cp-105696 and Disease-Models--Animal

Age-related macular degeneration (AMD), a progressive chronic disease of the central retina, is associated with aging and is a leading cause of blindness worldwide. Here, we demonstrate that leukotriene B4 (LTB4) receptor 1 (BLT1) promotes laser-induced choroidal neovascularization (CNV) in a mouse model for wet-type AMD. CNV was significantly less in BLT1-deficient (BLT1-KO) mice compared with BLT1-WT controls. Expression of several proangiogenic and profibrotic factors was lower in BLT1-KO eyes than in BLT1-WT eyes. LTB4 production in the eyes was substantially increased in the early phase after laser injury. BLT1 was highly expressed in M2 macrophages in vitro and in vivo, and ocular BLT1+ M2 macrophages were increased in the aged eyes after laser injury. Furthermore, M2 macrophages were rapidly attracted by LTB4 and subsequently produced VEGF-A- through BLT1-mediated signaling. Consequently, intravitreal injection of M2 macrophages augmented CNV formation, which was attenuated by BLT1 deficiency. Thus, laser-induced injury to the retina triggered LTB4 production and attracted M2 macrophages via BLT1, leading to development of CNV. A selective BLT1 antagonist (CP105696) and 3 LTB4 inhibitors (zileuton, MK-886, and bestatin) reduced CNV in a dose-dependent manner. CP105696 also inhibited the accumulation of BLT1+ M2 macrophages in the laser-injured eyes of aged mice. Together, these results indicate that the LTB4-BLT1 axis is a potentially novel therapeutic target for CNV of wet-type AMD.

Topics: Animals; Benzopyrans; Carboxylic Acids; Choroidal Neovascularization; Disease Models, Animal; Eye; Eye Injuries; Hydroxyurea; Indoles; Lasers; Leucine; Leukotriene B4; Macrophages; Macular Degeneration; Male; Mice; Mice, Knockout; Neovascularization, Pathologic; Receptors, Leukotriene B4; Signal Transduction

Leukotriene B4 (LTB4) mediates different inflammatory events such as neutrophil migration and pain. The present study addressed the mechanisms of LTB4-mediated joint inflammation-induced hypernociception. It was observed that zymosan-induced articular hypernociception and neutrophil migration were reduced dose-dependently by the pretreatment with MK886 (1-9 mg/kg; LT synthesis inhibitor) as well as in 5-lypoxygenase-deficient mice (5LO(-/-)) or by the selective antagonist of the LTB(4) receptor (CP105696; 3 mg/kg). Histological analysis showed reduced zymosan-induced articular inflammatory damage in 5LO(-/-) mice. The hypernociceptive role of LTB4 was confirmed further by the demonstration that joint injection of LTB4 induces a dose (8.3, 25, and 75 ng)-dependent articular hypernociception. Furthermore, zymosan induced an increase in joint LTB4 production. Investigating the mechanism underlying LTB4 mediation of zymosan-induced hypernociception, LTB4-induced hypernociception was reduced by indomethacin (5 mg/kg), MK886 (3 mg/kg), celecoxib (10 mg/kg), antineutrophil antibody (100 mug, two doses), and fucoidan (20 mg/kg) treatments as well as in 5LO(-/-) mice. The production of LTB4 induced by zymosan in the joint was reduced by the pretreatment with fucoidan or antineutrophil antibody as well as the production of PGE2 induced by LTB4. Therefore, besides reinforcing the role of endogenous LTB4 as an important mediator of inflamed joint hypernociception, these results also suggested that the mechanism of LTB4-induced articular hypernociception depends on prostanoid and neutrophil recruitment. Furthermore, the results also demonstrated clearly that LTB4-induced hypernociception depends on the additional release of endogenous LTs. Concluding, targeting LTB4 synthesis/action might constitute useful therapeutic approaches to inhibit articular inflammatory hypernociception.

