An increase in nitric oxide produced by rat peritoneal neutrophils is not involved in cell apoptosis

Polymorphonuclear neutrophils (PMN) obtained from carrageenin-stimulated peritoneal cavities of rats, but not blood PMN, spontaneously produced nitric oxide (NO) when incubated in vitro. Incubation of the cells with the NO synthase inhibitors, L-imino-ethyl-L-ornithine (L-NIO) or NG-monomethyl-L-arginine (L-NMMA), inhibited NO production. This inhibition could be reversed by L-arginine. Incubation of PMN with lipopolysaccharide (LPS) failed to enhance NO production. Pretreatment of the rats with dexamethasone (DEXA) prior to carrageenin injection or incubation of PMN with the glucocorticoid in vitro partially inhibited the spontaneous release of NO. On the other hand, when PMN obtained from DEXA pretreated rats were incubated in vitro with DEXA, NO synthase activity and hence NO generation were almost abolished. A similar inhibition was also observed following the addition of L-NIO or cycloheximide to cultures of carrageenin-elicited PMN. The NO production by PMN did not appear to be related to cell viability or apoptosis. Indeed, neither the blockade of NO generation by L-NIO nor the incubation of the neutrophils with a NO donor, S-nitroso-acetylpenicillamine (SNAP) modified the pattern of LDH release or DNA fragmentation. In summary, it appears that PMN migration triggers a continuous NO synthesis, and that NO produced by these cells is not related to their apoptosis

Heme modulates smooth muscle cell proliferation and migration via NADPH oxidase: A counter-regulatory role for heme oxygenase system

AbstractAccumulation of vascular smooth muscle cells (VSMC) in response to inflammatory stimuli is a key event in atherogenesis, which commonly occurs in sinuous vessels with turbulent blood flow what leads to hemolysis and consequent free heme accumulation, a known pro-oxidant and pro-inflammatory molecule.In this work, we investigated the effects of free heme on VSMC, and the molecular mechanisms underlying this process.Free heme induces a concentration-dependent migration and proliferation of VSMC which depends on the production of reactive oxygen species (ROS) derived from NADPH oxidase (NADPHox) activity. Additionally, heme activates redox-sensitive proliferation-related signaling routes, such as Mitogen Activated Protein Kinase (MAPK) and NF-κB, and induces Heme Oxygenase-1 (HO-1) expression. NADPHox-dependent proliferative effect of heme seems to be endogenously modulated by HO since the pretreatment of VSMC with HO inhibitors potentiates heme-induced proliferation and, in parallel, increases ROS production. These effects were no longer observed in the presence of heme metabolites, carbon monoxide and biliverdin.The data indicate that VSMC proliferation induced by heme is endogenously modulated by a critical counter-regulatory crosstalk between NADPHox and HO systems

Heme and innate immunity: new insights for an old molecule

Hemolytic episodes such as sickle cell disease, malaria and ischemia-reperfusion occurrence are often associated to the statement of an inflammatory response which may develop or not to a chronic inflammatory status. Although these pathological states are triggered by distinct etiological agents, all of them are associated to high levels of free heme in circulation. In this review, we aim to focus the very recent achievements that have led to the statement of free heme as a proinflammatory molecule, which may play a central role during the onset and/or persistance of inflammation during these pathologies

Procoagulant properties of human MV3 melanoma cells

A correlation between cancer and prothrombotic states has long been described. More recently, a number of studies have focused on the procoagulant mechanisms exhibited by tumor cells. In the present study, we dissected the molecular mechanisms responsible for the procoagulant activity of MV3, a highly aggressive human melanoma cell line. It was observed that tumor cells strongly accelerate plasma coagulation as a result of: i) expression of the blood clotting initiator protein, a tissue factor, as shown by flow cytometry and functional assays (factor Xa formation in the presence of cells and factor VIIa), and ii) direct activation of prothrombin to thrombin by cells, as evidenced by hydrolysis of the synthetic substrate, S-2238, and the natural substrate, fibrinogen. This ability was highly potentiated by the addition of exogenous factor Va, which functions as a co-factor for the enzyme factor Xa. In contrast, prothrombin activation was not observed when cells were previously incubated with DEGR-factor Xa, an inactive derivative of the enzyme. Moreover, a monoclonal antibody against bovine factor Xa reduced the prothrombin-converting activity of tumor cells. In conclusion, the data strongly suggest that MV3 cells recruit factor Xa from the culture medium, triggering an uncommon procoagulant mechanism

Flavonoids Bearing an O-arabinofuranosyl-(1 -> 3)-rhamnoside Moiety from Cladocolea micrantha: Inhibitory Effect on Human Melanoma Cells

Eleven known triterpenes (alpha-amyrin, beta-amyrin, lupeol, and their respective acetates, 3-O-acetyl derivatives of betulinic, oleanolic, and ursolic acids, cycloartenol, and tirucall-7,24-dicnol), two new flavonols presenting an uncommon interglycosidic O-[1 -> 3) linkage (kaempferol 3-O-alpha-L-arabinofuranosyl-(1 -> 3)-alpha-L-rhamnoside and quercetin 3-O-alpha-L-arabinofuranosyl-(1 -> 3)-alpha-L-rhamnoside), beta-sitosterol, stigmasterol, quercetin, and gallic acid were isolated from the Amazonian medicinal mistletoe, Cladocolea micrantha Kuijt (Loranthaceae). Their structures were established by spectral methods and eventual chromatographic comparisons. The quercetin derivative was not cytotoxic to MV3 human melanoma cells, but was able, when administered at 1 mu g/mL, to promote a twofold inhibition of the migration of the cells through the transwell system when compared with paclitaxel at 5 mu M.71013111314Fundação de Amparo à Pesquisa do Estado do Rio de Janeiro (FAPERJ)FAPERJ Grant [E-26/110.032/2011

Lipoxin A4 selectively programs the profile of M2 tumor-associated macrophages which favour control of tumor progression

In tumor microenvironments, the macrophage population is heterogeneous, but some macrophages can acquire tumor-promoting characteristics. These tumor-associated macrophages (TAM) exhibit an M2-like profile, with deficient production of NO and ROS, characteristics of pro-inflammatory M1 cytotoxic macrophages. The results show an unexpected effect of lipoxin, which induces in TAM a change from an M2- to an M1-like profile, thereby triggering tumor cell apoptosis and down-modulating the tumor progression