"Consulted to death": Personal stress as a major barrier to environmental co-management.

Co-management is widely seen as a way of improving environmental governance and empowering communities. When successful, co-management enhances the validity and legitimacy of decision-making, while providing stakeholders with influence over processes and outcomes that directly impact them. However, our research with participants in co-management across several cases leads us to argue that many of the individuals who contribute to co-management are subject to significant personal stress arising from both the logistical and social/emotional demands of participation in these processes. We argue that the literature on co-management has touched on this only indirectly, and that personal stress is a major challenge for participants that ought to be integrated into research agendas and addressed by policy-makers. In this article, we review the contours of the personal stress issue as it has appeared in our observations of co-management events and interviews with participants. While these findings are partial and preliminary, we argue that personal stress has theoretical and practical significance to the broader literature and process design. We conclude the article with recommendations for participants, researchers and policy-makers about how to consider and respond to problems of personal stress.

Omega-3 Monoacylglyceride Effects on Longevity, Mitochondrial Metabolism and Oxidative Stress: Insights from Drosophila melanogaster.

During the last decade, essential polyunsaturated fatty acids (PUFAs) such as eicosatetraenoic acid (EPA) and docosahexaenoic acid (DHA) derived from marine sources have been investigated as nonpharmacological dietary supplements to improve different pathological conditions, as well as aging. The aim of this study was to determine the effects of dietary n-3 PUFA monoacylglycerides (MAG, both EPA and DHA) on the mitochondrial metabolism and oxidative stress of a short-lifespan model, Drosophila melanogaster, sampled at five different ages. Our results showed that diets supplemented with MAG-EPA and MAG-DHA increased median lifespan by 14.6% and decreased mitochondrial proton leak resulting in an increase of mitochondrial coupling. The flies fed on MAG-EPA also had higher electron transport system capacity and mitochondrial oxidative capacities. Moreover, both n-3 PUFAs delayed the occurrence of lipid peroxidation but only flies fed the MAG-EPA diet showed maintenance of superoxide dismutase activity during aging. Our study therefore highlights the potential of n-3 PUFA monoacylglycerides as nutraceutical compounds to delay the onset of senescence by acting directly or indirectly on the mitochondrial metabolism and suggests that Drosophila could be a relevant model for the study of the fundamental mechanisms linking the effects of n-3 PUFAs to aging.

The Efficacy of MAG-DHA for Correcting AA/DHA Imbalance of Cystic Fibrosis Patients.

Omega-3 polyunsaturated fatty acid (n-3 PUFA) supplementations are thought to improve essential fatty acid deficiency (EFAD) as well as reduce inflammation in Cystic Fibrosis (CF), but their effectiveness in clinical studies remains unknown. The aim of the study was to determine how the medical food containing docosahexaenoic acid monoglyceride (MAG-DHA) influenced erythrocyte fatty acid profiles and the expression levels of inflammatory circulating mediators. We conducted a randomized, double blind, pilot trial including fifteen outpatients with Cystic Fibrosis, ages 18⁻48. The patients were divided into 2 groups and received MAG-DHA or a placebo (sunflower oil) for 60 days. Patients took 8 × 625 mg MAG-DHA softgels or 8 × 625 mg placebo softgels every day at bedtime for 60 days. Lipid analyses revealed that MAG-DHA increased docosahexaenoic acid (DHA) levels and decrease arachidonic acid (AA) ratio (AA/DHA) in erythrocytes of CF patients following 1 month of daily supplementation. Data also revealed a reduction in plasma human leukocyte elastase (pHLE) complexes and interleukin-6 (IL-6) expression levels in blood samples of MAG-DHA supplemented CF patients. This pilot study indicates that MAG-DHA supplementation corrects erythrocyte AA/DHA imbalance and may exert anti-inflammatory properties through the reduction of pHLE complexes and IL6 in blood samples of CF patients. TRIAL REGISTRATION: Pro-resolving Effect of MAG-DHA in Cystic Fibrosis (PREMDIC), NCT02518672.

Docosahexaenoic Acid Monoglyceride Increases Carboplatin Activity in Lung Cancer Models by Targeting EGFR.

