[show abstract][hide abstract] ABSTRACT: Pharmacological treatments targeting CXC chemokines and the associated neutrophil activation and recruitment into atherosclerotic plaques hold promise for treating cardiovascular disorders. Therefore, we investigated whether FK866, a nicotinamide phosphoribosyltransferase (NAMPT) inhibitor with anti-inflammatory properties that we recently found to reduce neutrophil recruitment into the ischaemic myocardium, would exert beneficial effects in a mouse atherosclerosis model. Atherosclerotic plaque formation was induced by carotid cast implantation in ApoE-/- mice that were fed with a Western-type diet. FK866 or vehicle were administrated intraperitoneally from week 8 until week 11 of the diet. Treatment with FK866 reduced neutrophil infiltration and MMP-9 content and increased collagen levels in atherosclerotic plaques compared to vehicle. No effect on other histological parameters, including intraplaque lipids or macrophages, was observed. These findings were associated with a reduction in both systemic and intraplaque CXCL1 levels in FK866-treated mice. In vitro, FK866 did not affect MMP-9 release by neutrophils, but it strongly reduced CXCL1 production by endothelial cells which, in the in vivo model, were identified as a main CXCL1 source at the plaque level. CXCL1 synthesis inhibition by FK866 appears to reflect interference with nuclear factor-κB signalling as shown by reduced p65 nuclear levels in endothelial cells pre-treated with FK866. In conclusion, pharmacological inhibition of NAMPT activity mitigates inflammation in atherosclerotic plaques by reducing CXCL1-mediated activities on neutrophils. These results support further assessments of NAMPT inhibitors for the potential prevention of plaque vulnerability.
Thrombosis and Haemostasis 11/2013; 111(2). · 6.09 Impact Factor
[show abstract][hide abstract] ABSTRACT: Myocardial reperfusion injury is mediated by several processes including increase of reactive oxygen species (ROS). The aim of the study is to identify potential sources of ROS contributing to myocardial ischemia-reperfusion injury. For this purpose, we investigated myocardial ischemia/reperfusion pathology in mice deficient in various NADPH oxidase isoforms (Nox1, Nox2, Nox4, as well as Nox1/2 double knockout). Following 30minutes of ischemia and 24hours reperfusion, a significant decrease in the size of myocardial infarct was observed in Nox1-, Nox2- and Nox1/Nox2-, but not in Nox4-deficient mice. However, no protection was observed in a model of chronic ischemia, suggesting that NOX1 and NOX2-mediated oxidative damage occurs during reperfusion. Cardioprotective effect of Nox1 and Nox2 deficiencies was associated with decrease of neutrophil invasion, but, on the other hand an improved reperfusion injury was also observed in isolated perfused hearts (Langendorff model) suggesting that inflammatory cells were not the major source of oxidative damage. A decrease in global post-reperfusion oxidative stress was clearly detected in Nox2-, but not in Nox1-deficient hearts. Analysis of key signalling pathways during reperfusion suggest distinct cardioprotective patterns: increased phosphorylation was seen for Akt and Erk in Nox1-deficient mice and for Stat3 and Erk in Nox2-deficient mice. Consequently, NOX1 and NOX2 represent interesting drug targets for controlling reperfusion damage associated with revascularization in coronary disease.
