Coagulation factor Xa appears involved in the pathogenesis of pulmonary fibrosis. Through its interaction with protease activated receptor-1, this protease signals myofibroblast differentiation in lung fibroblasts. Although fibrogenic stimuli induce factor X synthesis by alveolar cells, the mechanisms of local posttranslational factor X activation are not fully understood. Cell-derived microparticles are submicron vesicles involved in different physiological processes, including blood coagulation; they potentially activate factor X due to the exposure on their outer membrane of both phosphatidylserine and tissue factor. We postulated a role for procoagulant microparticles in the pathogenesis of interstitial lung diseases. Nineteen patients with interstitial lung diseases and 11 controls were studied. All subjects underwent bronchoalveolar lavage; interstitial lung disease patients also underwent pulmonary function tests and high resolution CT scan. Microparticles were enumerated in the bronchoalveolar lavage fluid with a solid-phase assay based on thrombin generation. Microparticles were also tested for tissue factor activity. In vitro shedding of microparticles upon incubation with H₂O₂ was assessed in the human alveolar cell line, A549 and in normal bronchial epithelial cells. Tissue factor synthesis was quantitated by real-time PCR. Total microparticle number and microparticle-associated tissue factor activity were increased in interstitial lung disease patients compared to controls (84±8 vs. 39±3 nM phosphatidylserine; 293±37 vs. 105±21 arbitrary units of tissue factor activity; mean±SEM; p

There is increasing evidence that an elevation of oxidative stress and associated oxidative damages are mediators of vascular injury in various cardiovascular pathologies, including hypertension. Accumulation of oxidative damage is thought to play an important role in aging and age-associated diseases such as hypertension and oxidative stress may function as a common trigger for activation of the senescence programme. In this regard, the role of telomeres in the onset, development and prognosis of hypertension has generated considerable interest. These structures may deteriorate in the onset and development of arterial hypertension in which their length may be a predictor of outcome. As telomere length by its nature is a marker of cell senescence, this parameter is of particular interest when studying the lifespan and fate of endothelial cells, cardiomyocytes and smooth muscle cells, especially so because telomere length seems to be regulated by various factors notably certain cardiovascular risk factors, such as smoking, sex and obesity that are associated with high levels of oxidative stress. This review focuses on the vascular effects of reactive oxygen species and the role of oxidative stress in hypertension-associated vascular damage. In addition it reviewes the considerable amount of data published recently on the role of telomeres to gain insights into the links between telomere length and hypertension, and assesses the usefulness of telomere length as a new marker of cardiovascular risk.

The aim of this study was to investigate hypoxia effects on vascular endothelial growth factor (VEGF) and its receptors VEGFR-1 and VEGFR-2 in human umbilical vein endothelial cells (HUVEC) and to determine their modulation by the peptide somatostatin (SRIF) and its analogues. The involvement of signal transducer and activator of transcription (STAT) 3 and hypoxia inducible factor (HIF)-1 was also investigated. Quantitative real-time PCR, Western blot and ELISA were used. Hypoxia upregulated VEGF expression and release, whereas it downregulated VEGFR-1 and VEGFR-2. In contrast, neither the expression nor the phosphorylation of the platelet-derived growth factor receptor (PDGFR) β was affected by hypoxia. SU1498 at 1 μM did not affect pVEGFR-2 and pPDGFRβ, whereas at 20 μM it inhibited pVEGFR-2, but not pPDGFRβ. Upregulated VEGF expression and release were prevented by SU1498, which also inhibited the hypoxia-induced pSTAT3 and HIF-1α. Blocking pSTAT3 with S3I-201 inhibited HIF-1α and VEGF upregulation, suggesting the existence of an autocrine loop involving STAT3, HIF-1, VEGF and VEGFR-2. Endothelial cells express somatostatin (SRIF) receptors (sst1–5) although less is known in HUVEC. We found that sst1 and sst4 were expressed by HUVEC with sst1 more expressed than sst4 mRNA. Hypoxia downregulated sst1, whereas it upregulated sst4. The sst1 downregulation, but not the sst4 upregulation, was prevented by SU1498, S3I-201 or YC-1, an inhibitor of HIF-1α. SRIF and the sst1 agonist CH-275, but not the sst4 agonist L803,087 and the sst2/sst3/sst5 agonist octreotide, prevented hypoxia effects on VEGF and its receptors. In addition, SRIF and CH-275 inhibited the hypoxia-induced pSTAT3 and HIF-1α accumulation. Our results suggest that SRIF acting at sst1 limits upregulated VEGF expression and release through a control on the activity of STAT3 and HIF-1, supporting the possible use of sst1 agonists in antiangiogenic therapies.

Tobacco smoking remains the second largest preventable cause of mortality and morbidity worldwide. Exposure to tobacco smoke causes coronary disease, atherosclerosis and ischemic vessel disease. The degree of this risk is proportional to the amount of smoking and it varies from individual to individual because of between-individual differences in genetic background. While the chemical properties of tobacco smoke are relatively well characterized, the mechanisms by which smoking leads to disease and the genetic factors that determine susceptibility to these diseases are not well understood. The purpose of the present review is to describe the interaction between DNA variants in some important genes and cardiovascular diseases; and how the exposure to cigarette smoke significantly modifies the association between genetic variants and cardiovascular risk. A great number of gene-enzymes that usually protect against cardiovascular events may be adversely influenced by tobacco smoke and, through this way, exert less effective action.