Topics: Animals; Arachidonate 5-Lipoxygenase; Benzopyrans; Carboxylic Acids; Cell Movement; Dinoprostone; Disease Models, Animal; Dose-Response Relationship, Drug; Indoles; Leukotriene B4; Male; Mice; Mice, Inbred C57BL; Mice, Knockout; Neutrophils; Temporomandibular Joint; Temporomandibular Joint Disorders; Time Factors; Zymosan

To investigate the role of leukotriene B4 (LTB4) and its receptor BLT1 in the pathogenesis of mouse uveitis.. Experimental autoimmune uveitis (EAU) was induced in B10RIII mice by immunization of interphotoreceptor retinoid binding protein (IRBP; peptide sequence 161-180) or in C57BL/6 (B6) mice by transfer of activated T cells specific for IRBP1-20. The animals were then treated with and without the BLT1 receptor antagonist, CP105696, at the disease onset after immunization or at day 0 or day 6 after T-cell transfer. EAU was also induced in wild-type B6 (WT) and BLT1-deficient (BLT1-/-) mice by reciprocal transfer of the T cells from B6 to BLT1-deficient mice and vise versa. Clinical signs of inflammation and ocular histology were compared. The chemotactic activity of LTB4 on naïve and IRBP-specific autoreactive T cells as well as effector leukocytes was examined.. The treatment of CP105696, greatly reduced the intensity of ongoing disease. IRBP1-20-specific T cells derived from wild-type B6 mice induced only mild uveitis in syngeneic BLT1-deficient mice and that IRBP1-20-specific T cells derived from BLT1-/- mice induced milder disease in wild-type B6 mice than those derived from wild-type B6 mice, suggesting that expression of the LTB4 receptor on both activated autoreactive T cells and effector leukocytes was necessary for ocular inflammation to occur. Consistent with these data, transfer of autoreactive T cells from B6 mice to 5-lipoxygenase-deficient (5-LO-/-) mice, which have a functional defect in LTB4 expression, also failed to induce uveitis in the recipient mice.. The results demonstrate a critical role for LTB4 in ocular inflammation and in the development and progression of EAU and suggest a new potential target for therapeutic intervention in this disease.

Topics: Adoptive Transfer; Animals; Arachidonate 5-Lipoxygenase; Autoimmune Diseases; Benzopyrans; Carboxylic Acids; Cells, Cultured; Chemotaxis, Leukocyte; Disease Models, Animal; Eye Proteins; Female; Immunization; Leukotriene B4; Lymphocyte Activation; Mice; Mice, Inbred BALB C; Mice, Knockout; Peptide Fragments; Purinergic P2 Receptor Antagonists; Receptors, Leukotriene B4; Receptors, Purinergic P2; Retinol-Binding Proteins; T-Lymphocytes; Uveitis

The production of nitric oxide (NO) by gamma interferon (IFN-gamma)-activated macrophages is a major effector mechanism during experimental Trypanosoma cruzi infection. In addition to IFN-gamma, chemoattractant molecules, such as platelet-activating factor (PAF) and CC chemokines, may also activate macrophages to induce NO and mediate the killing of T. cruzi in an NO-dependent manner. Here we investigated the ability of leukotriene B(4) (LTB(4)) to induce the production of NO by macrophages infected with T. cruzi in vitro and whether NO mediated LTB(4)-induced parasite killing. The activation of T. cruzi-infected but not naive murine peritoneal macrophages with LTB(4) induced the time- and concentration-dependent production of NO. In addition, low concentrations of LTB(4) acted in synergy with IFN-gamma to induce NO production. The NO produced mediated LTB(4)-induced microbicidal activity in macrophages, as demonstrated by the inhibitory effects of an inducible NO synthase inhibitor. LTB(4)-induced NO production and parasite killing were LTB(4) receptor dependent and were partially blocked by a PAF receptor antagonist. LTB(4) also induced significant tumor necrosis factor alpha (TNF-alpha) production, and blockade of TNF-alpha suppressed LTB(4)-induced NO release and parasite killing. A blockade of LTB(4) or PAF receptors partially inhibited IFN-gamma-induced NO and TNF-alpha production but not parasite killing. Finally, daily treatment of infected mice with CP-105,696 was accompanied by a significantly higher level of blood parasitemia, but not lethality, than that seen in vehicle-treated animals. In conclusion, our results suggest a role for LTB(4) during experimental T. cruzi infection. Chemoattractant molecules such as LTB(4) not only may play a major role in leukocyte migration into sites of inflammation in vivo but also, in the event of an infection, may play a relevant role in the activation of recruited leukocytes to kill the invading microorganism in an NO-dependent manner.