Journal of Molecular and Cellular Cardiology 09/2013; · 5.15 Impact Factor
[show abstract][hide abstract] ABSTRACT: Chemokines trigger leukocyte trafficking and are implicated in cardiovascular disease pathophysiology. Chemokine-binding proteins, called "Evasins" have been shown to inhibit both CC and CXC chemokine-mediated bioactivities. Here, we investigated whether treatment with Evasin-3 (CXC chemokine inhibitor) and Evasin-4 (CC chemokine inhibitor) could influence post-infarction myocardial injury and remodelling. C57Bl/6 mice were submitted in vivo to left coronary artery permanent ligature and followed up for different times (up to 21 days). After coronary occlusion, three intraperitoneal injections of 10 μg Evasin-3, 1 μg Evasin-4 or equal volume of vehicle (PBS) were performed at 5 minutes, 24 hours (h) and 48 h after ischaemia onset. Both anti-chemokine treatments were associated with the beneficial reduction in infarct size as compared to controls. This effect was accompanied by a decrease in post-infarction myocardial leukocyte infiltration, reactive oxygen species release, and circulating levels of CXCL1 and CCL2. Treatment with Evasin-4 induced a more potent effect, abrogating the inflammation already at one day after ischaemia onset. At days 1 and 21 after ischaemia onset, both anti-chemokine treatments failed to significantly improve cardiac function, remodelling and scar formation. At 21-day follow-up, mouse survival was exclusively improved by Evasin-4 treatment when compared to control vehicle. In conclusion, we showed that the selective inhibition of CC chemokines (i.e. CCL5) with Evasin-4 reduced cardiac injury/inflammation and improved survival. Despite the inhibition of CXC chemokine bioactivities, Evasin-3 did not affect mouse survival. Therefore, early inhibition of CC chemokines might represent a promising therapeutic approach to reduce the development of post-infarction heart failure in mice.
Thrombosis and Haemostasis 08/2013; 110(4). · 6.09 Impact Factor
[show abstract][hide abstract] ABSTRACT: Percutaneous transluminal angioplasty is frequently used in patients with severe arterial narrowing due to atherosclerosis. However, it induces severe arterial injury and an inflammatory response leading to restenosis. Here, we studied a potential activation of the endocannabinoid system and the effect of fatty acid amide hydrolase (FAAH) deficiency, the major enzyme responsible for endocannabinoid anandamide degradation, in arterial injury. We performed carotid balloon injury in atherosclerosis-prone apolipoprotein E knockout (ApoE-/-) and ApoE-/-FAAH-/- mice. Anandamide levels were systemically elevated in ApoE-/- mice after balloon injury. ApoE-/-FAAH-/- mice had significantly higher baseline anandamide levels and enhanced neointima formation compared to ApoE-/- controls. The latter effect was inhibited by treatment with CB1 antagonist AM281. Similarly, ApoE-/- mice treated with AM281 had reduced neointimal areas, reduced lesional vascular smooth muscle cell (SMC) content and proliferating cell counts. The lesional macrophage content was unchanged. In vitro proliferation rates were significantly reduced in CB1-/- SMCs or when treating ApoE-/- or ApoE-/-FAAH-/- SMCs with AM281. Macrophage in vitro adhesion and migration was marginally affected by CB1 deficiency. Reendothelialization was not inhibited by treatment with AM281. In conclusion, endogenous CB1 activation contributes to vascular SMC proliferation and neointima formation in response to arterial injury.
The Journal of Lipid Research 03/2013; · 4.39 Impact Factor
[show abstract][hide abstract] ABSTRACT: Neutrophilic inflammation might have a pathophysiological role in both carotid plaque rupture and ischemic stroke injury. Here, we investigated the potential benefits of the CXC chemokine-binding protein Evasin-3, which potently inhibits chemokine bioactivity and related neutrophilic inflammation in two mouse models of carotid atherosclerosis and ischemic stroke, respectively. In the first model, the chronic treatment with Evasin-3 as compared with Vehicle (phosphate-buffered saline (PBS)) was investigated in apolipoprotein E-deficient mice implanted of a 'cast' carotid device. In the second model, acute Evasin-3 treatment (5 minutes after cerebral ischemia onset) was assessed in mice subjected to transient left middle cerebral artery occlusion. Although CXCL1 and CXCL2 were upregulated in both atherosclerotic plaques and infarcted brain, only CXCL1 was detectable in serum. In carotid atherosclerosis, treatment with Evasin-3 was associated with reduction in intraplaque neutrophil and matrix metalloproteinase-9 content and weak increase in collagen as compared with Vehicle. In ischemic stroke, treatment with Evasin-3 was associated with reduction in ischemic brain neutrophil infiltration and protective oxidants. No other effects in clinical and histological outcomes were observed. We concluded that Evasin-3 treatment was associated with reduction in neutrophilic inflammation in both mouse models. However, Evasin-3 administration after cerebral ischemia onset failed to improve poststroke outcomes.Journal of Cerebral Blood Flow & Metabolism advance online publication, 19 December 2012; doi:10.1038/jcbfm.2012.198.
