Exercise and cardiovascular health: get active to "AKTivate" your endothelial nitric oxide synthase.
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Arteries↗
Biomedical subjects
Andreas M Zeiher
Publications and source records attributed to Andreas M Zeiher.
Erythropoietin is a potent physiologic stimulus for endothelial progenitor cell mobilization.
Increasing evidence suggests that postnatal neovascularization involves the recruitment of circulating endothelial progenitor cells (EPCs). Hematopoietic and endothelial cell lineages share common progenitors. Cytokines formerly thought to be specific for the hematopoietic system have only recently been shown to affect several functions in endothelial cells. Accordingly, we investigated the stimulatory potential of erythropoietin (Epo) on EPC mobilization and neovascularization. The bone marrow of Epo-treated mice showed a significant increase in number and proliferation of stem and progenitor cells as well as in colony-forming units. The number of isolated EPCs and CD34+/flk-1+ precursor cells was significantly increased in spleen and peripheral blood of Epo-treated mice compared with phosphate-buffered saline-treated mice. In in vivo models of postnatal neovascularization, Epo significantly increased inflammation- and ischemia-induced neovascularization. The physiologic relevance of these findings was investigated in patients with coronary heart disease. In a multivariate regression model, serum levels of Epo and vascular endothelial growth factor were significantly associated with the number of stem and progenitor cells in the bone marrow as well as with the number and function of circulating EPCs. In conclusion, the present study suggests that Epo stimulates postnatal neovascularization at least in part by enhancing EPC mobilization from the bone marrow. Epo appears to physiologically regulate EPC mobilization in patients with ischemic heart disease. Thus, Epo serum levels may help in identifying patients with impaired EPC recruitment capacity.
Animals↗
Assessment of the tissue distribution of transplanted human endothelial progenitor cells by radioactive labeling.
BACKGROUND: Transplantation of endothelial progenitor cells (EPCs) improves vascularization and left ventricular function after experimental myocardial ischemia. However, tissue distribution of transplanted EPCs has not yet been monitored in living animals. Therefore, we tested whether radioactive labeling allows us to detect injected EPCs. METHODS AND RESULTS: Human EPCs were isolated from peripheral blood, characterized by expression of endothelial marker proteins, and radioactively labeled with [111In]indium oxine. EPCs (106) were injected in athymic nude rats 24 hours after myocardial infarction (n=8) or sham operation (n=8). Scintigraphic images were acquired after 1, 24, 48, and 96 hours after EPC injection. Animals were then killed, and specific radioactivity was measured in different tissues. At 24 to 96 hours after intravenous injection of EPCs, approximately 70% of the radioactivity was localized in the spleen and liver, with only approximately 1% of the radioactivity identified in the heart of sham-operated animals. After myocardial infarction, the heart-to-muscle radioactivity ratio increased significantly, from 1.02+/-0.19 in sham-operated animals to 2.03+/-0.37 after intravenous administration of EPCs. Injection of EPCs into the left ventricular cavity increased this ratio profoundly, from 2.69+/-1.54 in sham-operated animals to 4.70+/-1.55 (P<0.05) in rats with myocardial infarction. Immunostaining of cryosections from infarcted hearts confirmed that EPCs homed predominantly to the infarct border zone. CONCLUSIONS: Although only a small proportion of radiolabeled EPCs are detected in nonischemic myocardium, myocardial infarction increases homing of transplanted EPCs in vivo profoundly. Radiolabeling might eventually provide an useful tool for monitoring the fate of transplanted progenitor cells and for clinical cell therapy.
Animals↗
HMG-CoA reductase inhibitors reduce senescence and increase proliferation of endothelial progenitor cells via regulation of cell cycle regulatory genes.
