Navegando por Palavras-chave "endothelial cell"
Agora exibindo 1 - 6 de 6
Resultados por página
Opções de Ordenação
- ItemSomente MetadadadosAntimicrobial Action of Biguanides on the Viability of Acanthamoeba Cysts and Assessment of Cell Toxicity(Assoc Research Vision Ophthalmology Inc, 2013-09-01) Mafra, Cecilia Sales Pires [UNIFESP]; Carrijo-Carvalho, Linda Christian [UNIFESP]; Chudzinski-Tavassi, Ana Marisa; Taguchi, Felipe Marques de Carvalho [UNIFESP]; Foronda, Annette Silva [UNIFESP]; Carvalho, Fabio Ramos de Souza [UNIFESP]; Freitas, Denise de [UNIFESP]; Universidade Federal de São Paulo (UNIFESP); Butantan InstPURPOSE. To assess dose- and concentration-dependent rates of biguanides on the viability of Acanthamoeba cysts isolated from severe ulcerative keratitis, and to correlate cysticidal activites with cytotoxic profiles in corneal and endothelial cells.METHODS. Cysticidal activities of polyhexamethylene biguanide and chlorhexidine digluconate were evaluated in the Acanthamoeba castellanii strain and clinical isolates of Acanthamoeba spp obtained from two severe and recurrent cases of ulcerative keratitis. the molecular characterization of protozoa used in the experimental assays was performed by sequencing reactions of the 18S rDNA gene. Acanthamoeba cysts were exposed at different dosages and concentrations of both biguanides; the application of double-biguanides was also evaluated. Automated cell viability assessment of cysts was performed using the trypan blue dye exclusion method. Cytotoxicity assays of biguanides were conducted using primary cultures of endothelial cells alone or in coculture with Acanthamoeba cysts. Human corneal epithelial cells were used as a comparative pattern to assess the toxicity of biguanide compounds. Cell viability was measured using both quantitative and qualitative methods. Statistical analyses were applied to the data.RESULTS. the in vitro study showed that all dosages, concentrations, and combinations of biguanides tested had a cysticidal effect on Acanthamoeba spp strains tested compared with control cultures not exposed to any antimicrobials; the difference in response was statistically significant. the use of both biguanides in combination demonstrated the best cysticidal effect. the use of isolated biguanides was associated with greater cytotoxic effects than with biguanides used in combination. Chlorhexidine digluconate used alone tended to have greater cytotoxicity than polyhexamethylene biguanide. Furthermore, the double-biguanide application had a statistically significant decrease in the deleterious effect on endothelial cells at higher dosage and concentration. Quantitative and qualitative analyses demonstrated the toxic effect of biguanide compounds on the viability of corneal epithelial cells, under single or in combination usage.CONCLUSIONS. We demonstrated that the combined use of biguanides had greater cysticidal activity than individual drug application as well as a possible protective effect on endothelial cells. the biguanide compounds tested were able to induce corneal epithelial cell death in time and concentration-independent fashions. Findings support the hypothesis concerning the cysticidal effect and the differential patterns of toxicity expressed by polyhexamethylene biguanide and chlorhexidine digluconate on the endothelial and corneal cells.
- ItemAcesso aberto (Open Access)The blockade of cyclooxygenases-1 and-2 reduces the effects of hypoxia on endothelial cells(Assoc Bras Divulg Cientifica, 2006-09-01) Gloria, Maria Aparecida da [UNIFESP]; Cenedeze, Marcos Antonio [UNIFESP]; Pacheco-Silva, Alvaro [UNIFESP]; Camara, Niels Olsen Saraiva [UNIFESP]; Universidade Federal de São Paulo (UNIFESP); Universidade de São Paulo (USP)Hypoxia activates endothelial cells by the action of reactive oxygen species generated in part by cyclooxygenases (COX) production enhancing leukocyte transmigration. We investigated the effect of specific COX inhibition on the function of endothelial cells exposed to hypoxia. Mouse immortalized endothelial cells were subjected to 30 min of oxygen deprivation by gas exchange. Acridine orange/ ethidium bromide dyes and lactate dehydrogenase activity were used to monitor cell viability. the mRNA of COX-1 and -2 was amplified and semi-quantified before and after hypoxia in cells treated or not with indomethacin, a non-selective COX inhibitor. Expression of RANTES ( regulated upon activation, normal T cell expressed and secreted) protein and the protective role of heme oxygenase-1 (HO-1) were also investigated by PCR. Gas exchange decreased partial oxygen pressure (PaO2) by 45.12 +/- 5.85% (from 162 +/- 10 to 73 +/- 7.4 mmIIg). Thirty minutes of hypoxia decreased cell viability and enhanced lactate dehydrogenase levels compared to control (73.1 +/- 2.7 vs 91.2 +/- 0.9%, P < 0.02; 35.96 +/- 11.64 vs 22.19 +/- 9.65%, P = 0.002, respectively). COX-2 and HO-1 mRNA were up-regulated after hypoxia. Indomethacin (300 mu M) decreased COX-2, HO-1, hypoxiainducible factor-1 alpha and RANTES mRNA and increased cell viability after hypoxia. We conclude that blockade of COX up-regulation can ameliorate endothelial injury, resulting in reduced production of chemokines.