Journal of cerebral blood flow and metabolism: official journal of the International Society of Cerebral Blood Flow and Metabolism 12/2012; · 5.46 Impact Factor
[show abstract][hide abstract] ABSTRACT: OBJECTIVE: Endocannabinoid levels are elevated in human and mouse atherosclerosis, but their causal role is not well understood. Therefore, we studied the involvement of fatty acid amide hydrolase (FAAH) deficiency, the major enzyme responsible for endocannabinoid anandamide degradation, in atherosclerotic plaque vulnerability. METHODS AND RESULTS: We assessed atherosclerosis in apolipoprotein E-deficient (ApoE(-/-)) and ApoE(-/-)FAAH(-/-) mice. Before and after 5, 10, and 15 weeks on high-cholesterol diet, we analyzed weight, serum cholesterol, and endocannabinoid levels, and atherosclerotic lesions in thoracoabdominal aortas and aortic sinuses. Serum levels of FAAH substrates anandamide, palmitoylethanolamide (PEA), and oleoylethanolamide (OEA) were 1.4- to 2-fold higher in case of FAAH deficiency. ApoE(-/-)FAAH(-/-) mice had smaller plaques with significantly lower content of smooth muscle cells, increased matrix metalloproteinase-9 expression, and neutrophil content. Circulating and bone marrow neutrophil counts were comparable between both genotypes, whereas CXCL1 levels were locally elevated in aortas of FAAH-deficient mice. We observed enhanced recruitment of neutrophils, but not monocytes, to large arteries of ApoE(-/-) mice treated with FAAH inhibitor URB597. Spleens of ApoE(-/-)FAAH(-/-) mice had reduced CD4+FoxP3+regulatory T-cell content, and in vitro stimulation of splenocytes revealed significantly elevated interferon-γ and tumor necrosis factor-α production in case of FAAH deficiency. CONCLUSIONS: Increased anandamide and related FAAH substrate levels are associated with the development of smaller atherosclerotic plaques with high neutrophil content, accompanied by an increased proinflammatory immune response.
Arteriosclerosis Thrombosis and Vascular Biology 12/2012; · 6.34 Impact Factor
[show abstract][hide abstract] ABSTRACT: Cannabinoid receptor CB(2) activation inhibits inflammatory proliferation and migration of vascular smooth muscle cells in vitro. The potential in vivo relevance of these findings is unclear. We performed carotid balloon distension injury in hypercholesterolemic apolipoprotein E knockout (ApoE(-/-)) mice receiving daily intraperitoneal injection of the CB(2) agonist JWH133 (5 mg/kg) or vehicle, with the first injection given 30 min before injury. Alternatively, we subjected CB(2)(-/-) and wild-type (WT) mice to balloon injury. We determined CB(2) mRNA and protein expression in dilated arteries of ApoE(-/-) mice. Neointima formation was assessed histologically. We used bone marrow-derived murine CB(2)(-/-) and WT macrophages to study adhesion to plastic, fibronectin, or collagen, and migration was assayed by modified Boyden chamber. Aortic smooth muscle cells were isolated to determine in vitro proliferation rates. We found increased vascular CB(2) expression in ApoE(-/-) mice in response to balloon injury. Seven to twenty-one days after dilatation, injured vessels of JWH133-treated mice had less intimal nuclei numbers as well as intimal and medial areas, associated with less staining for proliferating cells, smooth muscle cells, and macrophages. Complete endothelial repair was observed after 14 days in both JWH133- and vehicle-treated mice. CB(2) deficiency resulted in increased intima formation compared with WT, whereas JWH133 did not affect intimal formation in CB(2)(-/-) mice. Apoptosis rates assessed by in situ terminal deoxynucleotidyl transferase dUTP-mediated nick-end labeling staining 1 h postballooning were significantly higher in the CB(2) knockouts. In vitro, bone marrow-derived CB(2)(-/-) macrophages showed enhanced adherence and migration compared with WT cells and elevated mRNA levels of adhesion molecules, chemokine receptors CCR1 and 5, and chemokine CCL2. Proliferation rates were significantly increased in CB(2)(-/-) smooth muscle cells compared with WT. In conclusion, pharmacological activation or genetic deletion of CB(2) receptors modulate neointima formation via protective effects in macrophages and smooth muscle cells.