Endothelial progenitor cells (EPCs) play an important role in postnatal neovascularization of ischemic tissue. Ex vivo expansion of EPCs might be useful for potential clinical cell therapy of myocardial ischemia. However, cultivation of primary cells leads to cellular aging (senescence), thereby severely limiting the proliferative capacity. Therefore, we investigated whether statins might be able to prevent senescence of EPCs. EPCs were isolated from peripheral blood and characterized. After ex vivo cultivation, EPCs became senescent as determined by acidic beta-galactosidase staining. Atorvastatin or mevastatin dose-dependently inhibited the onset of EPC senescence in culture. Moreover, atorvastatin increased proliferation of EPCs as assessed by BrdU incorporation and colony-forming capacity. Whereas geranylgeranylpyrophosphate or farnesylpyrophosphate reduced the senescence inhibitory effect of atorvastatin, NO synthase inhibition, antioxidants, or Rho kinase inhibitors had no effect. To get further insights into the underlying downstream effects of statins, we measured telomerase activity and determined the expression of various cell cycle regulatory genes by using a microarray assay. Whereas telomerase activity did not change, atorvastatin modulated expression of cell cycle genes including upregulation of cyclins and downregulation of the cell cycle inhibitor p27Kip1. Taken together, statins inhibited senescence of EPCs independent of NO, reactive oxygen species, and Rho kinase, but dependent on geranylgeranylpyrophosphate. Atorvastatin-mediated prevention of EPC senescence appears to be mediated by the regulation of various cell cycle proteins. The inhibition of EPC senescence and induction of EPC proliferation by statins in vitro may importantly improve the functional activity of EPCs for potential cell therapy.
Atorvastatin↗
Serum level of the antiinflammatory cytokine interleukin-10 is an important prognostic determinant in patients with acute coronary syndromes.
BACKGROUND: Convincing evidence suggests that atherosclerosis is an inflammatory disease. The inflammatory response is an important determinant of atherosclerotic plaque instability. Therefore, we investigated the prognostic impact of key inflammatory players, namely the inflammatory marker C-reactive protein (CRP) and the antiinflammatory cytokine interleukin-10 (IL-10), in patients with acute coronary syndromes. METHODS AND RESULTS: IL-10, CRP, and troponin T were measured at baseline and before discharge in 547 patients enrolled in the placebo group of the c7E3 Anti Platelet Therapy in Unstable Refractory angina (CAPTURE) trial. Death and nonfatal myocardial infarction were recorded during 6-month follow-up. IL-10 levels did not correlate with troponin T concentrations but were inversely correlated with CRP levels (P<0.001). Patients with elevated IL-10 levels (>3.5 pg/mL; n=276) were at significantly lower risk compared with patients with elevated IL-10 levels (hazard ratio, 0.33; 95% confidence interval [CI], 0.25 to 0.76; P=0.002). The predictive value of IL-10 was independent of myocardial necrosis but significantly interacted with CRP levels. CRP-positive patients with IL-10 serum levels above the calculated threshold value of 3.5 pg/mL were protected from the increased cardiac risk of CRP-positive patients with low IL-10 levels (adjusted hazard ratio, 0.25; 95% CI, 0.10 to 0.63; P=0.003). Moreover, discharge IL-10 levels >2.5 pg/mL were associated with lower cardiac risk during 6-month follow-up (hazard ratio, 0.38; 95% CI, 0.19 to 0.83; P=0.005). CONCLUSIONS: Elevated IL-10 serum levels are associated with a more favorable prognosis in patients with acute coronary syndromes and elevated CRP levels. These data demonstrate the importance of the balance between proinflammatory and antiinflammatory markers as a major determinant of patients' outcome in acute coronary syndromes.
Acute Disease↗
Soluble CD40 ligand in acute coronary syndromes.
