title: Insulin increases glomerular filtration barrier permeability through PKGIα-dependent mobilization of BKCa channels in cultured rat podocytes.
abstract: Podocytes are highly specialized cells that wrap around glomerular capillaries and comprise a key component of the glomerular filtration barrier. They are uniquely sensitive to insulin; like skeletal muscle and fat cells, they exhibit insulin-stimulated glucose uptake and express glucose transporters. Podocyte insulin signaling is mediated by protein kinase G type I (PKGI), and it leads to changes in glomerular permeability to albumin. Here, we investigated whether large-conductance Ca²⁺-activated K⁺ channels (BKCa) were involved in insulin-mediated, PKGIα-dependent filtration barrier permeability. Insulin-induced glomerular permeability was measured in glomeruli isolated from Wistar rats. Transepithelial albumin flux was measured in cultured rat podocyte monolayers. Expression of BKCa subunits was detected by RT-PCR. BKCa, PKGIα, and upstream protein expression were examined in podocytes with Western blotting and immunofluorescence. The BKCa-PKGIα interaction was assessed with co-immunoprecipitation. RT-PCR showed that primary cultured rat podocytes expressed mRNAs that encoded the pore-forming α subunit and four accessory β subunits of BKCa. The BKCa inhibitor, iberiotoxin (ibTX), abolished insulin-dependent glomerular albumin permeability and PKGI-dependent transepithelial albumin flux. Insulin-evoked albumin permeability across podocyte monolayers was also blocked with BKCa siRNA. Moreover, ibTX blocked insulin-induced disruption of the actin cytoskeleton and changes in the phosphorylation of PKG target proteins, MYPT1 and RhoA. These results indicated that insulin increased filtration barrier permeability through mobilization of BKCa channels via PKGI in cultured rat podocytes. This molecular mechanism may explain podocyte injury and proteinuria in diabetes.
authors.affiliation: Mossakowski Medical Research Centre, Polish Academy of Sciences, Laboratory of Molecular and Cellular Nephrology, Gdańsk, Poland. Electronic address: apiwkowska@imdik.pan.pl.
authors.forename: Agnieszka
authors.initials: A
authors.lastname: Piwkowska
authors.affiliation: Mossakowski Medical Research Centre, Polish Academy of Sciences, Laboratory of Molecular and Cellular Nephrology, Gdańsk, Poland.
authors.forename: Dorota
authors.initials: D
authors.lastname: Rogacka
authors.affiliation: Mossakowski Medical Research Centre, Polish Academy of Sciences, Laboratory of Molecular and Cellular Nephrology, Gdańsk, Poland.
authors.forename: Irena
authors.initials: I
authors.lastname: Audzeyenka
authors.affiliation: Medical University of Gdańsk, Department of Therapy Monitoring and Pharmacogenetics, Poland.
authors.forename: Małgorzata
authors.initials: M
authors.lastname: Kasztan
authors.affiliation: Mossakowski Medical Research Centre, Polish Academy of Sciences, Laboratory of Molecular and Cellular Nephrology, Gdańsk, Poland.
authors.forename: Stefan
authors.initials: S
authors.lastname: Angielski
authors.affiliation: Mossakowski Medical Research Centre, Polish Academy of Sciences, Laboratory of Molecular and Cellular Nephrology, Gdańsk, Poland; Medical University of Gdańsk, Department of Clinical Chemistry, Poland.
authors.forename: Maciej
authors.initials: M
authors.lastname: Jankowski
mesh_terms: D000418:Albumins / Q000378:metabolism; D000818:Animals; D020013:Calcium Signaling / Q000187:drug effects; D002478:Cells, Cultured; D062965:Cyclic GMP-Dependent Protein Kinase Type I / Q000502:physiology*; D005260:Female; D060730:Glomerular Filtration Barrier / Q000187:drug effects* / Q000378:metabolism*; D007328:Insulin / Q000494:pharmacology*; D051037:Large-Conductance Calcium-Activated Potassium Channel alpha Subunits / Q000378:metabolism*; D010455:Peptides / Q000494:pharmacology; D010539:Permeability / Q000187:drug effects; D050199:Podocytes / Q000187:drug effects* / Q000378:metabolism; D021381:Protein Transport / Q000187:drug effects; D051381:Rats; D017208:Rats, Wistar
chemical_list: D000418:Albumins; D007328:Insulin; C494842:Kcnma1 protein, rat; D051037:Large-Conductance Calcium-Activated Potassium Channel alpha Subunits; D010455:Peptides; C064719:iberiotoxin; D062965:Cyclic GMP-Dependent Protein Kinase Type I
keywords: Constriction apparatus; Filtration barrier permeability; Insulin; Large-conductance Ca(2+)-activated K(+) channels; Podocyte