The intake of ω-3 polyunsaturated fatty acids (PUFAs), which are abundant in marine fish meat and oil, has been shown to exert many beneficial effects. The mechanisms behind those effects are numerous, including interference with the arachidonic acid cascade that produces pro-inflammatory eicosanoids, formation of novel bioactive lipid mediators, and change in the pattern of secreted adipocytokines. In our study, we show that eicosapentaenoic acid (EPA) increases secreted adiponectin from 3T3-L1 adipocytes and in plasma of mice as early as 4 days after initiation of an EPA-rich diet. Using 3T3-L1 adipocytes, we report for the first time that 15-deoxy-δ12,14-PGJ3 (15d-PGJ3), a product of EPA, also increases the secretion of adiponectin. We demonstrate that the increased adiponectin secretion induced by 15d-PGJ3 is partially peroxisome proliferator-activated receptor-gamma (PPAR-γ)-mediated. Finally, we show that 3T3-L1 adipocytes can synthesize 15d-PGJ3 from EPA. 15d-PGJ3 was also detected in adipose tissue from EPA-fed mice. Thus, these studies provide a novel mechanism(s) for the therapeutic benefits of ω-3 polyunsaturated fatty acids dietary supplementation.
BackgroundFibrin provides a temporary matrix at the site of vascular injury. The aims of the present work were (1) to follow fibrin formation and lysis onto the surface of human dermal microvascular endothelial cells (HMEC-1), and (2) to quantify the secretion of fibrinolytic components in the presence of fibrin.MethodsFibrin clots at different fibrinogen concentrations were formed on top of (model 1) or beneath (model 2) the endothelial cells. Fibrin formation or lysis onto the surface of HMEC-1 cells, was followed by turbidity. Clot structure was visualized by laser scanning confocal microscopy (LSCM). The secretion of uPA and PAI-1 by HMEC-1 cells was quantified by ELISA.ResultsThe rate of fibrin formation increased approximately 1.5-fold at low fibrinogen content (0.5 and 1 mg/mL; p < 0.05) compared to the condition without cells; however, it was decreased at 2 mg/mL fibrinogen (p < 0.05) and no differences were found at higher fibrinogen concentrations (3 and 5 mg/mL). HMEC-1 retarded dissolution of clots formed onto their surface at 0.5 to 3 mg/mL fibrinogen (p < 0.05). Secretion of uPA was 13 × 10−6 ng/mL per cell in the absence of RGD and 8 × 10−6 ng/mL per cell in the presence of RGD, when clots were formed on the top of HMEC-1. However, the opposite was found when cells were grown over fibrin: 6 × 10−6 ng/mL per cell without RGD vs. 17 × 10−6 ng/mL per cell with RGD. The secretion of PAI-1 by HMEC-1 cells was unrelated to the presence of fibrin or RGD, 7 × 10−6 μg/mL per cell and 5 × 10−6 μg/mL per cell, for the apical (model 1) and basal clots (model 2), respectively.ConclusionsHMEC-1 cells influence fibrin formation and dissolution as a function of the fibrin content of clots. Clot degradation was accentuated at high fibrin concentrations. The secretion of fibrinolytic components by HMEC-1 cells seemed to be modulated by integrins that bind RGD ligands.
1 Endothelial cells play an important role in the modulation of vascular tone because of their ability to produce vasoactive substances such as prostacyclin (PGI 2 ). Cell -cell contact between human umbilical vein endothelial cells (HUVEC) and peripheral blood lymphocytes has been shown to stimulate endothelial PGI 2 synthesis by increasing free arachidonic acid availability through endothelial cytosolic phospholipase A2 (cPLA 2 ) activation. In this study, we sought to determine whether phospholipase C (PLC) and D (PLD) activation also contributes, besides cPLA 2 , to the lymphocyte-induced PGI 2 synthesis in HUVEC, and to delineate further the potential mechanisms of cPLA 2 activation triggered by the interaction of HUVEC with lymphocytes. 2 Pretreatment of endothelial cells with the PI-PLC inhibitor U-73122 before the coincubation with lymphocytes markedly inhibited the PGI 2 output whereas the diacylglycerol (DAG) lipase inhibitor RHC 80267 and ethanol had no effect. These results suggest that PLC may be involved through inositol trisphosphate generation and calcium mobilization, and that neither DAG nor phosphatidic acid (PtdOH) was used as sources of arachidonic acid. 3 The stimulated PGI 2 synthesis was protein kinase C (PKC)-independent but strongly inhibited by the mitogen-activated protein kinase kinase (MEK) inhibitors PD98059 and U-0126 and by the Src kinase inhibitor PP1. 4 Immunoblot experiments showed an increased phosphorylation of the extracellular signalregulated kinases 1/2 (ERK1/2) upon lymphocyte addition till 4 h coincubation. Phosphorylation was markedly inhibited by U-0126 and PP1 addition. 5 Collectively, these results suggest that the signaling cascade triggered by lymphocytes in endothelial cells involves an Src kinase/ERK1/2 pathway leading to endothelial cPLA 2 activation.
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