BACKGROUND: CD40 ligand is expressed on platelets and released from them on activation. We investigated the predictive value of soluble CD40 ligand as a marker for clinical outcome and the therapeutic effect of glycoprotein IIb/IIIa receptor inhibition in patients with acute coronary syndromes. METHODS: Serum levels of soluble CD40 ligand were measured in 1088 patients with acute coronary syndromes who had previously been enrolled in a randomized trial comparing abciximab with placebo before coronary angioplasty and in 626 patients with acute chest pain. RESULTS: The levels of soluble CD40 ligand were elevated (above 5.0 microg per liter) in 221 patients with acute coronary syndromes (40.6 percent). Among patients receiving placebo, elevated soluble CD40 ligand levels indicated a significantly increased risk of death or nonfatal myocardial infarction during six months of follow-up (adjusted hazard ratio as compared with patients with low levels of the ligand [< or =5.0 microg per liter], 2.71; 95 percent confidence interval, 1.51 to 5.35; P=0.001). The prognostic value of this marker was validated in the patients with chest pain, among whom elevated soluble CD40 ligand levels identified those with acute coronary syndromes who were at high risk for death or nonfatal myocardial infarction (adjusted hazard ratio as compared with those with low levels of the ligand, 6.65; 95 percent confidence interval, 3.18 to 13.89; P<0.001). The increased risk in patients with elevated soluble CD40 ligand levels was significantly reduced by treatment with abciximab (adjusted hazard ratio as compared with those receiving placebo, 0.37; 95 percent confidence interval, 0.20 to 0.68; P=0.001), whereas there was no significant treatment effect of abciximab in patients with low levels of soluble CD40 ligand. CONCLUSIONS: In patients with unstable coronary artery disease, elevation of soluble CD40 ligand levels indicated an increased risk of cardiovascular events. Elevation of soluble CD40 ligand identifies a subgroup of patients at high risk who are likely to benefit from antiplatelet treatment with abciximab.
Abciximab↗
Transdifferentiation of blood-derived human adult endothelial progenitor cells into functionally active cardiomyocytes.
BACKGROUND: Further to promoting angiogenesis, cell therapy may be an approach for cardiac regeneration. Recent studies suggest that progenitor cells can transdifferentiate into other lineages. However, the transdifferentiation potential of endothelial progenitor cells (EPCs) is unknown. METHODS AND RESULTS: EPCs were obtained from peripheral blood mononuclear cells of healthy adults or coronary artery disease (CAD) patients by cultivating with endothelial cell medium and growth factors. After 3 days, >95% of adherent cells were functionally and phenotypically EPCs. Diacetylated LDL-labeled EPCs were then cocultivated with rat cardiomyocytes for 6 days, resulting in significant increases of EPC cell length and size to a cardiomyocyte-like morphology. Biochemically, 9.94+/-1.39% and 5.04+/-1.09% of EPCs from healthy adults (n=15) or CAD patients (n=14, P<0.01 versus healthy adults), respectively, expressed alpha-sarcomeric actinin as measured by flow cytometry. Immunocytochemistry showed that human EPCs expressed alpha-sarcomeric actinin, cardiac troponin I (both with partial sarcomeric organization), atrial natriuretic peptide, and myocyte enhancer factor 2. Fluo 4 imaging demonstrated calcium transients synchronized with adjacent rat cardiomyocytes in transdifferentiated human EPCs. Single-cell microinjection of Lucifer yellow and calcein-AM labeling of cardiomyocytes demonstrated gap junctional communication between 51+/-7% of EPCs (16 hours after labeling, n=4) and cardiomyocytes. EPC transdifferentiation into cardiomyocytes was not observed with conditioned medium but in coculture with paraformaldehyde-fixed cardiomyocytes. CONCLUSIONS: EPCs from healthy volunteers and CAD patients can transdifferentiate in vitro into functionally active cardiomyocytes when cocultivated with rat cardiomyocytes. Cell-to-cell contact but not cellular fusion mediates EPC transdifferentiation. The therapeutic use of autologous EPCs may aid cardiomyocyte regeneration in patients with ischemic heart disease.
Adult↗
Regulation of telomerase activity and anti-apoptotic function by protein-protein interaction and phosphorylation.
The enzyme telomerase is necessary for the synthesis and maintenance of telomere length. The catalytic subunit, telomerase reverse transcriptase (TERT), is regulated by interaction with the 90 kDa heat shock protein (HSP90) and by Akt-dependent phosphorylation. Here, we demonstrate that HSP90 and Akt physically interact with TERT. Treatment of cells with novobiocin, which blocks C-terminal interaction of HSP90, disrupted HSP90 binding to Akt, induced Akt dephosphorylation and significantly reduced telomerase activity. The reduction of TERT activity by novobiocin was associated with an increase in apoptosis. Likewise, the induction of Akt dephosphorylation by protein phosphatase 2A (PP2A) reduced telomerase activity. HSP90 is known to prevent PP2A-mediated dephosphorylation of Akt. To investigate whether the effect of novobiocin is due to the reduction of Akt or TERT phosphorylation, we overexpressed a phospho-mimetic, active Akt (T308D/S473D). Akt (T308D/S473D) prevented novobiocin-induced reduction of telomerase activity and the stimulation of apoptosis. Moreover, overexpression of a dominant negative PP2A construct (PP2A(L199P)) as well as incubation with the PP2A inhibitor okadaic acid blocked the inhibition of telomerase activity by novobiocin. These data suggest that the association between HSP90, Akt and TERT in concert with the phosphorylation of TERT is necessary for maintaining telomerase activity and inhibition of apoptosis.