BACKGROUND/AIM: Omega 3 polyunsaturated fatty acids (PUFAs) have been shown to inhibit the induction and progression of many tumor types. The aim of this study was to determine the anticancer effect of docosahexaenoic acid monoglyceride (MAG-DHA) alone and in combination with the chemotherapeutic agent carboplatin (CBT) on lung cancer models. MATERIALS AND METHODS: Adenocarcinoma cell lines A549 and H1299 were used to evaluate the effect of combined MAG-DHA and CBT treatments both in vitro and in vivo in xenograft models. RESULTS: MAG-DHA+CBT treatment decreased cell proliferation and invasion abilities of A549 and H1299 cells. Furthermore, MAG-DHA+CBT treatment resulted in a decreased activation of epithelial growth factor receptor (EGFR) and its downstream extracellular signal-regulated kinase (ERK) in cell lysates. In A549 and H1299 xenograft mouse models, MAG-DHA+CBT treatment reduced tumor growth. CONCLUSION: Combined MAG-DHA and CBT treatment inhibited tumor growth by suppressing EGFR and ERK signaling pathways in lung carcinoma cells.

Potential Application of Eicosapentaenoic Acid Monoacylglyceride in the Management of Colorectal Cancer.

BACKGROUND: There is increasing evidence that marine omega-3 oils are involved in the reduction of cancer risk and progression. However, the anticancer effect of omega-3 monoglyceride on colorectal cancer has yet to be assessed. The goal of this study was to evaluate the anti-cancer effects of eicosapentaenoic acid monoglyceride (MAG-EPA) in HCT116 colorectal carcinoma cells. METHODS: The effect of MAG-EPA was evaluated in vitro on HCT116 cells and in vivo on mouse model of HCT116 xenograft. RESULTS: Our data reveal that MAG-EPA decreased cell proliferation and induced apoptosis in HCT116 cells. In a xenograft mouse model, daily per os administration of MAG-EPA reduced tumor growth. Furthermore, MAG-EPA treatments decreased EGFR, VEGFR, and AKT activation pathways and reduced VEGF and HIF1α expression levels in tumors. CONCLUSION: MAG-EPA may promote apoptosis and inhibit growth of tumors by suppressing EGFR and VEGFR activation pathways. Altogether, these data provide new evidence regarding the mode of action of MAG-EPA in colorectal cancer cells.

Eicosapentaenoic acid monoglyceride resolves inflammation in an ex vivo model of human peripheral blood mononuclear cell.

Phosphorylation and activation of p38 MAPK and NFκB pathways, along with the resulting overproduction of interleukin IL-1ß, IL-6, and tumor necrosis factor a (TNFα) is a hallmark of inflammatory disorders. Omega-3 polyunsaturated fatty acid (n-3 PUFA) supplementations are known to exert anti-inflammatory properties by reduction of keys cytokines and enzymes involved in inflammation. Here, we investigated the anti-inflammatory pathways and mediators modulated by eicosapentaenoic acid monoglyceride (MAG-EPA) on human peripheral blood mononuclear cells (PBMCs) from healthy donors and stimulated, ex vivo, with lipopolysaccharide (LPS). LPS stimulation increased p38 MAPK and NFκB phosphorylation, which was abolished by MAG-EPA treatments. Concomitantly, MAG-EPA also abolished LPS-induced inflammation in PBMCs by reducing IL-1ß, IL-6, and TNFα cytokines at protein and transcript levels. Moreover, MAG-EPA decreased the levels of HIF1α in LPS-induced human PBMCs. Results also revealed a decreased of pro-inflammatory enzymes such as Cyclooxygenase-2 (COX-2) and 5-lipoxygenase (5-LOX) in LPS-induced PBMCs. Altogether, the present data suggest that MAG-EPA, represents a new potential therapeutic strategy for resolving inflammation in inflammatory disorders including autoimmune diseases, allergies, asthma, arthritis and cancer.

Comparison of the effects of EPA and DHA alone or in combination in a murine model of myocardial infarction.

The aim of this project was to investigate the impact of two dietary omega-3 polyunsaturated fatty acids (PUFAs), eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA), alone or in combination, on infarct size. Adult, male Sprague-Dawley rats were fed for 14 days with different omega-3 diets. The animals were subjected to ischemia for 40min followed by reperfusion. Infarct size, Akt (protein kinase B) activation level, caspase-3 activity and mitochondrial permeability transition pore (mPTP) opening were measured. The results indicate that EPA or DHA alone significantly reduced infarct size compared to the other diets. Akt activity was increased in the group fed EPA or DHA alone, whereas no significant activation was observed in the other groups compared to no omega-3 PUFA. DHA alone reduced caspase-3 activity and conferred resistance to mPTP opening. In conclusion, our results demonstrate that EPA and DHA are individually effective in diminishing infarct size in our experimental model while their combination is not.

