title: High glucose increases glomerular filtration barrier permeability by activating protein kinase G type Iα subunits in a Nox4-dependent manner.
abstract: Hyperglycemia is a primary factor that disturbs podocyte function in the glomerular filtration process; this disturbance leads to the development of diabetic nephropathy, and ultimately, renal failure. Podocyte function may also be altered by biological agents that modify protein kinase activity, including the cGMP-activated protein kinase type Iα (PKGIα). We hypothesized that hyperglycemia-induced podocyte protein hyperpermeability was dependent on PKGIα activation, and that PKGIα was activated via dimerization induced by reactive oxygen species. This hypothesis was investigated in rat podocytes cultured in high glucose (HG, 30 mM). Protein expression was measured with Western blot and immunofluorescence. Podocyte permeability was measured with a transmembrane albumin flux assay. We found that HG increased podocyte permeability in long-term incubations (1, 3, and 5 days); permeability was increased by 66% on day 5. This effect was abolished with apocynin, a NAD(P)H inhibitor, and Rp-8-Br-cGMPS, a PKG inhibitor. It was also abolished by introducing small interfering RNAs (siRNAs) against Nox4 and PKGIα into cultured podocytes. Furthermore, HG increased PKGIα dimerization by 138% (0.23 ± 0.04 vs. 0.54 ± 0.09; P<0.05); this effect was abolished with a siRNA against Nox4. Our observations suggested that HG could increase albumin permeability across the podocyte filtration barrier via Nox4-dependent PKGIα dimerization.
authors.affiliation: Mossakowski Medical Research Centre Polish Academy of Sciences, Laboratory of Molecular and Cellular Nephrology Dębinki 7, Gdańsk 80-211, Poland. Electronic address: apiwkowska@cmdik.pan.pl.
authors.forename: Agnieszka
authors.initials: A
authors.lastname: Piwkowska
authors.forename: Dorota
authors.initials: D
authors.lastname: Rogacka
authors.forename: Irena
authors.initials: I
authors.lastname: Audzeyenka
authors.forename: Stefan
authors.initials: S
authors.lastname: Angielski
authors.forename: Maciej
authors.initials: M
authors.lastname: Jankowski
mesh_terms: D000818:Animals; D002478:Cells, Cultured; D062965:Cyclic GMP-Dependent Protein Kinase Type I / Q000378:metabolism*; D004305:Dose-Response Relationship, Drug; D005260:Female; D005919:Glomerular Filtration Rate / Q000187:drug effects*; D005947:Glucose / Q000494:pharmacology*; D006943:Hyperglycemia / Q000378:metabolism*; D007678:Kidney Glomerulus / Q000187:drug effects* / Q000201:enzymology; D000074663:NADPH Oxidase 4; D019255:NADPH Oxidases / Q000378:metabolism*; D010539:Permeability / Q000187:drug effects*; D050199:Podocytes / Q000166:cytology / Q000187:drug effects; D051381:Rats; D017208:Rats, Wistar; D013329:Structure-Activity Relationship
chemical_list: D000074663:NADPH Oxidase 4; D019255:NADPH Oxidases; C498911:Nox4 protein, rat; D062965:Cyclic GMP-Dependent Protein Kinase Type I; D005947:Glucose
keywords: Filtration barrier permeability; High glucose concentration; NAD(P)H oxidase; Podocyte; Protein kinase G type Iα