Apoptosis↗
Nicotine strongly activates dendritic cell-mediated adaptive immunity: potential role for progression of atherosclerotic lesions.
BACKGROUND: Antigen-presenting cells (APCs) such as monocytes and dendritic cells (DCs) stimulate T-cell proliferation and activation in the course of adaptive immunity. This cellular interaction plays a role in the growth of atherosclerotic plaques. Nicotine has been shown to increase the growth of atherosclerotic lesions. Therefore, we investigated whether nicotine can stimulate APCs and their T cell-stimulatory capacity using human monocyte-derived DCs and murine bone marrow-derived DCs as APCs. METHODS AND RESULTS: Nicotine dose-dependently (10(-8) to 10(-4) mol/L) induced DC expression of costimulatory molecules (ie, CD86, CD40), MHC class II, and adhesion molecules (ie, LFA-1, CD54). Moreover, nicotine induced a 7.0-fold increase in secretion of the proinflammatory T(H)1 cytokine interleukin-12 by human DCs. These effects were abrogated by the nicotinic receptor antagonist alpha-bungarotoxin and mecamylamine, respectively. The effects of nicotine were mediated in part by the phosphorylation of the PI3 kinase downstream target Akt and the mitogen-activated kinases ERK and p38 MAPK. Nicotine-stimulated APCs had a greater capacity to stimulate T-cell proliferation and cytokine secretion, as documented by mixed lymphocyte reactions and ovalbumin-specific assays with ovalbumin-transgenic DO10.11 mice. In a murine model of atherosclerosis, nicotine significantly enhanced the recruitment of DCs to atherosclerotic lesions in vivo. CONCLUSIONS: Nicotine activates DCs and augments their capacity to stimulate T-cell proliferation and cytokine secretion. These effects of nicotine may contribute to its influence on the progression of atherosclerotic lesions.
Animals↗
Prognostic significance of angiogenic growth factor serum levels in patients with acute coronary syndromes.
BACKGROUND: In patients with acute coronary syndromes, compensatory processes are initiated, including angiogenesis and endothelial regeneration of ruptured or eroded plaques. Angiogenic growth factors like vascular endothelial growth factor (VEGF), hepatocyte growth factor (HGF), and basic fibroblast growth factor (bFGF) are upregulated during ischemia. However, it is unknown whether their serum levels are related to clinical outcome. METHODS AND RESULTS: We measured VEGF, HGF, and bFGF levels in 1090 patients with acute coronary syndromes. Angiographic evaluation was performed at baseline as well as death, and nonfatal myocardial infarctions were recorded during 6-month follow-up. HGF and VEGF, but not bFGF, were significantly and independently associated with the patients' outcome. Patients with elevated VEGF serum levels suffered from adverse outcome (adjusted hazard ratio, 2.50 [1.52 to 4.82]; P=0.002). VEGF elevation was associated with evidence of ischemia and was a significant predictor of the effect of glycoprotein IIb/IIIa inhibition. In contrast, patients with high HGF levels had a significantly lower event rate compared with patients with low HGF levels (adjusted hazard ratio, 0.33 [0.21 to 0.51]; P<0.001). HGF levels did not correlate with evidence of ischemia and did not predict the effect of abciximab. Intriguingly, however, HGF levels significantly correlated with angiographically visible collateralization of the target vessel (22.4% versus 10.5%; P<0.001). CONCLUSIONS: The angiogenic growth factors VEGF and HGF are independent predictors of the patients' prognosis in acute coronary syndromes. Whereas VEGF elevation correlated with the evidence of myocardial ischemia and indicated an adverse outcome, HGF elevation was independent of ischemia and associated with improved collateralization as well as a favorable prognosis.
