RESUMO
Intravascular hemolysis, a major manifestation of sickle cell disease (SCD) and other diseases, incurs the release of hemoglobin and heme from red blood cells, in turn triggering inflammatory processes. This study investigated the in vitro effects of heme, a major inflammatory DAMP, on the adhesive properties of isolated human neutrophils. Heme (20 and 50 µM) significantly increased the adhesion of neutrophils to fibronectin and to recombinant ICAM-1, under static conditions, even more efficiently than the potent pro-inflammatory cytokine, tumor necrosis factor-α (TNF); a microfluidic assay confirmed that heme stimulated neutrophil adhesion under conditions of shear stress. Heme-induced neutrophil adhesion was associated with the increased activities, but not expressions, of the Mac-1 and LFA-1 integrin subunits, CD11b and CD11a, on the cell surface. Notably, heme (50 µM) significantly induced NFκB translocation in neutrophils, and inhibition of NFκB activity with the BAY11-7082 molecule abolished heme-induced cell adhesion to fibronectin and significantly decreased CD11a activity. Flow cytometric analysis demonstrated major reactive oxygen species (ROS) generation in neutrophils following heme stimulation that could be inhibited by the antioxidant, α-tocopherol, and by BAY11-7082. Furthermore, co-incubation with α-tocopherol abrogated both heme-stimulated neutrophil adhesion and CD11a/CD11b activation. Thus, our data indicate that heme, at clinically relevant concentrations, is a potent activator of neutrophil adhesion, increasing the ligand affinity of the ß2 integrins via a mechanism that may be partially mediated by an NFkB-dependent pathway and the generation of ROS. Given the fundamental role that the adhesion of neutrophils to the vascular wall plays in SCD vaso-occlusion and other vascular inflammatory processes, our findings provide further evidence that cell-free heme is a major therapeutic target in the hemolytic diseases.
Assuntos
Endotélio Vascular/efeitos dos fármacos , Heme/farmacologia , NF-kappa B/metabolismo , Neutrófilos/efeitos dos fármacos , Espécies Reativas de Oxigênio/metabolismo , Anemia Falciforme/metabolismo , Anemia Falciforme/patologia , Antígenos CD18/metabolismo , Adesão Celular/efeitos dos fármacos , Células Cultivadas , Endotélio Vascular/metabolismo , Endotélio Vascular/patologia , Hemólise , Humanos , Molécula 1 de Adesão Intercelular/metabolismo , Leucócitos Mononucleares , Neutrófilos/metabolismo , Neutrófilos/patologia , Transdução de SinaisRESUMO
BACKGROUND: Sickle cell disease (SCD) incurs vaso-occlusive episodes and organ damage, including nephropathy. Despite displaying characteristics of vascular dysfunction, SCD patients tend to present relatively lower systemic blood pressure (BP), via an unknown mechanism. We investigated associations between BP and renin-angiotensin-system (RAS) components in SCD and determined whether an inhibitor of angiotensin converting enzyme (ACE; often used to slow SCD glomerulopathy) further modulates BP and RAS components in a murine model of SCD. METHODS: BP was compared in human subjects and mice with/without SCD. Plasma angiotensin II, ACE and renin were measured by immunoassay. BP was reevaluated after treating mice with enalapril (25 mg/kg, 5x/week) for 5 weeks; plasma and organs were stored for angiotensin II and ACE activity measurement, and quantitative real-time PCR. RESULTS: Diastolic BP and systolic BP were significantly lower in patients and mice with SCD, respectively, compared to controls. Reduced BP was associated with increased plasma renin and markers of kidney damage (mice) in SCD, as well as significantly decreased plasma ACE concentrations and ACE enzyme activity. As expected, enalapril administration lowered BP, plasma angiotensin II and organ ACE activity in control mice. In contrast, enalapril did not further reduce BP or organ ACE activity in SCD mice; however, plasma angiotensin II and renin levels were found to be significantly higher in enalapril-treated SCD mice than those of treated control mice. CONCLUSION: Relative hypotension was confirmed in a murine model of SCD, in association with decreased ACE concentrations in both human and murine disease. Given that ACE inhibition has an accepted role in decreasing BP, further studies should investigate mechanisms by which ACE depletion, via both Ang II-dependent and alternative pathways, could contribute to reduce BP in SCD and understand how ACE inhibition confers Ang II-independent benefits on kidney function in SCD.
Assuntos
Anemia Falciforme/tratamento farmacológico , Anemia Falciforme/fisiopatologia , Inibidores da Enzima Conversora de Angiotensina/farmacologia , Pressão Sanguínea/efeitos dos fármacos , Peptidil Dipeptidase A/biossíntese , Adolescente , Adulto , Angiotensina II/metabolismo , Animais , Diástole , Modelos Animais de Doenças , Feminino , Humanos , Hidroxiureia/farmacologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Pessoa de Meia-Idade , Reação em Cadeia da Polimerase em Tempo Real , Renina/sangue , Sistema Renina-Angiotensina , Sístole , Adulto JovemRESUMO
A new acylamino acid, bunodosine 391 (BDS 391), was isolated from the venom of the sea anemone Bunodosoma cangicum. The structure was elucidated by spectroscopic analyses (2D NMR, ESIMS/MS) and verified by its synthesis. Intraplantar injection of BDS 391 into the hind paw of a rat induced a potent analgesic effect. This effect was not altered by naloxone (an opioid receptor antagonist), but was completely reversed by methysergide (a serotonin receptor antagonist), indicating that the effect is mediated by activation of serotonin receptors.
