Abstract: FR-PO1004

Klotho Enhances FoxO3-Mediated Manganese Superoxide Dismutase Expression by Negative Regulation of PI3K/AKT Pathway in Tacrolimus-Induced Oxidative Stress

Session Information

Category: Transplantation

  • 1701 Transplantation: Basic and Experimental

Authors

  • Lim, Sun Woo, The Catholic University of Korea, Seoul, Korea (the Republic of)
  • Shin, Yoo-Jin, The Catholic University of Korea, Seoul, Korea (the Republic of)
  • Luo, Kang, The Catholic University of Korea, Seoul, Korea (the Republic of)
  • Yang, Chul Woo, Seoul St. Mary's Hospital, Seoul, Korea (the Republic of)
Background

Mammalian forkhead members of the class O (FoxO) transcription factor are implicated in the regulation of oxidative stress, and FoxO proteins are negatively regulated by the phosphatidylinositol 3-kinase (PI3K)–AKT signaling pathway. We examined the effect of Klotho on PI3K/AKT pathway and manganese superoxide dismutase (MnSOD) in tacrolimus (Tac)-induced oxidative stress.

Methods

Mice were treated with Tac (1.5 mg/kg, subcutaneously) and recombinant Klotho (rKlotho, shedding form, 10 ug/kg once every 2 days, IP injection) for 4 weeks. For in vitro study, HK-2 cells were seeded in culture plates and treated with Tac (60 ug/mL) and rKlotho (1 ug/mL) with or without LY294003 (25 uM) for 12 h. We examined whether the oxidative stress and signaling pathway are involved in the protection by Klotho using immunoblot, immunostaining, RT-PCR, ChIP assay, oxygen consumption, ect.

Results

Klotho-treated mice showed decreased Tac-induced oxidative stress accompanied by functional and histological improvement. Klotho inhibited the PI3K/AKT-mediated phosphorylation of FoxO3a and enhanced FoxO3a binding to the MnSOD promoter. Klotho increased MnSOD mRNA and protein in mitochondria, and overexpressed MnSOD reduced Tac-induced toxicity in HK-2 cells. In mitochondria, Klotho improved Tac-induced mitochondrial dysfunction and decreased mitochondrial ROS, and this effect was enhanced by blocking PI3K activity with LY294002.

Conclusion

Collectively, Klotho protects Tac-induced oxidative stress by negative regulation of PI3K/AKT pathway, and subsequently enhances FoxO3a-mediated MnSOD expression. Through this mechanism, Klotho may protect against Tac-induced oxidative damage and apoptotic cell death. Finally, our results suggested that Klotho protein or Klotho-enhancing compounds may provide treatment options for nephrotoxicity in the future.

Klotho preserves Tac-induced mitochondrial dysfunction in HK-2 cells.