[show abstract][hide abstract] ABSTRACT: The activation of cannabinoid receptor type 2 (CB(2))-mediated pathways might represent a promising anti-atherosclerotic treatment. Here, we investigated the expression of the endocannabinoid system in human carotid plaques and the impact of CB(2) pharmacological activation on markers of plaque vulnerability in vivo and in vitro.
The study was conducted using all available residual human carotid tissues (upstream and downstream the blood flow) from our cohort of patients symptomatic (n = 13) or asymptomatic (n = 27) for ischaemic stroke. Intraplaque levels of 2-arachidonoylglycerol, anandamide N-arachidonoylethanolamine, N-palmitoylethanolamine, N-oleoylethanolamine, and their degrading enzymes (fatty acid amide hydrolase and monoacylglycerol lipase) were not different in human plaque portions. In the majority of human samples, CB(1) (both mRNA and protein levels) was undetectable. In downstream symptomatic plaques, CB(2) protein expression was reduced when compared with asymptomatic patients. In these portions, CB(2) levels were inversely correlated (r = -0.4008, P = 0.0170) with matrix metalloprotease (MMP)-9 content and positively (r = 0.3997, P = 0.0174) with collagen. In mouse plaques, CB(2) co-localized with neutrophils and MMP-9. Treatment with the selective CB(2) agonist JWH-133 was associated with the reduction in MMP-9 content in aortic root and carotid plaques. In vitro, pre-incubation with JWH-133 reduced tumour necrosis factor (TNF)-α-mediated release of MMP-9. This effect was associated with the reduction in TNF-α-induced ERK1/2 phosphorylation in human neutrophils.
Cannabinoid receptor type 2 receptor is down-regulated in unstable human carotid plaques. Since CB(2) activation prevents neutrophil release of MMP-9 in vivo and in vitro, this treatment strategy might selectively reduce carotid vulnerability in humans.
European Heart Journal 11/2011; 33(7):846-56. · 14.10 Impact Factor
[show abstract][hide abstract] ABSTRACT: The highly prevalent obstructive sleep apnea syndrome (OSA) with its main component intermittent hypoxia (IH) is a risk factor for cardiovascular mortality. The poor knowledge of its pathophysiology has limited the development of specific treatments, whereas the gold standard treatment, continuous positive airway pressure, may not fully reverse the chronic consequences of OSA and has limited acceptance in some patients.
To examine the contribution of IH-induced inflammation to the cardiovascular complications of OSA. Methods: We investigated systemic and vascular inflammatory changes in C57BL6 mice exposed to IH (21-5% Fi(O(2)), 60-s cycle) or normoxia 8 hours per day up to 14 days. Vascular alterations were reassessed in mice treated with a blocking antibody of regulated upon activation, normal T-cell expressed and secreted (RANTES)/CC chemokine ligand 5 (CCL5) signaling pathway, or with the IgG isotype control throughout the IH exposure.
IH induced systemic inflammation combining increased splenic lymphocyte proliferation and chemokine expression, with early and predominant RANTES/CCL5 alterations, and enhanced splenocyte migration toward RANTES/CCL5. IH also induced structural and inflammatory vascular alterations. Leukocyte-endothelium adhesive interactions were increased, attested by leukocyte rolling and intercellular adhesion molecule-1 expression in mesenteric vessels. Aortas had increased intima-media thickness with elastic fiber alterations, mucoid depositions, nuclear factor-κB-p50 and intercellular adhesion molecule-1 overexpression, hypertrophy of smooth-muscle cells overexpressing RANTES/CCL5, and adventitial-periadventitial T-lymphocyte infiltration. RANTES/CCL5 neutralization prevented both intima-media thickening and inflammatory alterations, independently of the IH-associated proatherogenic dyslipidemia.