- ItemSomente MetadadadosCytotoxic activity and degradation patterns of structural proteins by corneal isolates of Acanthamoeba spp(Springer, 2015-01-01) Sant'ana, Viviane Peracini [UNIFESP]; Carrijo-Carvalho, Linda Christian [UNIFESP]; Foronda, Annette Silva [UNIFESP]; Chudzinski-Tavassi, Ana Marisa; Freitas, Denise de [UNIFESP]; Souza de Carvalho, Fabio Ramos [UNIFESP]; Universidade Federal de São Paulo (UNIFESP); Butantan InstProteolytic enzymes secreted by trophozoites (amoebic secretome) are suggested as the main virulence factor involved in the severity of Acanthamoeba keratitis. the degradation profile of the main glycoprotein components of anterior and posterior portions of the cornea and the cytopathic effect of secretomes on endothelial cells by contact-independent mechanism were evaluated.Trophozoites were isolated primarily from corneal tissue samples (n = 11) and extracellular proteins were collected from axenic cell culture supernatants. the molecular weights of proteolytic enzymes were estimated by zymography. Enzymatic cleavage of laminin and fibronectin substrates by amoebic secretome was investigated and cluster analysis was applied to the proteolysis profiles. Primary cultures of endothelial cells were used in both qualitative and quantitative assays of cytophatogenicity.Differential patterns of proteolysis were observed among the Acanthamoeba secretomes that were analysed. the uniformity of laminin degradation contrasted with the diversity of the proteolysis profiles observed in the fibronectin substrate. Acanthamoeba secretome extracted from four clinical isolates was shown to be toxic when in contact with the endothelial cell monolayer (p < 0.01). Induction of apoptosis and membrane permeability, at different percentual values, were suggested as the main mechanisms that could induce endothelial cell death when in contact with amoebic secretome.Our results provide evidence that virulence factors secreted by Acanthamoeba trophozoites can be related to an increased pathogenicity pattern by an independent contact-trophozoite mechanism, through induction of endothelial cell death by apoptosis at a higher percentage than providing the lack of cell viability by the membrane-associated pore-forming toxin activity.
- ItemSomente MetadadadosA heparin mimetic isolated from a marine shrimp suppresses neovascularization(Wiley-Blackwell, 2010-08-01) Dreyfuss, Juliana Luporini [UNIFESP]; Regatieri, Caio Vinicius Saito [UNIFESP]; Lima, M. A. [UNIFESP]; Paredes-Gamero, E. J. [UNIFESP]; Brito, A. S. [UNIFESP]; Chavante, S. F.; Belfort, Rubens Junior [UNIFESP]; Farah, M. E. [UNIFESP]; Nader, Helena Bonciani [UNIFESP]; Universidade Federal de São Paulo (UNIFESP); Univ Fed Rio Grande do NorteBackground: Choroidal neovascularization (CNV) is the main cause of severe visual loss in age-related macular degeneration (AMD). Heparin/heparan sulfate are known to play important roles in neovascularization due to their abilities to bind and modulate angiogenic growth factors and cytokines. Previously, we have isolated from marine shrimp a heparin-like compound with striking anti-inflammatory action and negligible anticoagulant and hemorrhagic activities. Objectives: To investigate the role of this novel heparin-like compound in angiogenic processes. Methods and Results: the anti-angiogenic effect of this heparinoid in laser-induced CNV and in vitro models is reported. the compound binds to growth factors (FGF-2, EGF and VEGF), blocks endothelial cell proliferation and shows no cytotoxic effect. the decrease in proliferation is not related to cell death either by apoptosis or secondary necrosis. the results also showed that the heparinoid modified the 2-D network organization in capillary-like structures of endothelial cells in Matrigel and reduced the CNV area. the effect on CNV area correlates with decreases in the levels of VEGF and TGF-beta 1 in the choroidal tissue. the low content of 2-O-sulfate groups in this heparinoid may explain its potent anti-angiogenic effect. Conclusions: the properties of the shrimp heparinoid, such as potent anti-angiogenic and anti-inflammatory activities but insignificant anticoagulant or hemorrhagic actions, point to this compound as a compelling drug candidate for treating neovascular AMD and other angioproliferative diseases. A mechanism for the anti-angiogenic effect of the heparinoid is proposed.