Purpose: No studies have been addressed to the differences in inflammation kinetics between ST-segment elevation myocardial infarction (STEMI) and non-ST-segment elevation myocardial infarction (NSTEMI).Patients and methods: Forty consecutive patients with acute coronary syndrome (ACS) (n = 23 STEMI, age = 61.7 +/- 10.3 years; it = 17 NSTEMI, age = 65.6 +/- 11.3 years) were enrolled within 12 h after symptoms. All patients received therapy according to the current Guidelines.Blood samples were collected at admission (t0), on days 7 (t1) and 30 (t2) to evaluate CD40 ligand (CD40L), transforming growth factor (TGF)-beta, interleukin (IL)-6, tumor necrosis factor (TNF)-alpha and its receptors TNFRI and TNFRII, high sensitivity C-reactive protein (hs-CRP), serum amyloid A (SAA) and white blood cells (WBC). Echocardiographic parameters were also evaluated.Results: STEMI patients, at admission, had significantly higher median values of hs-CRP (p < 0.001), WBC (p < 0.01), ferritin (p < 0.0005) and IL-6 (p < 0.05) than NSTEMI. On the contrary, NSTEMI patients had lower median levels of every inflammatory marker except for CD40L (p < 0.05) that was significantly higher. Moreover, three out of four deceased patients presented levels of CD40L higher than the median. At admission, STEMI showed a reduced ejection fraction (EF, p < 0.01) and increased wall motion score index (WMSI, p < 0.001) and end-diastolic volume (EDV, p < 0.05) vs NSTEMI. An inverse correlation between admission values of inflammatory markers (SAA and WBC) and cardiac function was observed (p < 0.05). Moreover, the necrosis marker troponin I was positively correlated with both WMSI (p < 0.05) and hs-CRP (p < 0.05).Regarding the inflammation kinetics, a difference was observed in the two groups only for WBC (p < 0.05) and SAA (p < 0.05). SAA showed higher values in STEMI at t0 and t1. In both groups, TGF-beta had an increase at t1 and t2 with respect to admission, while IL-6 had a decreasing trend.The total incidence of major adverse clinical events (MACE) was 22.5% at t2, with a mortality rate of 10%.Conclusion: These observations suggest a differential inflammatory pattern in STEMI and NSTEMI patients. The absence of significant correlations between inflammatory indexes and myocardial infarction in NSTEMI supports the hypothesis that a different pattern of inflammation occurs in these patients. CD40L may have an important role as a marker for risk stratification in patients with ACS. (C) 2009 Elsevier Masson SAS. All rights reserved.

INTRODUCTION:Endothelial progenitor cells are circulating cells able to home to sites of vascular damage and to contribute to the revascularization of ischemic areas. We evaluated whether endothelial progenitor cells synthesize tissue factor, a procoagulant protein also involved in angiogenesis. MATERIALS AND METHODS:Endothelial progenitor cells were obtained from the peripheral blood mononuclear fraction of normal donors and cultured in endothelial medium supplemented with specific growth factors. The procoagulant activity expressed by cells disrupted by freeze-thaw cycles was assessed by a one stage clotting assay. Tissue factor mRNA expression was evaluated by RT-PCR. RESULTS:Endothelial progenitor cells do not express procoagulant activity in baseline conditions. However, lipopolysaccharide induces the expression of procoagulant activity. The effect is dose-dependent and reaches statistical significance at 100 ng/mL lipopolysaccharide. Inhibition with an anti-tissue factor antibody and amplification of cDNA with primers based on the tissue factor sequence confirm the identity of this activity with tissue factor. The kinetics of tissue factor expression by endothelial progenitor cells is identical to that of human umbilical vein endothelial cells showing maximal activity within 4 hours, and then decreasing; in contrast, tissue factor expression by mononuclear cells lasts for longer times. Both 5,6-dichloro-beta D-ribofuranosyl-benzimidazole and cycloheximide prevented the expression of procoagulant activity. Stimulation of endothelial progenitor cells with tumor necrosis factor-alpha did not elicit any detectable procoagulant activity. CONCLUSIONS:Endothelial progenitor cells can be stimulated by lipopolysaccharide to synthesize tissue factor. This protein might be involved in thrombotic phenomena and might contribute to endothelial progenitor cells related neovascularization.

The hyaluronan synthase 1 (HAS-1) gene encodes a plasma membrane protein that synthesizes hyaluronan (HA), an extracellular matrix molecule. Accumulating evidence emphasizes the relevance of HA metabolism in an increasing number of processes of clinical interest, including abdominal aortic aneurysm (AAA). The existence of aberrant splicing variants of the HAS-1 gene could partly explain the altered extracellular matrix architecture and influence various biological functions, resulting in progressive arterial wall failure in the development of AAA. In the present study, we assessed the hypothesis that HAS-1 genetic 833A/G polymorphism could be associated with the risk of AAA by performing a case-control association study, involving AAA patients and healthy matched donors.

