Over-expression of mitogen-activated protein kinase phosphatase-2 enhances adhesion molecule expression and protects against apoptosis in human endothelial cells.
OBJECTIVE
We assessed the effects of over-expressing the dual-specific phosphatase, mitogen-activated protein (MAP) kinase phosphatase-2 (MKP-2), in human umbilical vein endothelial cells (HUVECs) on inflammatory protein expression and apoptosis, two key features of endothelial dysfunction in disease.


METHODS
We infected HUVECs for 40 h with an adenoviral version of MKP-2 (Adv.MKP-2). Tumour necrosis factor (TNF)-α-stimulated phosphorylation of MAP kinase and protein expression was measured by Western blotting. Cellular apoptosis was assayed by FACS.


RESULTS
Infection with Adv.MKP-2 selectively abolished TNF-α-mediated c-Jun-N-terminal kinase (JNK) activation and had little effect upon extracellular signal-regulated kinase or p38 MAP kinase. Adv.MKP-2 abolished COX-2 expression, while induction of the endothelial cell adhesion molecules, intercellular adhesion molecule (ICAM) and vascular cell adhesion molecule (VCAM), two NFκB-dependent proteins, was not affected. However, when ICAM and VCAM expression was partly reduced by blockade of the NFκB pathway, Adv.MKP-2 was able to reverse this inhibition. This correlated with enhanced TNF-α-induced loss of the inhibitor of κB (IκB)α loss, a marker of NFκB activation. TNF-α in combination with NFκB blockade also increased HUVEC apoptosis; this was significantly reversed by Adv.MKP-2. Protein markers of cellular damage and apoptosis, H2AX phosphorylation and caspase-3 cleavage, were also reversed by MKP-2 over-expression.


CONCLUSIONS
Over-expression of MKP-2 had different effects upon the expression of inflammatory proteins due to a reciprocal effect upon JNK and NFκB signalling, and also prevented TNF-α-mediated endothelial cell death. These properties may make Adv.MKP-2 a potentially useful future therapy in cardiovascular diseases where endothelial dysfunction is a feature.
Al-Mutairi
Mashael
M
Division of Physiology and Pharmacology, University of Strathclyde, Strathclyde Institute for Biomedical Sciences, Glasgow, UK.
Al-Harthi
Sameer
S
Cadalbert
Laurence
L
Plevin
Robin
R
D000256:Adenoviridae / Q000235:genetics; D017209:Apoptosis; D002478:Cells, Cultured; D054637:Dual-Specificity Phosphatases / Q000235:genetics / Q000378:metabolism*; D042783:Endothelial Cells / Q000378:metabolism; D004730:Endothelium, Vascular / Q000378:metabolism*; D015870:Gene Expression; D006801:Humans; D018799:Intercellular Adhesion Molecule-1 / Q000378:metabolism; D048031:JNK Mitogen-Activated Protein Kinases / Q000378:metabolism; D054639:Mitogen-Activated Protein Kinase Phosphatases / Q000235:genetics / Q000378:metabolism*; D016328:NF-kappa B / Q000378:metabolism; D010766:Phosphorylation; D015398:Signal Transduction; D014409:Tumor Necrosis Factor-alpha / Q000008:administration & dosage / Q000378:metabolism; D014471:Umbilical Veins / Q000166:cytology / Q000378:metabolism; D019010:Vascular Cell Adhesion Molecule-1 / Q000378:metabolism
D016328:NF-kappa B; D014409:Tumor Necrosis Factor-alpha; D019010:Vascular Cell Adhesion Molecule-1; D018799:Intercellular Adhesion Molecule-1; D048031:JNK Mitogen-Activated Protein Kinases; D054639:Mitogen-Activated Protein Kinase Phosphatases; C490583:DUSP4 protein, human; D054637:Dual-Specificity Phosphatases