title: Activation of local aldosterone system within podocytes is involved in apoptosis under diabetic conditions.
abstract: Previous studies have shown that mineralocorticoid receptor (MCR) blocker reduces proteinuria in diabetic nephropathy (DN), but the role of aldosterone in podocyte injury has never been explored in DN. This study was undertaken to elucidate whether a local aldosterone system existed in podocytes and to examine its role in podocyte apoptosis under diabetic conditions. In vitro, immortalized podocytes were exposed to 5.6 mM glucose (NG), NG + 24.4 mM mannitol, and 30 mM glucose (HG) with or without 10(-7) M spironolactone (SPR). In vivo, 32 Sprague-Dawley rats were injected with diluent (C, n = 16) or streptozotocin intraperitoneally [diabetes mellitus (DM), n = 16], and 8 rats from each group were treated with SPR for 3 mo. Aldosterone synthase (CYP11B2) and MCR mRNA and protein expression were determined by real-time PCR and Western blot, respectively, and aldosterone levels by radioimmunoassay. Western blot for apoptosis-related molecules, Hoechst 33342 staining, and terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling (TUNEL) assay were performed to determine apoptosis. CYP11B2 and MCR expression were significantly higher in HG-stimulated podocytes and DM glomeruli compared with NG cells and C glomeruli, respectively, along with increased aldosterone levels. Western blot analysis revealed that cleaved caspase-3 and Bax expression was significantly increased, whereas Bcl-2 expression was significantly decreased in HG-stimulated podocytes and in DM glomeruli. Apoptosis determined by Hoechst 33342 staining and TUNEL assay were also significantly increased in podocytes under diabetic conditions. These changes in the expression of apoptosis-related proteins and the increase in apoptotic cells were inhibited by SPR treatment. These findings suggest that a local aldosterone system is activated and is involved in podocyte apoptosis under diabetic conditions.
authors.affiliation: Department of Internal Medicine, College of Medicine, Brain Korea 21 for Medical Science, Yonsei University, Seoul, Korea.
authors.forename: Sun Ha
authors.initials: SH
authors.lastname: Lee
authors.forename: Tae-Hyun
authors.initials: TH
authors.lastname: Yoo
authors.forename: Bo-Young
authors.initials: BY
authors.lastname: Nam
authors.forename: Dong Ki
authors.initials: DK
authors.lastname: Kim
authors.forename: Jin Ji
authors.initials: JJ
authors.lastname: Li
authors.forename: Dong-Sub
authors.initials: DS
authors.lastname: Jung
authors.forename: Seung-Jae
authors.initials: SJ
authors.lastname: Kwak
authors.forename: Dong-Ryeol
authors.initials: DR
authors.lastname: Ryu
authors.forename: Seung Hyeok
authors.initials: SH
authors.lastname: Han
authors.forename: Jung Eun
authors.initials: JE
authors.lastname: Lee
authors.forename: Sung Jin
authors.initials: SJ
authors.lastname: Moon
authors.forename: Dae Suk
authors.initials: DS
authors.lastname: Han
authors.forename: Shin-Wook
authors.initials: SW
authors.lastname: Kang
mesh_terms: D043209:11-beta-Hydroxysteroid Dehydrogenase Type 2 / Q000096:biosynthesis / Q000235:genetics; D000450:Aldosterone / Q000502:physiology*; D000818:Animals; D017209:Apoptosis / Q000502:physiology*; D051017:Apoptosis Regulatory Proteins / Q000096:biosynthesis / Q000235:genetics; D001562:Benzimidazoles; D015153:Blotting, Western; D053148:Caspase 3 / Q000096:biosynthesis / Q000235:genetics; D002452:Cell Count; D002478:Cells, Cultured; D019405:Cytochrome P-450 CYP11B2 / Q000096:biosynthesis / Q000235:genetics; D003921:Diabetes Mellitus, Experimental / Q000473:pathology*; D005455:Fluorescent Antibody Technique; D005456:Fluorescent Dyes; D020287:In Situ Nick-End Labeling; D007668:Kidney / Q000473:pathology; D007678:Kidney Glomerulus / Q000378:metabolism; D008297:Male; D050199:Podocytes / Q000502:physiology*; D051381:Rats; D017207:Rats, Sprague-Dawley; D018161:Receptors, Mineralocorticoid / Q000096:biosynthesis / Q000235:genetics; D020133:Reverse Transcriptase Polymerase Chain Reaction
chemical_list: D051017:Apoptosis Regulatory Proteins; D001562:Benzimidazoles; D005456:Fluorescent Dyes; D018161:Receptors, Mineralocorticoid; D000450:Aldosterone; D043209:11-beta-Hydroxysteroid Dehydrogenase Type 2; D019405:Cytochrome P-450 CYP11B2; D053148:Caspase 3; C017807:bisbenzimide ethoxide trihydrochloride