Abciximab↗
Interventional cardiology in Europe 1999.
AIMS: The purpose of this registry is to collect data on trends in interventional cardiology within Europe. Special interest focuses on relative increases and ratios in newer revascularization approaches and its distribution in different regions in Europe. METHODS AND RESULTS: Questionnaires distributed to delegates of the national societies of cardiology represented in the European Society of Cardiology to be completed by local institutions and operators yielded that 1,452,751 angiograms and 452,019 PTCAs were performed in 1999. This is an increase of 28% and 16%, respectively, compared with 1998. Most of these increases are due to high relative increases in eastern European countries. The number of PTCAs per 106 inhabitants rose to 714 in 1999. Coronary stenting increased by 31% to about 313,000 stents implanted in 1999. Complication rates remained stable, the need for emergency coronary artery bypass grafting showing a further slight decrease to currently 0.3%. CONCLUSION: Interventional cardiology in Europe is still growing, mainly due to rapid growth in countries with lower socio-economical levels. In some central European countries a saturation seemed to be reached with only minor increases in procedures performed. Coronary stenting remains the only noteworthy and growing complement or alternative to balloon angioplasty.
Angioplasty, Balloon, Coronary↗
Effects of cardiovascular risk factors on coronary artery remodeling in patients with mild atherosclerosis.
BACKGROUND: Vascular remodeling counteracts luminal encroachment during the progression of coronary artery disease (CAD) and modulates the manifestation of hemodynamically significant lesions. However, the role of cardiovascular risk factors for coronary remodeling has not been fully clarified. METHODS: Therefore, we investigated the role of local plaque burden and systemic risk factors on coronary vascular remodeling in 25 patients (49 segments) with angiographically normal or minimally diseased coronary arteries by intravascular ultrasound. In an additional 12 patients without coronary atherosclerosis, physiological vessel tapering was determined and used to calculate the extent of remodeling in diseased segments. RESULTS: An increase in local plaque burden was directly correlated with positive vascular remodeling (r = 0.54, P<0.001). However, cardiovascular risk factors like hypertension (P<0.001) and hypercholesterolemia (P = 0.03) were associated with reduced positive or even negative remodeling. Moreover, the total number of classical cardiovascular risk factors was a strong predictor for reduced positive remodeling (P for trend <0.001). In contrast, coronary flow reserve, a measure of shear stress imposed on the vessel wall, positively correlated with compensatory enlargement (r = 0.44, P = 0.002). By multivariate analysis, plaque burden (P = 0.001), hypertension (P = 0.001) and coronary flow reserve (P = 0.018) proved to be independent determinants of vascular remodeling of epicardial coronary arteries. CONCLUSIONS: Cardiovascular risk factors impair compensatory arterial enlargement and even predispose to shrinkage of epicardial arteries during the initial stage of atherosclerosis. Reduced positive vascular remodeling might contribute to the clinical manifestation of CAD by facilitating the development of flow-limiting stenoses in patients at risk.
Aged↗
Hydrogen peroxide triggers nuclear export of telomerase reverse transcriptase via Src kinase family-dependent phosphorylation of tyrosine 707.
The regulation of telomerase reverse transcriptase (TERT) plays an important role in the proliferative capacity and survival of cells. Here, we report that exogenously as well as endogenously induced oxidative stress leads to translocation of endogenous as well as overexpressed human TERT from the nucleus into the cytosol. TERT is transported through the nuclear pores in a leptomycin-sensitive and Ran GTPase-dependent process. H(2)O(2)-induced nuclear export of TERT is preceded by TERT tyrosine phosphorylation at position 707 and prevented by the Src kinase family inhibitor PP1. Oxidative stress-induced nuclear export of TERT depends on association with the Ran GTPase. In contrast, mutation of tyrosine 707 inhibits phosphorylation induced by oxidative stress and prevents association with Ran and nuclear export of TERT. Moreover, inhibition of tyrosine phosphorylation at 707 increases the antiapoptotic capacity of TERT. Taken together, depletion of nuclear TERT by tyrosine phosphorylation-dependent nuclear export of TERT is a novel mechanism for regulation of TERT localization, which reduces the antiapoptotic activity of TERT.
