High D-Glucose reduces SLC29A1 promoter activity and adenosine transport involving specific protein 1 in human umbilical vein endothelium

Puebla, C; Farias M.; Gonzalez, M.; Vecchiola, A; Aguayo, C.; Krause, B; Pastor-Anglada, M; Casanello, P; Sobrevia L.

Abstract

High D-glucose reduces human equilibrative nucleoside transporter 1 (hENT1)-mediated adenosine uptake involving endothelial nitric oxide synthase (eNOS), mitogen-activated protein (MAP) kinase kinases 1 and 2/MAP kinases p42/44 (MEK/ERKs), and protein kinase C (PKC) activation in human umbilical vein endothelium (HUVEC). Since NO represses SLC29A1 gene (hENT1) promoter activity we studied whether D-glucose-reduced hENT1-adenosine transport results from lower SLC29A1 expression in HUVEC primary cultures. HUVEC incubation (24 h) with high D-glucose (25 mM) reduced hENT1-adenosine transport and pGL3-hENT1 1114 construct SLC29A1 reporter activity compared with normal D-glucose (5 mM). High D-glucose also reduced pGL3-hENT1 1114 reporter activity compared with cells transfected with pGL3-hENT1 795 construct. NG-nitro-L-arginine methyl ester (L-NAME, NOS inhibitor), PD-98059 (MEK1/2 inhibitor), and/or calphostin C (PKC inhibitor) blocked D-glucose effects. Insulin (1 nM) and phorbol 12-myristate 13-acetate (PMA, 100 nM, PKC activator), but not 4α-phorbol 12,13-didecanoate (4αPDD, 100 nM, PMA less active analogue) reduced hENT1-adenosine transport. L-NAME and PD-98059 blocked insulin effects. L-NAME, PD-98059, and calphostin C increased hENT1 expression without altering protein or mRNA stability. High D-glucose increased Sp1 transcription factor protein abundance and binding to SLC29A1 promoter, phenomena blocked by L-NAME, PD-98059, and calphostin C. Sp1 overexpression reduced SLC29A1 promoter activity in normal D-glucose, an effect reversed by L-NAME and further reduced by S-nitroso-N-acetyl-L,D-penicillamine (SNAP, NO donor) in high D-glucose. Thus, reduced hENT1-mediated adenosine transport in high D-glucose may result from increased Sp1 binding to SLC29A1 promoter down-regulating hENT1 expression. This phenomenon depends on eNOS, MEK/ERKs, and PKC activity, suggesting potential roles for these molecules in hyperglycemia-associated endothelial dysfunction. © 2007 Wiley-Liss, Inc.

Más información

Título según WOS: High D-Glucose reduces SLC29A1 promoter activity and adenosine transport involving specific protein 1 in human umbilical vein endothelium
Título según SCOPUS: High D-glucose reduces SLC29A1 promoter activity and adenosine transport involving specific protein 1 in human umbilical vein endothelium
Título de la Revista: JOURNAL OF CELLULAR PHYSIOLOGY
Volumen: 215
Número: 3
Editorial: WILEY-BLACKWELL
Fecha de publicación: 2008
Página de inicio: 645
Página final: 656
Idioma: English
DOI:

10.1002/21347

Notas: ISI, SCOPUS