Topics: Animals; Benzopyrans; Carboxylic Acids; Cells, Cultured; Chagas Disease; Disease Models, Animal; Female; Immunity, Innate; Interferon-gamma; Leukotriene B4; Macrophage Activation; Macrophages, Peritoneal; Mice; Mice, Inbred BALB C; Nitric Oxide; Platelet Activating Factor; Receptors, Leukotriene B4; Time Factors; Trypanosoma cruzi; Tumor Necrosis Factor-alpha

Pharmacological strategies which limit neutrophil recruitment may also limit the damage induced by the reperfusion of an ischemic vascular territory. In the present study, we have investigated the effects of the BLT receptor antagonist, CP-105,696 ((+)-1-(3S,4R)-[3-(4-phenyl-benzyl)-4-hydroxy-chroman-7-yl]-cyclopentane carboxylic acid), on the local, remote and systemic inflammatory changes observed during severe intestinal ischemia (120 min) and reperfusion (120 min) injury. The post-ischemic treatment with CP-105,696 (3 mg/kg) virtually abolished the increase in vascular permeability, but not neutrophil accumulation, in the intestine and lungs. CP-105,696 partially inhibited the reperfusion-induced neutropenia, but failed to affect intestinal haemorrhage or lethality. CP-105,696 had no inhibitory effect on the local and systemic increases in the concentrations of tumour necrosis factor (TNF-alpha), interleukin-1 beta and interleukin-10, but markedly suppressed interleukin-6. Overall, our results show that activation of BLT receptor plays a minor role in the local, remote and systemic injuries following severe ischemia and reperfusion in rats.

Topics: Animals; Benzopyrans; Capillary Permeability; Carboxylic Acids; Disease Models, Animal; Interleukin-1; Interleukin-10; Interleukin-6; Intestinal Mucosa; Intestines; Lung; Male; Neutrophils; Rats; Rats, Wistar; Receptors, Leukotriene B4; Reperfusion Injury; Time Factors; Tumor Necrosis Factor-alpha

Reperfusion of ischemic vascular beds may lead to recruitment and activation of leukocytes, release of mediators of the inflammatory process and further injury to the affected vascular bed and to remote sites. Neutrophils appear to play a major role in the pathophysiology of reperfusion injury. Amongst inflammatory mediators shown to activate neutrophils and induce their recruitment in vivo, much interest has been placed on the role of leukotriene (LT)B(4). Here, we have assessed the effects of the BLT receptor antagonist (+)-1-(3S, 4R)-[3-(4-phenyl-benzyl)-4-hydroxy-chroman-7-yl]-cyclopentane carboxylic acid (CP 105,696) in a model of neutrophil-dependent ischemia and reperfusion injury in the rat. The superior mesenteric artery was isolated and ischemia was induced by its total occlusion for 30 min. After 30 min of reperfusion, injury was assessed by evaluating the extravasation of Evans blue, an index of vascular permeability, and the levels of myeloperoxidase, an index of neutrophil accumulation, in the intestine, mesentery and lung. The neutrophil-dependence of the local (intestine and mesentery) and remote (lung) injury was confirmed by using fucoidin, a selectin blocker, and WT-3, an anti-CD18 monoclonal antibody. Post-ischemic treatment with CP 105,696 dose-dependently inhibited vascular permeability and neutrophil accumulation in the intestine and mesentery. CP 105,696 also blocked the vascular permeability changes, but not neutrophil accumulation, in the lungs after reperfusion injury. Virtually identical results were obtained with another BLT receptor antagonist, 1-(5-ethyl-2-hydroxy-4-(6-methyl-6-(1H-tetrazol-5-yl)-heptoxy++ +)-phenyl )ethanone (LY255283). Our results suggest that post-ischemic treatment with BLT receptor antagonists may inhibit local and remote ischemia and reperfusion injury by blocking both the accumulation and/or activation of neutrophils.

Topics: Animals; Antibodies, Monoclonal; Benzopyrans; Capillary Permeability; Carboxylic Acids; CD18 Antigens; Disease Models, Animal; Dose-Response Relationship, Drug; Evans Blue; Intestinal Mucosa; Intestines; Leukotriene Antagonists; Lung; Male; Mesenteric Artery, Superior; Mesentery; Neutrophils; Peroxidase; Polysaccharides; Rats; Rats, Wistar; Receptors, Leukotriene B4; Reperfusion Injury; Tetrazoles