Active Transport, Cell Nucleus↗
Peptide-treated stent graft for the treatment of saphenous vein graft lesions: first clinical results.
UNLABELLED: Percutaneous interventions of degenerated saphenous vein grafts (SVG) are associated with high rates of distal embolic events and in-stent restenosis. A novel ePTFE-covered stent graft has been developed to improve outcomes for these difficult lesions. The stent graft has been treated with a 15-amino acid, cell adhesion peptide (P-15) to promote endothelialization on the inner surface post-implantation. Patients with de novo lesions in SVGs were treated with a peptide-coated ePTFE stent graft. Follow-up data was collected at 30 days, 3 and 6 months. An angiographic core lab analyzed an angiographic follow-up at 6 months. RESULTS: Procedural success was achieved in all cases. No device-related major adverse cardiac events occurred in-hospital, at 30 days, or at 3 months, despite the lack of distal protection. Six-month restenosis rates were 21%, and were comparable to similar devices. CONCLUSIONS: The peptide-treated stent graft appears to be a safe and effective treatment for SVG lesions, and has the potential to reduce complications without the use of adjunct distal protection in this patient population.
Aged↗
Transplantation of Progenitor Cells and Regeneration Enhancement in Acute Myocardial Infarction (TOPCARE-AMI).
BACKGROUND: Experimental studies suggest that transplantation of blood-derived or bone marrow-derived progenitor cells beneficially affects postinfarction remodeling. The safety and feasibility of autologous progenitor cell transplantation in patients with ischemic heart disease is unknown. METHODS AND RESULTS: We randomly allocated 20 patients with reperfused acute myocardial infarction (AMI) to receive intracoronary infusion of either bone marrow-derived (n=9) or circulating blood-derived progenitor cells (n=11) into the infarct artery 4.3+/-1.5 days after AMI. Transplantation of progenitor cells was associated with a significant increase in global left ventricular ejection fraction from 51.6+/-9.6% to 60.1+/-8.6% (P=0.003), improved regional wall motion in the infarct zone (-1.5+/-0.2 to -0.5+/-0.7 SD/chord; P<0.001), and profoundly reduced end-systolic left ventricular volumes (56.1+/-20 mL to 42.2+/-15.1 mL; P=0.01) at 4-month follow-up. In contrast, in a nonrandomized matched reference group, left ventricular ejection fraction only slightly increased from 51+/-10% to 53.5+/-7.9%, and end-systolic volumes remained unchanged. Echocardiography revealed a profound enhancement of regional contractile function (wall motion score index 1.4+/-0.2 at baseline versus 1.19+/-0.2 at follow-up; P<0.001). At 4 months, coronary blood flow reserve was significantly (P<0.001) increased in the infarct artery. Quantitative F-18-fluorodeoxyglucose-positron emission tomography analysis revealed a significant (P<0.01) increase in myocardial viability in the infarct zone. There were no differences for any measured parameter between blood-derived or bone marrow-derived progenitor cells. No signs of an inflammatory response or malignant arrhythmias were observed. CONCLUSIONS: In patients with AMI, intracoronary infusion of autologous progenitor cells appears to be feasible and safe and may beneficially affect postinfarction remodeling processes.
Cells, Cultured↗
Inhibitors of histone deacetylation downregulate the expression of endothelial nitric oxide synthase and compromise endothelial cell function in vasorelaxation and angiogenesis.
The histone deacetylase (HDAC) inhibitor trichostatin A (TSA) inhibits hypoxia-stimulated angiogenesis. Endothelial nitric oxide synthase (eNOS)-derived NO is central to angiogenesis signaling in endothelial cells (ECs). We hypothesized that the HDAC-dependent regulation of angiogenesis may involve a modulatory effect on eNOS expression. The HDAC inhibitors TSA, butyric acid (BuA), and MS-275 time- and concentration-dependently suppressed eNOS protein levels to 41+/-2%, 46+/-12%, and 40+/-12% of control, respectively. In parallel, TSA and BuA also downregulated eNOS mRNA expression to 21+/-4% and 37+/-4% of control. TSA also attenuated the NO-dependent relaxation of porcine coronary arteries (P<0.0001, TSA 1 micromol/L) and prevented tube formation in a human angiogenesis assay. Although vascular endothelial growth factor substitution did not compensate for the inhibitory effect of TSA, exogenous NO reversed the inhibition of angiogenesis by TSA. To address the underlying signaling mechanism, we characterized the effect of TSA on eNOS gene transcription and mRNA half-life. Although TSA decreased both eNOS protein and mRNA levels, TSA paradoxically enhanced the activity of the eNOS promoter, and did not alter the eNOS transcription rate in nuclear run-on experiments, suggesting that TSA posttranscriptionally targets eNOS mRNA. These data indicate that HDAC-dependent mechanisms contribute to the regulation of eNOS expression in ECs.
