title: High glucose suppresses expression of equilibrative nucleoside transporter 1 (ENT1) in rat cardiac fibroblasts through a mechanism dependent on PKC-zeta and MAP kinases.
abstract: Recently it was demonstrated that the elevated concentration of glucose but not lack of insulin is responsible for suppression of equilibrative nucleoside transporter (ENT1) in diabetic rat cardiac fibroblasts (CFs). The present study was undertaken to determine the signaling pathway utilized by glucose to regulate the expression of ENT1 in the primary culture of rat CFs. Pretreatment of CFs with Go 6983, an isozyme non-selective PKC inhibitor, prevented the high glucose (25 mM) effect on ENT1 mRNA level and nitrobenzylthioinosine (NBTI)-sensitive adenosine uptake. Similar effect was observed with a cell-permeable PKC-zeta pseudosubstrate, whereas Go 6976 a selective inhibitor of Ca(2+)-dependent PKC-alpha and PKC-beta isozymes had little effect on high glucose-induced suppression of ENT1 mRNA level. Incubation of CFs with nitric oxide (NO) donors (SNAPE, SNP) or NO synthase inhibitors (L-NAME, L-NMMA) prior to exposition of CFs to high glucose did not change the glucose effect on ENT1 mRNA level. The high glucose-induced suppression of ENT1 expression was blocked by PD9859 (an inhibitor of MEK), whereas neither wortmannin (an inhibitor of PI3K) nor rapamycin (an inhibitor of mTOR) affected the glucose action on ENT1 transcript level. Highly effective in preventing the high glucose effect on ENT1 mRNA level were GW 5074 (an inhibitor of Raf kinase) and SB 203580 (selective p38 MAPK inhibitor). These findings indicate that high glucose suppresses the expression of ENT1 in CFs by NO independent manner involving the signaling through PKC-zeta, Raf-1, MEK, and p38 MAPK pathways.
authors.affiliation: Department of Molecular Medicine, Medical University of Gdansk, Gdansk, Poland.
authors.forename: Marzena
authors.initials: M
authors.lastname: Grden
authors.forename: Marzena
authors.initials: M
authors.lastname: Podgorska
authors.forename: Katarzyna
authors.initials: K
authors.lastname: Kocbuch
authors.forename: Robert
authors.initials: R
authors.lastname: Rzepko
authors.forename: Andrzej
authors.initials: A
authors.lastname: Szutowicz
authors.forename: Tadeusz
authors.initials: T
authors.lastname: Pawelczyk
mesh_terms: D000818:Animals; D002352:Carrier Proteins / Q000235:genetics / Q000378:metabolism*; D002478:Cells, Cultured; D004305:Dose-Response Relationship, Drug; D033721:Equilibrative Nucleoside Transporter 1; D005347:Fibroblasts / Q000166:cytology / Q000187:drug effects / Q000201:enzymology*; D005786:Gene Expression Regulation / Q000187:drug effects; D005947:Glucose / Q000494:pharmacology*; D008297:Male; D020929:Mitogen-Activated Protein Kinase Kinases / Q000378:metabolism; D020928:Mitogen-Activated Protein Kinases / Q000378:metabolism*; D008954:Models, Biological; D009206:Myocardium / Q000166:cytology* / Q000201:enzymology*; D009569:Nitric Oxide / Q000378:metabolism; D020030:Nitric Oxide Donors / Q000494:pharmacology; D010766:Phosphorylation / Q000187:drug effects; D011493:Protein Kinase C / Q000378:metabolism*; D047428:Protein Kinase Inhibitors / Q000494:pharmacology; D019908:Proto-Oncogene Proteins c-raf / Q000378:metabolism; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D051381:Rats; D017208:Rats, Wistar; D013997:Time Factors; D048051:p38 Mitogen-Activated Protein Kinases / Q000378:metabolism; D000099221:Protein Kinase C zeta
chemical_list: D002352:Carrier Proteins; D033721:Equilibrative Nucleoside Transporter 1; D005947:Glucose; D020929:Mitogen-Activated Protein Kinase Kinases; D020928:Mitogen-Activated Protein Kinases; D009569:Nitric Oxide; D020030:Nitric Oxide Donors; D011493:Protein Kinase C; D047428:Protein Kinase Inhibitors; D019908:Proto-Oncogene Proteins c-raf; D012333:RNA, Messenger; D048051:p38 Mitogen-Activated Protein Kinases; D000099221:Protein Kinase C zeta; C438652:Slc29a1 protein, rat