Inflammation is a determinant mechanism for IH-induced preatherosclerotic remodeling involving RANTES/CCL5, a key chemokine in atherogenesis. Characterization of the inflammatory response could allow identifying at-risk patients for complications, and its pharmacologic manipulation may represent a potential complementary treatment of sleep apnea consequences.
American Journal of Respiratory and Critical Care Medicine 06/2011; 184(6):724-31. · 11.04 Impact Factor
[show abstract][hide abstract] ABSTRACT: Although beneficial for cardiomyocyte salvage and to limit myocardial damage and cardiac dysfunction, restoration of blood flow after prolonged ischemia exacerbates myocardial injuries. Several deleterious processes that contribute to cardiomyocyte death have been proposed, including massive release of reactive oxygen species, calcium overload and hypercontracture development or leukocyte infiltration within the damaged myocardium. Chemokines are known to enhance leukocyte diapedesis at inflammatory sites. The aim of the present study was to investigate the effect of chemokine CCL5/RANTES antagonism in an in vivo mouse model of ischemia and reperfusion. ApoE(-/-) mice were submitted to 30 min ischemia, by ligature of the left coronary artery, followed by 24 h reperfusion. Intraperitoneal injection of 10 mug of CCL5/RANTES antagonist [(44)AANA(47)]-RANTES, 5 min prior to reperfusion, reduced infarct size as well as Troponin I serum levels compared to PBS-treated mice. This beneficial effect of [(44)AANA(47)]-RANTES treatment was associated with reduced leukocyte infiltration into the reperfused myocardium, as well as decreased chemokines Ccl2/Mcp-1 and Ccl3/Mip-1alpha expression, oxidative stress, and apoptosis. However, mice deficient for the CCL5/RANTES receptor Ccr5 did not exhibit myocardium salvage in our model of ischemia-reperfusion. Furthermore, [(44)AANA(47)]-RANTES did not mediate cardioprotection in these ApoE(-/-) Ccr5(-/-) deficient mice, probably due to enhanced expression of compensatory chemokines. This study provides the first evidence that inhibition of CCL5/RANTES exerts cardioprotective effects during early myocardial reperfusion, through its anti-inflammatory properties. Our findings indicate that blocking chemokine receptor/ligand interactions might become a novel therapeutic strategy to reduce reperfusion injuries in patients during acute coronary syndromes.
Journal of Molecular and Cellular Cardiology 09/2009; 48(4):789-98. · 5.15 Impact Factor
[show abstract][hide abstract] ABSTRACT: Plasma soluble inflammatory molecules are associated with the risk of ischaemic cardiovascular events. We investigated whether HIV replication modified the levels of these proteins in a combination antiretroviral therapy (cART) interruption trial.
In 145 HIV-infected Thai patients (62% women, median CD4 cell count 271 cells/microl, median plasma HIV-RNA 4.66 log10 copies/ml) included in the Swiss-Thai-Australia Treatment Interruption Trial (STACCATO) trial, leptin, adiponectin, C-reactive protein, soluble vascular cell adhesion molecule-1 (s-VCAM-1), P-selectin, chemokine ligand 2, chemokine ligand 3, interleukin (IL)-6, IL-10, granulocyte macrophage colony-stimulating factor and D-dimer were measured before cART was initiated, after cART had suppressed HIV replication to less than 50 copies/ml plasma (median 8 months) and again 12 weeks after randomization to continued cART (n=48) or interrupted cART (n=97). Multiple linear regression and logistic regression were used to investigate the association between each cardiovascular marker and plasma HIV-RNA. Initiation of cART resulted in significant declines in s-VCAM-1, P-selectin, leptin and D-dimer, whereas mediators with anti-inflammatory properties, such as adiponectin and IL-10, increased. At 12 weeks after randomization, we found positive associations between levels of s-VCAM-1 and chemokine ligand 2 with an increase in plasma HIV-RNA (r=0.271, P=0.001 and r=0.24, P=0.005, respectively), whereas levels of adiponectin decreased for each 1 log increase in plasma HIVRNA (r=-0.24, P=0.002). Detectable IL-10 was less likely (odds ratio = 0.64, 95% confidence interval = 0.43-0.96) for each 1 log increase in plasma HIV-RNA.