Assuntos
Analgésicos/isolamento & purificação , Analgésicos/farmacologia , Anêmonas-do-Mar/química , Analgésicos/química , Animais , Venenos de Cnidários/síntese química , Venenos de Cnidários/química , Venenos de Cnidários/isolamento & purificação , Venenos de Cnidários/farmacologia , Edema/induzido quimicamente , Edema/tratamento farmacológico , Membro Posterior/efeitos dos fármacos , Masculino , Estrutura Molecular , Naloxona/farmacologia , Antagonistas de Entorpecentes/farmacologia , Ressonância Magnética Nuclear Biomolecular , Ratos , Ratos Wistar , Receptores de Serotonina/efeitos dos fármacos , Receptores de Serotonina/metabolismoRESUMO
Crotoxin (CTX), a neurotoxin isolated from the venom of the South American rattlesnake Crotalus durissus terrificus, induces analgesia. In this study, we evaluated the antinociceptive effect of CTX in a model of neuropathic pain induced by rat sciatic nerve transection. Hyperalgesia was detected 2 h after nerve transection and persisted for 64 days. Immersion of proximal and distal nerve stumps in CTX solution (0.01 mM for 10 s), immediately after nerve transection, blocked hyperalgesia. The antinociceptive effect of CTX was long-lasting, since it was detected 2 h after treatment and persisted for 64 days. CTX also delayed, but did not block, neurectomy-induced neuroma formation. The effect of CTX was blocked by zileuton (100 mg/kg, p.o.) and atropine (10 mg/kg, i.p.), and reduced by yohimbine (2 mg/kg, i.p.) and methysergide (5 mg/kg, i.p.). On the other hand, indomethacin (4 mg/kg, i.v.), naloxone (1 mg/kg, i.p.), and N-methyl atropine (30 mg/kg, i.p.) did not interfere with the effect of CTX. These results indicate that CTX induces a long-lasting antinociceptive effect in neuropathic pain, which is mediated by activation of central muscarinic receptors and partially, by activation of alpha-adrenoceptors and 5-HT receptors. Eicosanoids derived from the lipoxygenase pathway modulate the action of crotoxin.
Assuntos
Analgésicos não Narcóticos , Araquidonato 5-Lipoxigenase/fisiologia , Crotoxina/farmacologia , Dor/tratamento farmacológico , Dor/etiologia , Doenças do Sistema Nervoso Periférico/complicações , Receptores Muscarínicos/efeitos dos fármacos , Animais , Anti-Inflamatórios não Esteroides/farmacologia , Araquidonato 5-Lipoxigenase/metabolismo , Creatina Quinase/sangue , Creatina Quinase/metabolismo , Eicosanoides/metabolismo , Eicosanoides/fisiologia , Hiperalgesia/tratamento farmacológico , Hiperalgesia/psicologia , Indometacina/farmacologia , Masculino , Atividade Motora/efeitos dos fármacos , Agonistas Muscarínicos/farmacologia , Antagonistas Muscarínicos/farmacologia , Naloxona/farmacologia , Antagonistas de Entorpecentes/farmacologia , Limiar da Dor/efeitos dos fármacos , Ratos , Nervo Isquiático/lesões , Serotonina/fisiologiaRESUMO
Direct-acting cannabinoid receptor ligands are well known to reduce hyperalgesic responses after nerve injury, although their psychoactive side effects have damped enthusiasm for their therapeutic development. Hemopressin (Hp) is a nonapeptide that selectively binds CB1 cannabinoid receptors (CB1 receptors) and exerts antinociceptive action in inflammatory pain models. We investigated the effect of Hp on neuropathic pain in rats subjected to chronic constriction injury (CCI) of the sciatic nerve, and explored the mechanisms involved. Oral administration of Hp inhibits mechanical hyperalgesia of CCI-rats up to 6h. Hp treatment also decreases Egr-1 immunoreactivity (Egr-1Ir) in the superficial layer of the dorsal horn of the spinal cord of CCI rats. The antinociceptive effect of Hp seems to be independent of inhibitory descending pain pathway since methysergide (5HT1A receptor antagonist) and yohimbine (α-2 adrenergic receptor antagonist) were unable to prevent Hp antinociceptive effect. Hp decreased calcium flux on DRG neurons from CCI rats, similarly to that observed for AM251, a CB1 receptor antagonist. We also investigated the effect of Hp on potassium channels of CCI rats using UCL 1684 (a blocker of Ca(2+)-activated K(+) channels) which reversed Hp-induced antinociception. Furthermore, concomitant administration of URB-584 (FAAH inhibitor) but not JZL-184 (MAGL inhibitor) potentiates antinociceptive effect of Hp in CCI rats indicating an involvement of anadamide on HP-induced antinociception. Together, these data demonstrate that Hp displays antinociception in pain from neuropathic etiology through local effects. The release of anandamide and the opening of peripheral K(+) channels are involved in the antinociceptive effect.