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Abstract: PO2002

Analysis of the Relationship Between Proteasome and Autophagy in Podocytes Using Podocyte-Specific Proteasome Impairment Mice

Session Information

  • Podocyte Biology
    October 22, 2020 | Location: On-Demand
    Abstract Time: 10:00 AM - 12:00 PM

Category: Glomerular Diseases

  • 1204 Podocyte Biology

Authors

  • Makino, Shinichi, Chiba Daigaku Daigakuin Igaku Kenkyuin Igakubu, Chiba, Chiba, Japan
  • Yamada, Hiroyuki, Chiba Daigaku Daigakuin Igaku Kenkyuin Igakubu, Chiba, Chiba, Japan
  • Kadariswantiningsih, Ika N., Chiba Daigaku Daigakuin Igaku Kenkyuin Igakubu, Chiba, Chiba, Japan
  • Empitu, Maulana A., Chiba Daigaku Daigakuin Igaku Kenkyuin Igakubu, Chiba, Chiba, Japan
  • Yanagita, Motoko, Kyoto Daigaku Daigakuin Igaku Kenkyuka Igakubu, Kyoto, Japan
  • Asanuma, Katsuhiko, Chiba Daigaku Daigakuin Igaku Kenkyuin Igakubu, Chiba, Chiba, Japan
Background

Ubiquitin-proteasome system and autophagy-lysosome system are major intracellular protein degradation mechanism. The relationship of these systems in podocyte has not well been understood.

Methods

In this study, we generated podocyte-specific proteasome impaired mice (Rpt3pdKO) by deletion of Rpt3, which is essential for construction of 26S proteasome, using Cre-loxP system. Albuminuria and number of sclerotic glomeruli increased in the Rpt3pdKO mice compared with Rpt3control mice. Oxidative stress and podocytes apoptosis were related to podocyte injury. To evaluated autophagic activity, LC3 dots in podocytes were evaluated after administration of chloroquine, inhibitor of autophagic flux. In vitro experiment, cultured podocytes were treated with bortezomib (BTZ), proteasome inhibitor, which lead to podocyte apoptosis. The expression of LC3 was evaluated in podocytes after administration of bortezomib in the presence of E64d/pepstatinA, inhibitor of autophagic flux.

Results

In vivo, after administration of chloroquine, LC3 dots decreased in podocytes of Rpt3pdKO mice compared with Rpt3control mice. In vitro, the expression of LC3 decreased and the accumulation of p62 increased in cultured podocytes after treatment of BTZ in the presence of E64d/pepstatinA. These results indicated autophagic activity was suppressed in podocytes with proteasome impairment. pULK1, which is a down stream of mTOR signal, was phosphorylated in podocytes with proteasome impairment, suggested that suppression of autophagic activity was associated with mTOR activation. Pre-treatment of rapamycin, inhibitor of mTOR, ameliorated podocyte apoptosis induced by BTZ in vitro. In vivo, the number of sclerotic glomeruli decreased in Rpt3pdKO mice with rapamycin administration compared with in Rpt3pdKO mice without rapamycin administration.

Conclusion

Impairment of proteasome suppressed autophagic activity associated with mTOR activation in podocytes. Activation of autophagy have the protective effect on podocyte injury due to proteasome impairment.