Plasma levels of several inflammatory, anti-inflammatory and endothelial activation markers of cardiovascular disease are associated with HIV-RNA replication.
AIDS (London, England) 06/2009; 23(8):929-39. · 4.91 Impact Factor
[show abstract][hide abstract] ABSTRACT: In this study we analysed the possible modulation of endocannabinoids and related molecules during atherosclerosis development in mice. Wild-type and apolipoprotein E knockout (ApoE(-/-)) mice were fed either normal chow or high-cholesterol diet for 8-12 weeks, and tissue endocannabinoid levels were measured by liquid chromatography-mass spectrometry. We found increased levels of 2-AG in aortas and visceral adipose tissue (VAT) of ApoE(-/-) mice fed on high-cholesterol diet for 12 weeks as compared to ApoE(-/-) mice fed on normal chow or wild-type mice fed on cholesterol. No significant difference in 2-AG levels was observed after 8 weeks of diet, and no changes in anandamide levels were found in any group. The levels of the anandamide-related mediators with anti-inflammatory or anti-lipogenic properties, palmitoylethanolamide (PEA) and oleoylethanolamide (OEA), decreased or increased only in VAT or in both tissues, respectively. Endocannabinoid- and OEA/PEA-degrading enzymes were expressed by macrophages within atherosclerotic lesions. In vitro, 2-AG and OEA-induced monocyte migration at 0.3-1microM, which corresponds to the levels observed in aortas. PEA 1microM also induced monocyte migration but counteracted the effect of 2-AG, whereas OEA enhanced it. Enhanced 2-AG levels in advanced atherosclerotic lesions may trigger the inflammatory process by recruiting more inflammatory cells and inducing extracellular matrix degradation via CB(2) receptors, and this possibility was supported in vitro but not in vivo by experiments with the CB(2) antagonist, SR144528.
[show abstract][hide abstract] ABSTRACT: Preventive treatment with cannabinoid agonists has been reported to reduce the infarct size in a mouse model of myocardial ischemia/reperfusion. Here we investigated the possible cardioprotective effect of selective CB(2) cannabinoid receptor activation during ischemia. We performed left coronary artery ligature in C57Bl/6 mice for 30 min, followed by 24 h of reperfusion. Five minutes before reperfusion, mice received intraperitoneal injection of the CB(2) selective agonist JWH-133 (20 mg/kg) or vehicle. Infarct size was assessed histologically and by cardiac troponin I (cTnI) ELISA. Immunohistochemical analysis of leukocyte infiltration, oxidative stress in situ quantification, real-time RT-PCR analysis of inflammatory mediators as well as western blots for kinase phosphorylation was also performed. In addition, we studied chemotaxis and integrin expression of human neutrophils in vitro. JWH-133 significantly reduced the infarct size (I/area at risk: 19.27%+/-1.91) as compared to vehicle-treated mice (31.77%+/-2.7). This was associated with a reduction of oxidative stress and neutrophil infiltration in the infarcted myocardium, whereas activation of ERK 1/2 and STAT-3 was increased. Preinjection of PI3K inhibitor LY294002, MEK 1/2 inhibitor U0126 and JAK-2 inhibitor AG-490 partially abrogated the JWH-133 mediated infarct size reduction. No changes in cardiac CXCL1, CXCL2, CCL3, TNF-alpha, and ICAM-1 expression levels were found. Furthermore, JWH-133 inhibited the TNF-alpha induced chemotaxis and integrin CD18/CD11b (Mac-1) upregulation on human neutrophils. Our data suggest that JWH-133 administration during ischemia reduces the infarct size in a mouse model of myocardial ischemia/reperfusion through a direct cardioprotective activity on cardiomyocytes and neutrophils.