- ItemSomente MetadadadosInflammation-induced modulation of cellular galectin-1 and-3 expression in a model of rat peritonitis(Birkhauser Verlag Ag, 2006-03-01) Gil, C. D.; Cooper, D.; Rosignoli, G.; Perretti, M.; Oliani, S. M.; UNESP; Universidade de São Paulo (USP); Universidade Federal de São Paulo (UNIFESP); Queen Mary Sch Med & DentObjective and design: To investigate the effect of galectin-1 (Gal-1) and -3 (Gal-3) on leukocyte migration and analyze the expression of both galectins in inflammatory cells using a model of rat peritonitis.Material or Subjects: Sprague-Dawley rats (n = 4 per group).Treatment: Peritonitis was induced in animals through intraperitoneal injection of carrageenin (1.5 mg/kg) and rat mesenteries were analyzed at different time points (0, 4, 24 and 48h). for pharmacological treatment, rats received intravenous injection of Gal-1 or -3 (3 mu g/kg) followed by carrageenin.Methods: Western blotting and immunoelectron microscopy analysis. Statistical analysis was performed using ANOVA followed by Bonferroni test.Results: Pharmacological treatment with Gal-1, but not Gal-3, inhibited (similar to 50%) leukocyte recruitment into the peritoneal cavity at 4h time-point. in this early phase, immunogold staining of mesenteries showed a diminished Gal-3 expression in degranulated mast cells and Gal-1 in transmigrated neutrophils (similar to 20% reduction compared to intravascular cells). in the later phases (24 and 48 h), leukocyte turnover was associated with augmented Gal-1 expression in neutrophils and macrophages and Gal-3 in mast cells and macrophages.Conclusions: These results point to a balanced expression of cell-associated-Gal-1/Gal-3 and might impact on the development of new therapeutic strategies for inflammatory diseases.
- ItemSomente MetadadadosInsularinase A, a prothrombin activator from Bothrops insularis venom, is a metalloprotease derived from a gene encoding protease and disintegrin domains(Walter de Gruyter Gmbh, 2005-06-01) Modesto, JCD; Junqueira-de-Azevedo, ILM; Neves-Ferreira, AGC; Fritzen, M.; Oliva, MLV; Ho, P. L.; Perales, J.; Chudzinski-Tavassi, A. M.; Inst Butantan; Inst Burantan; Fiocruz MS; Universidade Federal de São Paulo (UNIFESP); Universidade de São Paulo (USP)The first low-molecular-mass metalloprotease presenting prothrombin activating activity was purified from Bothrops insularis venom and named insularinase A. It is a single-chain protease with a molecular mass of 22 639 Da. cDNA sequence analysis revealed that the disintegrin domain of the precursor protein is post-translationally processed, producing the mature insularinase A. Analysis of its deduced amino acid sequence showed a high similarity with several fibrin(ogen)olytic metalloproteases and only a moderate similarity with prothrombin activators. However, SIDS-PAGE of prothrombin after activation by insularinase A showed fragment patterns similar to those generated by group A prothrombin activators, which convert prothrombin into meizothrombin independently of the prothrombinase complex. in addition, insularinase A activates factor X and hydrolyses fibrinogen and fibrin. Chelating agents fully inhibit all insularinase A activities. Insularinase A induced neither detachment nor apoptosis of human endothelial cells and was also not able to trigger an endothelial proinflammatory cell response. Nitric oxide and prostacyclin levels released by endothelial cells were significantly increased after treatment with insularinase A. Our results show that, although its primary structure is related to class P-I fibrin(ogen)olytic metalloproteases, insularinase A is functionally similar to group A prothrombin activators.