Eicosapentaenoic acid and docosahexaenoic acid modulate mitogen-activated protein kinase activity in endothelium

Abstract

Omega-3 polyunsaturated fatty acids (PUFA) regulate inflammation and immunoreaction partially via affecting endothelial functions. However, the intracellular signaling mechanisms for inhibiting endothelial activation by omega-3 PUFA remain unclear. We investigated the effects of eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) on mitogen-activated protein kinases (MAPK) of endothelium. We analyzed the expression of extracellular signal-related kinases (ERK1/2), Jun amino-terminal kinases (JNK), and p38 mRNA by real-time RT–PCR and the kinases activity by western blotting in tumor necrosis factor-alpha (TNF-α)-activated human umbilical vein endothelial cells (HUVEC). We observed that EPA or DHA alone significantly reduced the TNF-α-induced activation of p38 and JNK kinases at a concentration of 20 μM, but EPA is a more potent inhibitor than DHA. In contrast, both EPA and DHA significantly counteracted the TNF-α-mediated deactivation of ERK1/2 kinases. Meanwhile, both EPA and DHA significantly attenuated the TNF-α-induced expression of p38 and ERK1/2 mRNA, and DHA but not EPA also reduced the TNF-α-induced JNK mRNA expression. We present data show that both EPA and DHA alone diminish activation of p38 and JNK kinases, while maintaining the activation of ERK1/2 kinases of TNF-α-stimulated HUVEC. This may contribute to the inhibiting effects of omega-3 PUFA on endothelial activation by proinflammatory stimuli.

Introduction

Over the past 25 years, there has been a dramatic increase in the scientific scrutiny of and public interest in omega-3 polyunsaturated fatty acids (PUFA) and their impact on personal health. It has been recognized that omega-3 PUFA, especially eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA), possess immunoregulatory and antiinflammatory properties. The beneficial effect of EPA and DHA has been demonstrated in several human diseases including atherosclerotic, autoimmune, inflammatory diseases, and sepsis (Calder, 1998, Grimm et al., 2002, Simopoulos, 2002, Yaqoob, 2004). The endothelium acts to maintain vascular homeostasis through multiple complex interactions with cells in the vessel wall and lumen. The endothelial cells play a key role in inflammation and immunoreaction (Cines et al., 1998). It has become apparent that EPA and DHA regulate inflammation and immunoreaction partially via affecting endothelial functions. EPA and DHA significantly decreased the cytokine-induced adhesion molecule expression by endothelial cells (De Caterina et al., 1994, De Caterina et al., 1995, Chen et al., 2003a, Weber et al., 1995), diminished the adhesion of leukocytes to the stimulated endothelial cells (Weber et al., 1995, Khalfoun et al., 1996, Sanderson and Calder, 1998, Mayer et al., 2002), inhibited endothelial cells production of cytokines (Khalfoun et al., 1997), and enhanced nitric oxide production by endothelial cells (Okuda et al., 1997, Omura et al., 2001). However, the intracellular signaling mechanisms for inhibiting endothelial activation by omega-3 PUFA remain unclear.

Many studies have shown that the mitogen-activated protein kinases (MAPK) are important signal enzymes witch involved in many facets of cellular regulation. The MAPK cascade is a major signaling system by which cells transduce an extracellular stimulus into an intracellular response (Chang and Karin, 2001, Cowan and Storey, 2003). The key role of MAPK in endothelial activation by proinflammatory cytokines has been shown. While the extracellular signal-related kinases (ERK1/2) are primarily involved in growth and cytoprotective functions, Jun amino-terminal kinases (JNK) and p38 proteins play an important role in inflammatory and stress responses (Hoefen and Berk, 2002). Several studies have shown that EPA and DHA modulated MAPK signaling activity in T cell (Denys et al., 2001), macrophage (Lo et al., 2000), neuron (Mirnikjoo et al., 2001) and myocyte (Chen et al., 2003b).

Tumor necrosis factor-alpha (TNF-α) is the prototypic proinflammatory cytokine and endothelial cells are the principal cellular targets of its actions. TNF-α-mediated endothelial activation is a central event in inflammation (Pober, 2002). Recently, studies have shown that superoxide dismutase (SOD) (Lin et al., 2005) and fluid shear stress (Yamawaki et al., 2003) can inhibit TNF-α-induced proinflammatory responses by modulating MAPK signaling pathways in endothelial cells.

Thus, we hypothesized that EPA and DHA may modulate TNF-α-mediated endothelial activation by altering the MAPK signaling activity. In the present study, we have analyzed the role of EPA and DHA in MAPK cascades transcription and activation in TNF-α-stimulated human umbilical vein endothelial cells (HUVEC). We show here for the first time, to our knowledge, that both EPA and DHA alone diminish activation of the p38 and JNK pathway, while maintaining the activation of ERK1/2 pathway of TNF-α-stimulated HUVEC. This may contribute to the inhibiting effects of omega-3 PUFA on endothelial activation by proinflammatory stimuli.