Journal of Molecular and Cellular Cardiology 02/2009; 46(5):612-20. · 5.15 Impact Factor
[show abstract][hide abstract] ABSTRACT: We have recently shown that CRP induces chemokine secretion and adhesion molecule up-regulation in human primary monocytes cultured in adherence. Given the increasing evidence on direct immunomodulatory properties of statins, we investigated their possible anti-inflammatory role on CRP-treated human monocytes.
Monocytes were isolated by Ficoll-Percoll gradients and cultured in adherence to polystyrene. Chemokine secretion and adhesion molecule expression were detected by ELISA and flow cytometry. Migration assays were performed in modified Boyden chambers. Intracellular kinase activation was assessed by western blot.
Treatment with simvastatin or atorvastatin decreased CRP-induced release of CCL2, CCL3 and CCL4. In addition, both statins reduced CRP-induced intercellular adhesion molecule (ICAM-1) up-regulation, but had no effects on CD11b and CD18. Treatments with 1 microM simvastatin or atorvastatin significantly inhibited monocyte migration in response to CRP. CD32 and CD64 (CRP receptors) expression on monocytes was not affected by statins. Statin-induced inhibition of CRP-mediated chemokine secretion, ICAM-1 up-regulation and migration occurred through the inhibition of extracellular signal-regulated kinase (ERK) 1/2. Treatment with L-mevalonate or farnesylpyrophosphate, but not geranylgeranyl-pyrophosphate reversed the statin-induced effect on CRP-mediated functions and ERK 1/2 phosphorylation, confirming that statins blocked CRP-induced ERK 1/2 phosphorylation through the inhibition of 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase.
Statins inhibited CRP-induced chemokine secretion, ICAM-1 up-regulation and migration in human adherent monocytes, through the inhibition of HMG-CoA reductase-ERK 1/2 pathway. This pathway could represent a very promising target to reduce CRP-induced activities in monocyte-mediated diseases, such as atherosclerosis or RA.
[show abstract][hide abstract] ABSTRACT: Several studies support C-reactive protein (CRP) as a systemic cardiovascular risk factor. The recent detection of CRP in arterial intima suggests a dual activity in atherosclerosis as a circulating and tissue mediator on vascular and immune cells. In the present paper, we focused on the inflammatory effects of CRP on human monocytes, which were isolated by Ficoll-Percoll gradients and cultured in adherence to polystyrene, endothelial cell monolayer, or in suspension. Chemokine levels, adhesion molecule, and chemokine receptor expression were detected by ELISA, flow cytometry, and real-time RT-PCR. Migration assays were performed in a Boyden chamber. Stimulation with CRP induced release of CCL2, CCL3, and CCL4 in adherent monocytes through the binding to CD32a, CD32b, and CD64, whereas no effect was observed in suspension culture. This was associated with CRP-induced up-regulation of adhesion molecules membrane-activated complex 1 (Mac-1) and ICAM-1 on adherent monocytes. Blockade of Mac-1/ICAM-1 interaction inhibited the CRP-induced chemokine secretion. In addition, CRP reduced mRNA and surface expression of corresponding chemokine receptors CCR1, CCR2, and CCR5 in adherent monocytes. This effect was a result of chemokine secretion, as coincubation with neutralizing anti-CCL2, anti-CCL3, and anti-CCL4 antibodies reversed the effect of CRP. Accordingly, a reduced migration of CRP-treated monocytes to CCL2 and CCL3 was observed. In conclusion, our data suggest an in vitro model to study CRP activities in adherent and suspension human monocytes. CRP-mediated induction of adhesion molecules and a decrease of chemokine receptors on adherent monocytes might contribute to the retention of monocytes within atherosclerotic lesions and recruitment of other circulating cells.
