High glucose-induced upregulation of osteopontin is mediated via Rho/Rho kinase pathway in cultured rat aortic smooth muscle cells.
OBJECTIVE
Osteopontin is upregulated in the diabetic vascular wall and in vascular smooth muscle cells cultured under high glucose concentration. In the present study, we analyzed the mechanism of high glucose-induced upregulation of osteopontin in cultured rat aortic smooth muscle cells.


RESULTS
We found that an inhibitor of Rho-associated protein kinase, Y-27632, suppressed osteopontin mRNA expression under high glucose concentration. Transfection of cells with a constitutive active Rho mutant, pSRalpha-myc-RhoDA, enhanced osteopontin mRNA expression. Furthermore, incubation of cells under high glucose concentration activated Rho, indicating that Rho/Rho kinase pathway mediates high-glucose-stimulated osteopontin expression. Treatment of cells with an inhibitor of protein kinase C, GF109203X, and azaserine, an inhibitor of the hexosamine pathway, suppressed high glucose-induced Rho activation. Glucosamine treatment was shown to activate Rho. Treatment of cells with an inhibitor of MEK1, PD98059, suppressed osteopontin mRNA expression under high glucose concentration. Incubation of cells under high glucose concentration activated ERK. Finally, transfection of cells with pSRalpha-myc-RhoDA also activated ERK.


CONCLUSIONS
In conclusion, our present findings support a notion that Rho/Rho kinase pathway functions downstream of protein kinase C and the hexosamine pathways and upstream of ERK in mediating high-glucose-induced upregulation of osteopontin expression.
Kawamura
Harukiyo
H
Department of Clinical Cell Biology and Medicine, Chiba University Graduate School of Medicine, 1-8-1 Inohana, Chiba 260-8670, Japan.
Yokote
Koutaro
K
Asaumi
Sunao
S
Kobayashi
Kazuki
K
Fujimoto
Masaki
M
Maezawa
Yoshiro
Y
Saito
Yasushi
Y
Mori
Seijiro
S
D000577:Amides / Q000494:pharmacology; D000818:Animals; D001011:Aorta / Q000166:cytology*; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005947:Glucose / Q000494:pharmacology*; D019161:Hydroxymethylglutaryl-CoA Reductase Inhibitors / Q000494:pharmacology; D007211:Indoles / Q000494:pharmacology; D047908:Intracellular Signaling Peptides and Proteins; D020935:MAP Kinase Signaling System / Q000187:drug effects / Q000502:physiology; D008297:Male; D008301:Maleimides / Q000494:pharmacology; D020928:Mitogen-Activated Protein Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism; D009131:Muscle, Smooth, Vascular / Q000166:cytology / Q000187:drug effects / Q000201:enzymology* / Q000378:metabolism*; D053495:Osteopontin; D011493:Protein Kinase C / Q000037:antagonists & inhibitors / Q000378:metabolism; D017368:Protein Prenylation / Q000502:physiology; D017346:Protein Serine-Threonine Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism*; D011725:Pyridines / Q000494:pharmacology; D011804:Quinolines / Q000494:pharmacology; D051381:Rats; D017208:Rats, Wistar; D012795:Sialoglycoproteins / Q000378:metabolism*; D015854:Up-Regulation / Q000502:physiology*; D054460:rho-Associated Kinases; D020742:rhoA GTP-Binding Protein / Q000378:metabolism*
D000577:Amides; D004791:Enzyme Inhibitors; D019161:Hydroxymethylglutaryl-CoA Reductase Inhibitors; D007211:Indoles; D047908:Intracellular Signaling Peptides and Proteins; D008301:Maleimides; D011725:Pyridines; D011804:Quinolines; D012795:Sialoglycoproteins; C506897:Spp1 protein, rat; D053495:Osteopontin; C108830:Y 27632; D017346:Protein Serine-Threonine Kinases; D054460:rho-Associated Kinases; D011493:Protein Kinase C; D020928:Mitogen-Activated Protein Kinases; D020742:rhoA GTP-Binding Protein; D005947:Glucose; C070515:bisindolylmaleimide I; C108475:pitavastatin