Animals↗
CD40 ligand inhibits endothelial cell migration by increasing production of endothelial reactive oxygen species.
BACKGROUND: The CD40/CD40 ligand system is involved in atherogenesis. Activated T lymphocytes and platelets, which express high amounts of CD40 ligand (CD40L) on their surface, contribute significantly to plaque instability with ensuing thrombus formation, leading to acute coronary syndromes. Because reendothelialization may play a pivotal role for plaque stabilization, we investigated a potential role of CD40L on endothelial cell (EC) migration. METHODS AND RESULTS: Stimulation of ECs with recombinant CD40L prevented vascular endothelial growth factor (VEGF)-induced EC migration, as determined by a "scratched wound assay." In addition, activated T lymphocytes and platelets significantly inhibited VEGF-induced EC migration and tube formation in vitro. Because the activation of endothelial nitric oxide (NO) synthase and the release of NO are required for EC migration and angiogenesis, we analyzed the effect of NO. Coincubation with the NO donor S-nitroso-N-acetyl-penicillamine (SNAP) did not reverse the inhibitory effect of CD40L on VEGF-induced EC migration and tube formation. In addition, EC migration induced by SNAP was completely inhibited by CD40L. CD40L, however, induced the production of reactive oxygen species and reduced endothelial NO bioavailability. This reactive oxygen species-dependent effect of CD40L stimulation was reversed with vitamin C or N-acetylcysteine. CONCLUSIONS: The activation of the CD40 receptor inhibits EC migration by increasing reactive oxygen species. The blockade of EC migration by CD40L may critically affect endothelial regeneration after plaque erosion and thereby may contribute to the increased risk for development of acute coronary events in patients with high circulating levels of CD40L.
Antioxidants↗
Hypoxic induction of the hypoxia-inducible factor is mediated via the adaptor protein Shc in endothelial cells.
Tyrosine kinase cascades may play a role in the hypoxic regulation of hypoxia-inducible factor (HIF)-1. We investigated the role of tyrosine kinase phosphorylation and of the Shc/Ras cascade on hypoxic HIF-1 stabilization. Exposure of human umbilical vein endothelial cells to hypoxia results in HIF protein stabilization as early as 10 minutes, with a maximum at 3 hours, and also in Shc tyrosine phosphorylation, with a maximum at 10 minutes. To test whether Shc directly mediates hypoxia-induced HIF stabilization, human umbilical vein endothelial cells were transfected with a dominant-negative Shc mutant (dnShc), resulting in significantly reduced HIF protein levels compared with control. Similar results were obtained with cells transfected with dominant-negative Ras, a known downstream effector of Shc. Hypoxia-induced Ras activity was significantly reduced in cells transfected with dnShc compared with control levels, indicating that Ras indeed acts downstream from Shc. Moreover, cells pretreated with a specific Raf-1 kinase inhibitor, a known downstream effector of Ras, exhibited reduced HIF protein levels. To examine the functional consequences of Shc in hypoxic signaling, HIF-1 ubiquitination, protein stabilization, and endothelial cell migration were assessed. Overexpression of dnShc increased ubiquitination of HIF-1 and reduced the half-life of the protein. Moreover, dnShc, dominant-negative Ras, or the Raf-1 kinase inhibitor significantly inhibited migration under hypoxia. Thus, Shc in concert with Ras and Raf-1 contributes to hypoxia-induced HIF-1alpha protein stabilization and endothelial cell migration.
Adaptor Proteins, Signal Transducing↗