Journal of Leukocyte Biology 07/2008; 84(4):1109-19. · 4.57 Impact Factor
[show abstract][hide abstract] ABSTRACT: Atherosclerosis is a chronic inflammatory disease that represents the primary cause of death through coronary disease and stroke. Chemokines are known to play a crucial role in this disease by recruiting inflammatory leukocytes to the endothelium. Recently, the chemokine variant [44AANA47]-RANTES was shown to impair inflammatory cell recruitment in vivo by interfering with heparin binding and oligomerization.
In this study we report that curative treatment with [44AANA47]-RANTES limits atherosclerotic plaque formation in LDLr-/- mice. This was associated with reduced infiltration of T cells and macrophages and reduced production of matrix metalloproteinase (MMP)-9. By contrast, the relative smooth muscle cell and collagen content was increased, indicating a more stable plaque phenotype. In addition, we provide evidence for direct inhibition of leukocyte recruitment into aortic root lesions, attenuated leukocyte rolling and arrest in mesenteric vessels, as well as a reduced proinflammatory response following Con A stimulation in vitro.
Interference with chemokine oligomerization and chemokine/heparin interactions is a powerful novel approach that inhibits progression of established atherosclerosis in mice. By inhibiting leukocyte recruitment into plaques, [44AANA47]-RANTES mediates a less inflammatory plaque phenotype and thus reduced systemic inflammatory state.
[show abstract][hide abstract] ABSTRACT: Strong evidence suggests that neutrophils may play an active role in acute and chronic inflammatory disorders, such as rheumatoid arthritis and atherosclerosis. Given the role of pro-inflammatory cytokine TNF-alpha in these inflammatory processes, we planned the present study to investigate the effect of short term incubation with TNF-alpha on neutrophil migration to CCL3, a chemokine produced in inflammatory sites and normally devoid of neutrophil chemotactic properties. We found that TNF-alpha primed neutrophils for migration to CCL3 via CCR5. TNF-alpha-induced migration was a consequence of the TNF-alpha-induced up-regulation of integrin CD11b/CD18 (Mac-1) on neutrophil surface. Furthermore, TNF-alpha activity was found to be strictly dependent on the activation of ERK 1/2 p44, cooperating with the intracellular pathways involving Src kinases, PI3K/Akt, p38 MAPK, well known as activated in response to classical chemoattractants (CXCL8) or priming agents (GM-CSF). On the contrary, the effect of TNF-alpha on neutrophil migration to CCL3 was not dependent on JNK 1/2. In conclusion, the present report shows that TNF-alpha unveils a previously unknown capacity of neutrophils to migrate to CCL3 through the intervention of Mac-1. TNF-alpha regulates Mac-1 up-regulation through signalling pathways, involving various kinases, but not JNK 1/2. Although highly speculative, ERK 1/2 p44 may represent a selective target for the pharmacologic manipulation of neutrophil-mediated adverse activities in TNF-alpha-mediated inflammatory states.
[show abstract][hide abstract] ABSTRACT: Recruitment of leukocytes to inflammatory sites is crucial in the pathogenesis of chronic inflammatory diseases. The aim of this study was to investigate if activation of CB2 cannabinoid receptors would modulate the chemotactic response of human monocytes. Human monocytes treated with the CB2 agonist JWH-015 for 12-18 h showed significantly reduced migration to chemokines CCL2 and CCL3, associated with reduced mRNA and surface expression of their receptors CCR2 and CCR1. The induction of ICAM-1 in response to IFN-gamma was inhibited by JWH-015. Moreover, JWH-015 cross-desensitized human monocytes for migration in response to CCL2 and CCL3 by its own chemoattractant properties. The CB2-selective antagonist SR-144528, but not the CB1 antagonist SR-147778, reversed JWH-015-induced actions, whereas the CB2 agonist JWH-133 mimicked the effects of JWH-015. The investigation of underlying pathways revealed the involvement of phosphatidylinositol 3-kinase/Akt and ERK1/2 but not p38 MAPK. In conclusion, selective activation of CB2 receptors modulates chemotaxis of human monocytes, which might have crucial effects in chronic inflammatory disorders such as atherosclerosis or rheumatoid arthritis.