MAG-EPA reduces severity of DSS-induced colitis in rats.

Ulcerative colitis (UC) is a chronic disease characterized by diffuse inflammation of the intestinal mucosa of the large bowel. Omega-3 (ω3) fatty acid supplementation has been associated with a decreased production of inflammatory cytokines involved in UC pathogenesis. The aim of this study was to determine the preventive and therapeutic potential of eicosapentaenoic acid monoglyceride (MAG-EPA) in an in vivo rats model of UC induced by dextran sulfate sodium (DSS). DSS rats were untreated or treated per os with MAG-EPA. Morphological, histological, and biochemical analyses were performed following MAG-EPA administrations. Morphological and histological analyses revealed that MAG-EPA pretreatment (12 days pre-DSS) and treatment (6 days post-DSS) exhibited strong activity in reducing severity of disease in DSS rats. Following MAG-EPA administrations, tissue levels of the proinflammatory cytokines TNF-α, IL-1ß, and IL-6 were markedly lower compared with rats treated only with DSS. MAG-EPA per os administration decrease neutrophil infiltration in colon tissues, as depicted by myelohyperoxidase activity. Results also revealed a reduced activation of NF-κB pathways correlated with a decreased expression of COX-2 in colon homogenates derived from MAG-EPA-pretreated and treated rats. Tension measurements performed on colon tissues revealed that contractile responses to methacholine and relaxing effect induced by sodium nitroprusside were largely increased following MAG-EPA treatment. The combined treatment of MAG-EPA and vitamin E displayed an antagonistic effect on anti-inflammatory properties of MAG-EPA in DSS rats.

Resolvin E1 normalizes contractility, Ca2+ sensitivity and smooth muscle cell migration rate in TNF-α- and IL-6-pretreated human pulmonary arteries.

Pulmonary hypertension (PH) is a rare disease in which pathophysiology is characterized by an increase in proinflammatory mediators, chronic endothelial dysfunctions, and a high migration rate of smooth muscle cells (SMC). Over the course of the last decade, various treatments have been proposed to relax the pulmonary arteries, none of which have been effective in resolving PH. Our hypothesis is that artery-relaxing drugs are not the long-term solution, but rather the inhibition of tissue inflammation, which underlies human pulmonary artery (HPA) dysfunctions that lead to abnormal vasoconstriction. The goal of the present study was to assess the anti-inflammatory effects of resolvin E1 (RvE1) with concomitant effects on SMC migration and on HPA reactivity. The role and mode of action of RvE1 and its precursor, monoacylglyceride eicosapentaenoic acid were assessed on HPA under proinflammatory conditions, involving a combined pretreatment with 10 ng/ml TNF-α and 10 ng/ml IL-6. Our results show that TNF-α and IL-6 treatment induced hyperreactivity and Ca(2+) hypersensitivity in response to pharmaco-mechanical stimuli, including 80 mM KCl, 1 µM phorbol 12-13-dibutyrate, and 30 nM U-46619. Furthermore, the proinflammatory treatment increased the migration rate of SMC isolated from HPA. The phosphorylation level of regulatory contractile proteins (CPI-17, MYPT-1), and proinflammatory signaling pathways (c-Fos, c-Jun, NF-κB) were also significantly increased compared with control conditions. Conversely, 300 nM RvE1 was able to normalize all of the above abnormal events triggered by proinflammation. In conclusion, RvE1 can resolve human arterial hyperreactivity via the resolution of inflammatory markers.

Eicosapentaenoic acid and docosapentaenoic acid monoglycerides are more potent than docosahexaenoic acid monoglyceride to resolve inflammation in a rheumatoid arthritis model.