Patients with critical limb ischemia (CLI) have low levels of endothelial progenitor cells (EPC). Iloprost has been demonstrated to stimulate vascular endothelial growth factor (VEGF) and promote angiogenesis. We investigated the effects of iloprost on EPC levels in vivo in CLI patients. Twenty-three patients with stage III and IV CLI were treated with iloprost for four weeks, improving clinical and instrumental parameters. Mononuclear cells isolated from peripheral blood were cultured to obtain "early" EPC, evaluated counting adherent cells with double positivity for acetylated low-density lipoprotein uptake and Ulex Europaeus lectin at flow cytometry. These cells also co-expressed the monocyte markers CD14 and CD45. Iloprost increased EPC number in the whole patient population: pre-treatment median: 13,812/ml; range: 1,263-83,648/ml; post-treatment median: 23,739/ml; range: 3,385-99,251/ml; p = 0.035, irrespective of age, sex, disease stage or atherosclerosis risk factors. In conclusion, iloprost increases EPC number in peripheral blood in vivo. Such an effect may have therapeutic relevance.

Abdominal aortic aneurysm (AAA) has a multifactorial aetiology and the importance of genetic components is getting increasing interest. Alteration in the structure of the vascular extracellular matrix has been described in AAA. Matrix metalloproteinases (MMPs) degrade extracellular matrix proteins which alter the vessel wall stability. We evaluated two different polymorphisms, a CA repeat and a cytosine to thymidine transition in the promoter sequence of MMP-9 gene for frequency in 146 patients with AAA. We compared the results with those of 156 healthy subjects. No difference was found in the allelic distribution of either polymorphisms. We therefore found no evidence that MMP-9 is a marker of susceptibility for AAA.

Somatostatin (SRIF)-14 is recognized as an important mediator between the nervous and the immune system, although the functional role of its receptors (sst(1)-sst(5)) is poorly understood in humans. In our study, we demonstrate that human macrophages, differentiated from PBMC-derived monocytes, express sst(1) and sst(2) mRNAs. sst(1) and sst(2) are mostly localized at the cell surface and display active binding sites. In particular, sst(1)/sst(2) activation results in a weak internalization of sst(1), and the sst(2) internalization appears more efficient. At the functional level, the activation of SRIF receptors by the multiligand analogs SOM230 and KE108, but not by SRIF-14 or cortistatin-14, reduces macrophage viability. Their effects are mimicked by the selective activation of sst(1) and sst(2) using CH-275 and SMS 201-995/L-779,976, respectively. Further, sst(1)- and sst(2)-mediated effects are reversed by the sst(1) antagonist SRA-880 or the sst(2) antagonist CYN 154806, respectively. CH-275, SMS 201-995, and L-779,976, but not SRIF-14, decrease mRNA expression and secretion of the MCP-1. In addition, SRIF-14, CH-275, SMS 201-995, and L-779,976 decrease IL-8 secretion, and they do not affect IL-8 mRNA expression. In contrast, SRIF-14 and sst(1)/sst(2) agonists do not affect the secretion of matrix metalloproteinase-9. Collectively, our results suggest that the SRIF system, through sst(1) and sst(2), exerts mainly an immunosuppressive effect in human macrophages and may, therefore, represent a therapeutic window that can be exploited for the development of new strategies in pharmacological therapy of inflammation.

Several lines of evidence, including an increased level of lipid peroxidation and the depletion of antioxidant molecules like as glutathione (GSH), indicate that oxidative stress plays an important role in the pathogenesis of several neurodegenerative disorders, such as Parkinson's disease (PD) and Alzheimer's disease (AD). We previously observed a significant increased level of DNA oxidative damage in peripheral blood cells of PD patients, with respect to controls, moreover, the activity of glutathione transferases (GSTs) measured in circulating plasma was higher in controls than in PD patients, suggesting a lower enzymatic protection in PD individuals. Among human GSTs, glutathione transferase A4-4 displays a high catalitic activity towards 4-hydroxy-2-nonenal (HNE), a marker of lipid peroxidation whose levels have been found significantly increased in the substantia nigra of Parkinson's disease patients, in respect to controls. We performed this study to determine the presence of allelic variants of functional interest in the coding region of the hGSTA4 gene on 60 PD patients and 60 healthy controls. By the combined effort of polymerase chain reaction/single-strand conformation polymorphisms (PCR/SSCP) techniques, we observed a single nucleotide polymorphism (SNP) G351A leading to the silent mutation Gln117Gln. No significant difference was observed in the distribution of this polymorphism between PD individuals and controls, moreover, we did not observe any other polymorphism in the hGSTA4 gene in our population. Further studies are required to test the role played by both factors regulating the level of the expression of the hGSTA4 gene and any possible post-translational modification of the protein, in the protection against oxidative damage in neuronal cells.