title: Adrenomedullin protects against oxidative stress-induced podocyte injury as an endogenous antioxidant.
abstract: BACKGROUND
  We previously reported that puromycin aminonucleoside (PAN) increased adrenomedullin (AM) secretion and AM mRNA expression in podocytes, through overproduction of oxidative stress. To clarify the cytoprotective role of AM as antioxidative and antiapoptotic substance in podocytes, we investigated the effect of exogenous AM and AM antagonist on PAN-induced apoptosis in conditionally immortalized murine podocytes.
  
  
  METHODS
  The expression of AM, RAMP 2 and RAMP 3 was investigated using real-time PCR, western blotting analysis and immunofluorescence microscopy. Reactive oxygen species (ROS) production was measured by CM-H(2)DCFDA fluorescence intensity method. The percentage of apoptotic cells was measured by Hoechst 33342 staining.
  
  
  RESULTS
  PAN (100 microg/ml) significantly (P < 0.01) increased ROS production, associated with an increase in apoptosis; the percentage of apoptotic cells is 5.3% + 0.05% (P < 0.01) with 36 h treatment of PAN compared to 0.24 + 0.16% with no treatment. Several antioxidants could markedly reduce PAN-induced apoptosis in cultured podocytes, suggesting that PAN-induced apoptosis might be attributable to the overproduction of ROS. Accordingly, the administration of exogenous AM (10(-6) M) could significantly reduce not only ROS production via a PKA-dependent pathway, but also the resultant apoptosis induced by PAN. AM antagonists, CGRP8-37, augmented PAN-induced apoptosis, associated with increased ROS production, 2.2- and 2.3-Fold, respectively. RAMP 2 and RAMP 3 could be detected in podocytes and glomeruli.
  
  
  CONCLUSIONS
  This suggests that ROS-induced up-regulation of AM with PAN could counteract ROS-induced apoptosis, by the suppression of ROS production. Therefore, AM might have the endogenous antioxidant potential to protect against ROS-induced podocyte injury.
authors.affiliation: Department of Nephrology and Endocrinology, University of Tokyo Graduate School of Medicine, Hongo, Bunkyo-ku, Tokyo 113-8655, Japan.
authors.forename: Shigeyoshi
authors.initials: S
authors.lastname: Oba
authors.forename: Masayo
authors.initials: M
authors.lastname: Hino
authors.forename: Toshiro
authors.initials: T
authors.lastname: Fujita
mesh_terms: D053607:Adrenomedullin / Q000627:therapeutic use*; D000818:Animals; D000975:Antioxidants / Q000627:therapeutic use*; D002478:Cells, Cultured; D051379:Mice; D018384:Oxidative Stress / Q000187:drug effects*; D050199:Podocytes / Q000187:drug effects*; D011692:Puromycin Aminonucleoside / Q000494:pharmacology
chemical_list: D000975:Antioxidants; D053607:Adrenomedullin; D011692:Puromycin Aminonucleoside