INTRODUCTION: Rheumatoid arthritis (RA) is a chronic inflammatory autoimmune disease of the joints and bones. Omega-3 (ω3) fatty acid supplementation has been associated with a decreased production of inflammatory cytokines and eicosanoids involved in RA pathogenesis. The aim of this study was to determine the therapeutic potential of ω3 monoglyceride (MAG-ω3) compounds in an in vivo rat model of RA induced by Complete Freund's Adjuvant (CFA). METHOD: CFA rats were untreated or treated per os with three specific compounds, namely, MAG-docosahexaenoic acid (MAG-DHA), MAG-eicosapentaenoic acid (MAG-EPA) and MAG-docosapentaenoic acid (MAG-DPA). Morphological and histological analyses, as well as pro-inflammatory marker levels were determined following MAG-ω3 treatments. RESULTS: Morphological and histological analyses revealed that MAG-EPA and MAG-DPA exhibited strong activity in reducing the progression and severity of arthritic disease in CFA rats. Following MAG-EPA and MAG-DPA treatments, plasma levels of the pro-inflammatory cytokines; interleukin 17A (IL-17A), IL-1ß, IL-6 and tumor necrosis factor α (TNFα) were markedly lower when compared to CFA-untreated rats. Results also revealed a decreased activation of p38 mitogen-activated protein kinases (p38 MAPK) and nuclear factor-kappa B (NFκB) pathways correlated with a reduced expression of TNFα, cyclooxygenase-2 (COX-2), matrix metalloproteinase-2 (MMP-2) and MMP-9 in paw homogenates derived from MAG-EPA and MAG-DPA-treated rats. Of interest, the combined treatment of MAG-EPA and vitamin E displayed an antagonistic effect on anti-inflammatory properties of MAG-EPA in CFA rats. CONCLUSION: Altogether, the present data suggest that MAG-EPA, without vitamin E, represents a new potential therapeutic strategy for resolving inflammation in arthritis.

Effect of docosahexaenoic acid monoacylglyceride on systemic hypertension and cardiovascular dysfunction.

ω-3 Fatty acid supplementation has been associated with lower blood pressure. Cardiovascular diseases are also known to be linked directly to an increase in ω-6 and a reduction in ω-3 fatty acid levels in blood circulation and tissues. To determine the effect of docosahexaenoic acid monoglycerides (MAG-DHA) on blood pressure, lipid profiles, and vascular remodeling in rats fed a high-fat/high-carbohydrate (HFHC) diet. Studies were performed in male rats subjected to 8 wk of HFHC diet supplemented or not with 3 g/day MAG-DHA. After 8 wk of daily MAG-DHA treatment, rats in the HFHC + MAG-DHA group had lower arterial blood pressure and heart rate compared with the HFHC group. Moreover, MAG-DHA prevented the increase aortic wall thickness, whereas lipid analysis of aortic tissues revealed an increase in DHA/AA ratio correlated with the production of resolvin D2 and D3 metabolites. Histological analysis revealed that MAG-DHA prevented the development of LVH in the HFHC group. Serum lipid profile analysis further showed a decrease in total cholesterol (TC) and LDL, including very low-density lipoprotein (VLDL) and triglyceride (TG) levels, together with an increase in HDL levels after 8 wk of MAG-DHA treatment compared with the HFHC group. Furthermore, daily MAG-DHA treatment resulted in reduced proinflammatory marker levels such as CRP, IL-6, TNFα, and IL-1ß. Altogether, these findings revealed that per os administration of MAG-DHA prevents HFHC-diet induced hypertension and LVH in rats.

Proresolving Action of Docosahexaenoic Acid Monoglyceride in Lung Inflammatory Models Related to Cystic Fibrosis.

Cystic fibrosis (CF) is a hereditary, chronic disease of the exocrine glands, characterized by the production of viscid mucus that obstructs the pancreatic ducts and bronchi, leading to infection and fibrosis. ω3 fatty acid supplementations are known to improve the essential fatty acid deficiency as well as reduce inflammation in CF. The objective of this study was to determine the effects of docosahexaenoic acid monoacylglyceride (MAG-DHA) on mucin overproduction and resolution of airway inflammation in two in vitro models related to CF. Isolated human bronchi reverse permeabilized with CF transmembrane conductance regulator (CFTR) silencing (si) RNA and stable Calu3 cells expressing a short hairpin (sh) RNA directed against CFTR (shCFTR) were used. Lipid analyses revealed that MAG-DHA increased DHA/arachidonic acid (AA) ratio in shCFTR Calu-3 cells. MAG-DHA treatments, moreover, resulted in a decreased activation of Pseudomonas aeruginosa LPS-induced NF-κB in CF and non-CF Calu-3 cells. Data also revealed a reduction in MUC5AC, IL-6, and IL-8 expression levels in MAG-DHA-treated shCFTR cells stimulated, or not, with LPS. Antiinflammatory properties of MAG-DHA were also investigated in a reverse-permeabilized human bronchi model with CFTR siRNA. After MAG-DHA treatments, messenger RNA transcript levels for MUC5AC, IL-6, and IL-8 were markedly reduced in LPS-treated CFTR siRNA bronchi. MAG-DHA displays antiinflammatory properties and reduces mucin overexpression in Calu-3 cells and human bronchi untreated or treated with P. aeruginosa LPS, a finding consistent with the effects of resolvinD1, a known antiinflammatory mediator.

