Sphingosine-1-phosphate inhibits high glucose-mediated ERK1/2 action in endothelium through induction of MAP kinase phosphatase-3.
Endothelial activation is a key early event in vascular complications of Type 1 diabetes. The nonobese diabetic (NOD) mouse is a well-characterized model of Type 1 diabetes. We previously reported that Type 1 diabetic NOD mice have increased endothelial activation, with increased production of monocyte chemoattractant protein (MCP)-1 and IL-6, and a 30% increase of surface VCAM-1 expression leading to a fourfold increase in monocyte adhesion to the endothelium. Sphingosine-1-phosphate (S1P) prevents monocyte:endothelial interactions in these diabetic NOD mice. Incubation of diabetic NOD endothelial cells (EC) with S1P (100 nmol/l) reduced ERK1/2 phosphorylation by 90%, with no significant changes in total ERK1/2 protein. In the current study, we investigated the mechanism of S1P action on ERK1/2 to reduce activation of diabetic endothelium. S1P caused a significant threefold increase in mitogen-activated kinase phosphatase-3 (MKP-3) expression in EC. MKP-3 selectively regulates ERK1/2 activity through dephosphorylation. Incubation of diabetic NOD EC with S1P and the S1P(1)-selective agonist SEW2871 significantly increased expression of MKP-3 and reduced ERK1/2 phosphorylation, while incubation with the S1P(1)/S1P(3) antagonist VPC23019 decreased the expression of MKP-3, both results supporting a role for S1P(1) in MKP-3 regulation. To mimic the S1P-mediated induction of MKP-3 diabetic NOD EC, we overexpressed MKP-3 in human aortic endothelial cells (HAEC) cultured in elevated glucose (25 mmol/l). Overexpression of MKP-3 in glucose-cultured HAEC decreased ERK1/2 phosphorylation and resulted in decreased monocyte:endothelial interactions in a static monocyte adhesion assay. Finally, we used small interfering RNA to MKP-3 and observed increased monocyte adhesion. Moreover, S1P was unable to inhibit monocyte adhesion in the absence of MKP-3. Thus, one mechanism for the anti-inflammatory action of S1P in diabetic EC is inhibition of ERK1/2 phosphorylation through induction of MKP-3 expression via the S1P-S1P(1) receptor axis.
Whetzel
Angela M
AM
Cardiovascular Research Center, Univ. of Virginia, P. O. Box 801394, 415 Lane Rd., MR5 Rm. G123, Charlottesville, VA 22908, USA.
Bolick
David T
DT
Hedrick
Catherine C
CC
D000818:Animals; D002448:Cell Adhesion; D002478:Cells, Cultured; D003922:Diabetes Mellitus, Type 1 / Q000201:enzymology*; D004195:Disease Models, Animal; D054642:Dual Specificity Phosphatase 6 / Q000235:genetics / Q000378:metabolism*; D042783:Endothelial Cells / Q000201:enzymology*; D005947:Glucose / Q000378:metabolism*; D006801:Humans; D007249:Inflammation / Q000201:enzymology / Q000517:prevention & control; D008246:Lysophospholipids / Q000378:metabolism*; D051379:Mice; D016688:Mice, Inbred NOD; D019950:Mitogen-Activated Protein Kinase 1 / Q000378:metabolism*; D048052:Mitogen-Activated Protein Kinase 3 / Q000378:metabolism*; D009000:Monocytes / Q000378:metabolism; D010766:Phosphorylation; D034622:RNA Interference; D049349:Receptors, Lysosphingolipid / Q000378:metabolism; D013110:Sphingosine / Q000031:analogs & derivatives* / Q000378:metabolism; D014162:Transfection
D008246:Lysophospholipids; D049349:Receptors, Lysosphingolipid; C060506:sphingosine 1-phosphate; D019950:Mitogen-Activated Protein Kinase 1; D048052:Mitogen-Activated Protein Kinase 3; C517035:DUSP6 protein, human; D054642:Dual Specificity Phosphatase 6; C517036:Dusp6 protein, mouse; D005947:Glucose; D013110:Sphingosine