimulated renal fibrosis in rTEC.

Results

The ferroptosis morphological changes including the shrucked volume of mitochondria, a reduced number of cristae and an increased density of bilayer membrane were observed in TGF-β-stimulated rTEC and renal tissue of UUO by EM. The expression of ferroptosis negatively indexes of GPX4 and SLC7A11 was significantly decreased in renal fibrosis both in vivo and in vitro . After ferroptosis induced by Erastin, the renal fibrosis markers of FN, Col I and α-SMA were significantly enhanced after TGF-β treatment for 24 hours, while renal fibrosis was decreased by Ferrostatin. In SIS3 specifically inhibited Smad3 rTEC and Smad3 KO UUO mice, GPX4 expression was significantly up-regulated. Confocal and CO-IP results suggested that the P-Smad3 and GPX4 was co-expressed in rTEC and physically combined with each other after TGF-β1 stimulated for 24h.

Conclusion

Ferroptosis is expressed in renal fibrosis both in vivo and in vitro. The intervention of ferroptosis by agonist and inhibitor has effect on the renal fibrosis in TECs. GPX4-mediated ferroptosis is dependent on TGF-β/Smad3 signaling in renal fibrosis. These data indicates that the potential role and mechanism of ferroptosis in TGF-β/Smad3-driven renal fibrosis.

Funding

  • Government Support - Non-U.S.