One-pot synthesis of polyunsaturated fatty acid amides with anti-proliferative properties.

A one-pot environmentally friendly transamidation of ω-3 fatty acid ethyl esters to amides and mono- or diacylglycerols was investigated via the use of a polymer-supported lipase. The method was used to synthesize a library of fatty acid monoglyceryl esters and amides. These new derivatives were found to have potent growth inhibition effects against A549 lung cancer cells.

Resolvin D1 reverses reactivity and Ca2+ sensitivity induced by ET-1, TNF-α, and IL-6 in the human pulmonary artery.

Pulmonary hypertension (PH) is a rare and progressive disease characterized by an inflammatory status and vessel wall remodeling, resulting in increased pulmonary artery resistance. During the last decade, treatments have been proposed; most of them target the endothelial pathways that stimulate smooth muscle cell relaxation. However, PH remains associated with significant morbidity. We hypothesized that inflammation plays a crucial role in the severity of the abnormal vasoconstriction in PH. The goal of this study was to assess the effects of resolvin D1 (RvD1), a potent anti-inflammatory agent, on the pharmacological reactivity of human pulmonary arteries (HPAs) via an in vitro model of induced hyperreactivity. The effects of RvD1 and monoacylglyceride compounds were measured on contractile activity and Ca(2+) sensitivity developed by HPAs that had been pretreated (or not) under proinflammatory conditions with either 10 ng/ml TNF-α or 10 ng/ml IL-6 or under hyperreactive conditions with 5 nM endothelin-1. The results demonstrated that, compared with controls, 24-h pretreatment with TNF-α, IL-6, or endothelin-1 increased reactivity and Ca(2+) sensitivity of HPAs as revealed by agonist challenges with 80 mM KCl, 1 µM serotonin (5-hydroxytryptamine), 30 nM U-46619, and 1 µM phorbol 12,13-dibutyrate. However, 300 nM RvD1 as well as 1 µM monoacylglyceride-docosapentaenoic acid monoglyceride strongly reversed the overresponsiveness induced by both proinflammatory and hyperreactive treatments. In pretreated pulmonary artery smooth muscle cells, Western blot analyses revealed that RvD1 treatment decreased the phosphorylation level of CPI-17 and expression of transmembrane protein member 16A while increasing the detection of G protein-coupled receptor 32. The present data demonstrate that RvD1, a trihydroxylated docosahexaenoic acid derivative, decreases induced overreactivity in HPAs via a reduction in CPI-17 phosphorylation and transmembrane protein member 16A expression.

Docosapentaenoic acid monoacylglyceride reduces inflammation and vascular remodeling in experimental pulmonary hypertension.

n-3 Polyunsaturated fatty acids (n-3 PUFA) have been shown to reduce inflammation and proliferation of pulmonary artery smooth muscle cells under pathophysiological conditions. However, the anti-inflammatory effect of the newly synthesized docosapentaenoic acid monoacylglyceride (MAG-DPA) on key signaling pathways in pulmonary hypertension (PH) pathogenesis has yet to be assessed. The aim of the present study was to determine the effects of MAG-DPA on pulmonary inflammation and remodeling occurring in a rat model of PH, induced by a single injection of monocrotaline (MCT: 60 mg/kg). Our results demonstrate that MAG-DPA treatment for 3 wk following MCT injection resulted in a significant improvement of right ventricular hypertrophy (RVH) and a reduction in Fulton's Index (FI). Morphometric analyses revealed that the wall thickness of pulmonary arterioles was significantly lower in MCT + MAG-DPA-treated rats compared with controls. This result was further correlated with a decrease in Ki-67 immunostaining. Following MAG-DPA treatments, lipid analysis showed a consistent increase in DPA together with lower levels of arachidonic acid (AA), as measured in blood and tissue samples. Furthermore, in MCT-treated rats, oral administration of MAG-DPA decreased NF-κB and p38 MAPK activation, leading to a reduction in MMP-2, MMP-9, and VEGF expression levels in lung tissue homogenates. Altogether, these data provide new evidence regarding the mode of action of MAG-DPA in the prevention of pulmonary hypertension induced by MCT.

