title: Glucose-induced regulation of NHEs activity and SGLTs expression involves the PKA signaling pathway.
abstract: The effect of glucose on the intracellular pH (pH(i)) recovery rate (dpH(i)/dt) and Na(+)-glucose transporter (SGLT) localization was investigated in HEK-293 cells, a cell line that expresses endogenous NHE1, NHE3, SGLT1, and SGLT2 proteins. The activity of the Na(+)/H(+) exchangers (NHEs) was evaluated by using fluorescence microscopy. The total and membrane protein expression levels were analyzed by immunoblotting. In cells cultivated in 5 mM glucose, the pH(i) recovery rate was 0.169 ± 0.020 (n = 6). This value did not change in response to the acute presence of glucose at 2 or 10 mM, but decreased with 25 mM glucose, an effect that was not observed with 25 mM mannitol. Conversely, the chronic effect of high glucose (25 mM) increased the pH(i) recovery rate (~40%, P < 0.05), without changes in the total levels of NHE1, NHE3, or SGLT1 expression, but increasing the total cellular (~50%, P < 0.05) and the plasma membrane (~100%, P < 0.01) content of SGLT2. Treatment with H-89 (10(-6) M) prevented the stimulatory effect of chronic glucose treatment on the pH(i) recovery rate and SGLT2 expression in the plasma membrane. Our results indicate that the effect of chronic treatment with a high glucose concentration is associated with increased NHEs activity and plasma membrane expression of SGLT2 in a protein kinase A-dependent way. The present results reveal mechanisms of glucotoxicity and may contribute to understanding the diabetes-induced damage of this renal epithelial cell.
authors.affiliation: Department of Physiology and Biophysics, Institute of Biomedical Sciences, University of São Paulo, São Paulo, SP 05508-900, Brazil.
authors.forename: Olívia
authors.initials: O
authors.lastname: Beloto-Silva
authors.forename: Ubiratan Fabres
authors.initials: UF
authors.lastname: Machado
authors.forename: Maria
authors.initials: M
authors.lastname: Oliveira-Souza
mesh_terms: D019921:Biotinylation / Q000187:drug effects; D027682:Cation Transport Proteins / Q000378:metabolism; D002460:Cell Line; D017868:Cyclic AMP-Dependent Protein Kinases / Q000378:metabolism*; D005947:Glucose / Q000494:pharmacology*; D006801:Humans; D015151:Immunoblotting; D015398:Signal Transduction / Q000187:drug effects*; D051269:Sodium-Glucose Transporter 1 / Q000378:metabolism; D051297:Sodium-Glucose Transporter 2 / Q000378:metabolism; D000074082:Sodium-Hydrogen Exchanger 1; D000074083:Sodium-Hydrogen Exchanger 3; D017923:Sodium-Hydrogen Exchangers / Q000378:metabolism*
chemical_list: D027682:Cation Transport Proteins; C495674:SLC5A1 protein, human; C089180:SLC5A2 protein, human; C490998:SLC9A1 protein, human; C000613385:SLC9A3 protein, human; D051269:Sodium-Glucose Transporter 1; D051297:Sodium-Glucose Transporter 2; D000074082:Sodium-Hydrogen Exchanger 1; D000074083:Sodium-Hydrogen Exchanger 3; D017923:Sodium-Hydrogen Exchangers; D017868:Cyclic AMP-Dependent Protein Kinases; D005947:Glucose