title: Angiotensin II induces nephrin dephosphorylation and podocyte injury: role of caveolin-1.
abstract: Nephrin, an important structural and signal molecule of podocyte slit-diaphragm (SD), has been suggested to contribute to the angiotensin II (Ang II)-induced podocyte injury. Caveolin-1 has been demonstrated to play a crucial role in signaling transduction. In the present study, we evaluated the role of caveolin-1 in Ang II-induced nephrin phosphorylation in podocytes. Wistar rats-receiving either Ang II (400 ng/kg/min) or normal saline (via subcutaneous osmotic mini-pumps, control) were administered either vehicle or telmisartan (3 mg/kg/min) for 14 or 28 days. Blood pressure, 24-hour urinary albumin and serum biochemical profile were measured at the end of the experimental period. Renal histomorphology was evaluated through light and electron microscopy. In vitro, cultured murine podocytes were exposed to Ang II (10(-6)M) pretreated with or without losartan (10(-5) M) for variable time periods. Nephrin and caveolin-1 expression and their phosphorylation were analyzed by Western-blotting and immunofluorescence. Caveolar membrane fractions were isolated by sucrose density gradient centrifugation, and then the distribution and interactions between Ang II type 1 receptor (AT1), nephrin, C-terminal Src kinase (Csk) and caveolin-1 were evaluated using Western-blotting and co-immunoprecipitation. Podocyte apoptosis was evaluated by cell nucleus staining with Hoechst-33342. Ang II-receiving rats displayed diminished phosphorylation of nephrin but enhanced glomerular/podocyte injury and proteinuria when compared to control rats. Under control conditions, podocyte displayed expression of caveolin-1 in abundance but only a low level of phospho moiety. Nonetheless, Ang II stimulated caveolin-1 phosphorylation without any change in total protein expression. Nephrin and caveolin-1 were co-localized in caveolae fractions. AT1 receptors and Csk were moved to caveolae fractions and had an interaction with caveolin-1 after the stimulation with Ang II. Transfection of caveolin-1 plasmid (pEGFPC3-cav-1) significantly increased Ang II-induced nephrin dephosphorylation and podocyte apoptosis. Furthermore, knockdown of caveolin-1 expression (using siRNA) inhibited nephrin dephosphorylation and prevented Ang II-induced podocyte apoptosis. These findings indicate that Ang II induces nephrin dephosphorylation and podocyte injury through a caveolin-1-dependent mechanism.
authors: Division of Nephrology, Renmin Hospital of Wuhan University, Wuhan, Hubei, China.
authors: Zhilong
authors: Z
authors: Ren
authors: Division of Nephrology, Renmin Hospital of Wuhan University, Wuhan, Hubei, China.
authors: Wei
authors: W
authors: Liang
authors: Division of Nephrology, Renmin Hospital of Wuhan University, Wuhan, Hubei, China.
authors: Cheng
authors: C
authors: Chen
authors: Division of Nephrology, Renmin Hospital of Wuhan University, Wuhan, Hubei, China.
authors: Hongxia
authors: H
authors: Yang
authors: Medicine, North Shore-Long Island Jewish Health System, Manhasset, NY, USA.
authors: Pravin C
authors: PC
authors: Singhal
authors: Division of Nephrology, Renmin Hospital of Wuhan University, Wuhan, Hubei, China. Electronic address: ghxding@gmail.com.
authors: Guohua
authors: G
authors: Ding
mesh_terms: D000804:Angiotensin II / Q000008:administration & dosage* / Q000009:adverse effects; D047228:Angiotensin II Type 1 Receptor Blockers / Q000494:pharmacology; D000818:Animals; D017209:Apoptosis / Q000187:drug effects; D001562:Benzimidazoles / Q000032:analysis; D015153:Blotting, Western; D000081247:CSK Tyrosine-Protein Kinase; D021941:Caveolae / Q000187:drug effects / Q000378:metabolism; D051242:Caveolin 1 / Q000235:genetics / Q000378:metabolism*; D047468:Immunoprecipitation; D019808:Losartan / Q000494:pharmacology; D008565:Membrane Proteins / Q000235:genetics / Q000378:metabolism*; D051379:Mice; D010766:Phosphorylation; D010957:Plasmids; D050199:Podocytes / Q000187:drug effects / Q000378:metabolism* / Q000473:pathology; D011505:Protein-Tyrosine Kinases / Q000235:genetics / Q000378:metabolism*; D011507:Proteinuria / Q000235:genetics / Q000378:metabolism / Q000473:pathology; D051381:Rats; D017208:Rats, Wistar; D044140:Receptor, Angiotensin, Type 1 / Q000235:genetics / Q000378:metabolism*; D015398:Signal Transduction; D014162:Transfection; D019061:src-Family Kinases
chemical_list: D047228:Angiotensin II Type 1 Receptor Blockers; D001562:Benzimidazoles; D051242:Caveolin 1; D008565:Membrane Proteins; D044140:Receptor, Angiotensin, Type 1; C113267:nephrin; D000804:Angiotensin II; D011505:Protein-Tyrosine Kinases; D000081247:CSK Tyrosine-Protein Kinase; D019061:src-Family Kinases; D019808:Losartan; C017807:bisbenzimide ethoxide trihydrochloride