Anti-proliferative effects of a new docosapentaenoic acid monoacylglyceride in colorectal carcinoma cells.

N-3 polyunsaturated fatty acids (n-3 PUFAs) have been shown to inhibit the induction and progression of many tumor types. However, the anticancer effect of n-3 PUFA monoglyceride on colorectal cancer has yet to be assessed. The aim of the present study was to determine the anti-tumorigenic effects of docosahexaenoic acid monoglyceride (MAG-DHA), eicosapentaenoic acid monoglyceride (MAG-EPA) and docosapentaenoic acid (22:5n-3) monoglyceride (MAG-DPA) in colorectal carcinoma cells. Our results demonstrate that MAG-DHA, MAG-EPA and MAG-DPA all decreased cell proliferation and induced apoptosis in HCT116 cells, with MAG-DPA having the higher anti-proliferative and pro-apoptotic effects in vitro. In a HCT116 xenograft mouse model, oral administration of MAG-DPA significantly inhibited tumor growth. Furthermore, MAG-DPA treatments decreased NFκB activation leading to a reduction in Bcl-2, CyclinD1, c-myc, COX-2, MMP9 and VEGF expression levels in tumor tissue sections. Altogether, these data provide new evidence regarding the mode of action of MAG-DPA in colorectal cancer cells.

Anti-cancer effects of a new docosahexaenoic acid monoacylglyceride in lung adenocarcinoma.

Lung cancer is the leading cause of cancer-related deaths worldwide. Despite advances in research, diagnosis and treatment, lung cancer remains a highly lethal disease, often diagnosed at advanced stages and with a very poor prognosis. Therefore, new strategies for the prevention and treatment of lung cancer are urgently needed. The aim of the present study was to determine the anti-tumorigenic effects of docosahexaenoic acid monoacylglyceride (MAG-DHA), a newly patented DHA derivative in lung adenocarcinoma. Our results demonstrate that MAG-DHA treatments decreased cell proliferation and induced apoptosis in A549 human lung carcinoma cells whereas MAG-DHA treatment did not induce apoptosis of normal bronchial epithelial BEAS-2B cells. MAG-DHA decreased NFκB activation leading to a reduction in COX-2 expression level in both A549 cells and lung adenocarcinoma tissues. Furthermore, MAG-DHA treatment increased PTEN expression and activation concomitant with a decrease in AKT phosphorylation levels and enhanced apoptosis. Oral administration of MAG-DHA significantly reduced tumor growth in a mouse A549 xenograft model. Lastly, MAG-DHA markedly decreased COX-2 and enhanced PTEN protein expression in tumor tissue sections. Altogether, these data provide new evidence regarding the mode of action of MAG-DHA and strongly suggest that this compound could be of clinical interest in cancer treatment.

Docosahexaenoic acid monoacylglyceride decreases endothelin-1 induced Ca(2+) sensitivity and proliferation in human pulmonary arteries.

BACKGROUND: Pulmonary artery vasoconstriction and vascular remodeling contribute to a sustained elevation of pulmonary vascular resistance and pressure in patients with pulmonary arterial hypertension (PH), an often fatal hemodynamic disease. The effect of docosahexaenoic acid monoacylglyceride (MAG-DHA) and the role of the 17 kDa protein kinase C-potentiated inhibitor protein (CPI-17) were determined on vasoconstriction and smooth muscle cell proliferation of human pulmonary arteries (HPA). METHODS: HPA were obtained from 16 patients undergoing lung resection for carcinoma. The mechanical tension and Ca(2+) sensitivity were measured on arterial rings treated with endothelin-1 (ET-1) in the absence or presence of MAG-DHA. The effect of MAG-DHA on the level of proliferation of smooth muscle cells isolated from HPA was evaluated in order to determine the role of CPI-17 protein. RESULTS: MAG-DHA treatment decreased the reactivity and Ca(2+) sensitivity induced by ET-1 in HPA. MAG-DHA treatment also decreased the expression of vascular endothelial growth factor (VEGF) induced by ET-1. Moreover, both VEGF inhibitor and MAG-DHA treatments reduced Ca(2+) hypersensitivity induced by ET-1, which was associated to a reduction in CPI-17 and myosin-binding subunit of the myosin light chain phosphatase (MYPT-1) phosphorylation levels. Proliferation of ET-1-stimulated HPA smooth muscle cells (PASMc) was also decreased following CPI-17 small interfering RNA transfection and MAG-DHA treatments. Western blot analyses revealed that MAG-DHA treatment resulted in decreased phosphorylation levels of CPI-17 and extracellular signal-regulated kinases (ERK) in PASMc treated with ET-1. CONCLUSIONS: We have demonstrated that VEGF interacts with CPI-17 signaling pathway resulting in an increase in Ca(2+) sensitivity and proliferation of PASMc, whereas MAG-DHA treatment reversed these effects.

