Objective—
Atherosclerosis, an inflammatory disease of arterial vessel walls, requires migration and matrix metalloproteinase (MMP)-9–dependent invasion of monocytes/macrophages into the vascular wall. MMP-9 expression is stimulated by transcription factor nuclear factor-κB, which is regulated by inhibitor κB (IκB) and thus IκB kinase. Regulators of nuclear factor-κB include serum- and glucocorticoid-inducible kinase 1 (SGK1). The present study explored involvement of SGK1 in vascular inflammation and atherogenesis.
Approach and Results—
Gene-targeted apolipoprotein E (ApoE)–deficient mice without (
apoe
−/−
sgk1
+/+
) or with (
apoe
−/−
sgk1
−/−
) additional SGK1 knockout received 16-week cholesterol-rich diet. According to immunohistochemistry atherosclerotic lesions in aorta and carotid artery, vascular CD45
+
leukocyte infiltration, Mac-3
+
macrophage infiltration, vascular smooth muscle cell content, MMP-2, and MMP-9 positive areas in atherosclerotic tissue were significantly less in
apoe
−/−
sgk1
−/−
mice than in
apoe
−/−
sgk1
+/+
mice. As determined by Boyden chamber, thioglycollate-induced peritonitis and air pouch model, migration of SGK1-deficient CD11b
+
F4/80
+
macrophages was significantly diminished in vitro and in vivo. Zymographic MMP-2 and MMP-9 production, MMP-9 activity and invasion through matrigel in vitro were significantly less in
sgk1
−/−
than in
sgk1
+/+
macrophages and in control plasmid–transfected or inactive
K127N
SGK1-transfected than in constitutively active
S422D
SGK1-transfected THP-1 cells. Confocal microscopy revealed reduced macrophage number and macrophage MMP-9 content in plaques of
apoe
−/−
sgk1
−/−
mice. In THP-1 cells, MMP-inhibitor GM6001 (25 μmol/L) abrogated
S422D
SGK1-induced MMP-9 production and invasion. According to reverse transcription polymerase chain reaction, MMP-9 transcript levels were significantly reduced in
sgk1
−/−
macrophages and strongly upregulated in
S422D
SGK1-transfected THP-1 cells compared with control plasmid–transfected or
K127N
SGK1-transfected THP-1 cells. According to immunoblotting and confocal microscopy, phosphorylation of IκB kinase and inhibitor κB and nuclear translocation of p50 were significantly lower in
sgk1
−/−
macrophages than in
sgk1
+/+
macrophages and significantly higher in
S422D
SGK1-transfected THP-1 cells than in control plasmid–transfected or
K127N
SGK1-transfected THP-1 cells. Treatment of
S422D
SGK1-transfected THP-1 cells with IκB kinase-inhibitor BMS-345541 (10 μmol/L) abolished
S422D
SGK1-induced increase of MMP-9 transcription and gelatinase activity.
Conclusions—
SGK1 plays a pivotal role in vascular inflammation during atherogenesis. SGK1 participates in the regulation of monocyte/macrophage migration and MMP-9 transcription via regulation of nuclear factor-κB.
Endomyocardial expression of SDF-1 is enhanced in inflammatory cardiomyopathy, positively correlates with myocardial fibrosis and identifies high-risk patients with suspected myocarditis.
Platelet adhesion, activation, and aggregation are essential for primary hemostasis, but are also critically involved in the development of acute arterial thrombotic occlusion. Stimulation of the collagen receptor glycoprotein VI (GPVI) leads to phospholipase Cγ2-dependent inositol triphosphate (IP) production with subsequent platelet activation, due to increased intracellular Ca concentration ([Ca]). Although tricyclic antidepressants have been shown to potentially impair platelet activation, nothing is hitherto known about potential effects of the tricyclic antidepressant doxepin on platelet Ca signaling and thrombus formation. As shown in the present study, doxepin significantly diminished the stimulatory effect of GPVI agonist collagen-related peptide (CRP) on intracellular Ca release as well as subsequent extracellular Ca influx. Doxepin was partially effective by impairment of CRP-dependent IP production. Moreover, doxepin abrogated CRP-induced platelet degranulation and integrin αβ activation and aggregation. Finally, doxepin markedly blunted in vitro platelet adhesion to collagen and thrombus formation under high arterial shear rates (1,700). In conclusion, doxepin is a powerful inhibitor of GPVI-dependent platelet Ca signaling, platelet activation, and thrombus formation.
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