Bronchial inflammation induced PKCζ over-expression: involvement in mechanical properties of airway smooth muscle.

Protein kinase C variants (PKCs) have been involved in the control of airway smooth muscle (ASM) tone, and abnormalities in PKC-dependent signaling have been associated with respiratory diseases such as asthma. In this study, the role of atypical PKCζ in airway hyperresponsiveness was investigated, using an in-vitro model of TNFα-treated human bronchi and an in vivo guinea pig model of chronic asthma. Our results demonstrated that PKCζ-specific inhibition produced a significant increase in isoproterenol sensitivity in TNFα-treated bronchi and ovalbumin (OVA)-sensitized guinea pig bronchi. The role of epoxy-eicosanoids, known to exert anti-inflammatory effects in lung, on PKCζ expression and activity in these models was evaluated. An enhanced PKCζ protein expression was delineated in TNFα-treated bronchi when compared with control (untreated) and epoxy-eicosanoid-treated bronchi. Measurements of Ca(2+) sensitivity, performed in TNFα-treated bronchi, demonstrated that treatment with myristoylated (Myr) PKCζ peptide inhibitor resulted in significant reductions of pCa-induced tension. Epoxy-eicosanoid treatments had similar effects on Ca(2+) sensitivity in TNFα-treated bronchi. In control and epoxy-eicosanoid-treated bronchi, the phosphorylated forms of p38MAPK and CPI-17 were significantly decreased compared with the TNFα-treated bronchi. An enhanced expression of PKCζ was ascertained in our in-vivo model of allergic asthma. Hence an increased Ca(2+) sensitivity could be explained by the phosphorylation of p38-MAPK, which in turn leads to phosphorylation and activation of the CPI-17 regulatory protein. This process was reversed upon treatment with the Myr-PKCζ-peptide inhibitor. The present data provide relevant evidence regarding the role of PKCζ in human and rodent models of airways inflammation.

19,20-EpDPE, a bioactive CYP450 metabolite of DHA monoacyglyceride, decreases Ca²âº sensitivity in human pulmonary arteries.

The aim of this study was to investigate the effect of docosahexaenoic acid monoacylglyceride (MAG-DHA) on human pulmonary arterial tone. Tension measurements on pulmonary arterial tissues demonstrated that MAG-DHA reduced U-46619-induced tone, which is highly sensitive to the H-1152 inhibitor. Results also showed that MAG-DHA treatments decreased RhoA activity levels, which in turn inactivated the Rho-kinase pathway, leading to a reduction in U-46619-induced Ca(2+) sensitivity of permeabilized pulmonary artery smooth muscle cells. According to the mechanical responses assessing U-46619-induced Ca(2+) sensitivity in the absence or presence of 3 µM MAG-DHA, MAG-DHA plus 1 µM N-methylsulfonyl-6-(2-propargyloxyphenyl) hexanamide (MS-PPOH, a cytochrome P-450 epoxygenase inhibitor) and 300 nM 19,20-epoxydocosapentaenoic acid (a cytochrome P-450 epoxygenase-dependent DHA metabolite), our data suggest that the MAG-DHA is metabolized in a bioactive epoxymetabolite. This epoxyeicosanoid in turn decreases active tone and Ca(2+) sensitivity of smooth muscles cells through an inhibition of the Rho-kinase pathway. Together, these data provide primary evidence regarding the mode of action of MAG-DHA in human pulmonary arteries and suggest that this compound may be of pharmacological interest in patients with pulmonary hypertension to generate intracellular bioactive metabolites.