RESUMO
The mechanisms through which Candida albicans is recognized by immune cells and how it triggers host defence are not completely understood. In this study, we evaluated the effect of Concanavalin-A on the clearance of C. albicans by infected mice and their production of proinflammatory cytokine tumor necrosis factor-alpha (TNF-alpha). Subgroups of 5 animals were pretreated with Con-A (250 mug mL(-1) PBS) and after 96 h were infected intraperitoneally with 10(7) cells of C. albicans CR15 (an isolate from a HIV+ person); 30 min, 2, 6, 24 or 72 h after infection the mice were sacrificed. Phagocytosis of C. albicans by peritoneal macrophages increased 30 min after infection in mice pretreated with Con-A. The liver presented the greatest number of CFUs, and this number was reduced by pretreatment with Con-A. Control animals infected with C. albicans presented a significant increase in plasmatic alanine aminotransferase, which was not observed in mice treated with Con-A. Two hours after infection the production of TNF-alpha in the liver of mice pretreated with Con-A was significantly increased. These results suggest that a single dose of Con-A caused a beneficial modulating action of the inflammatory response during infection with C. albicans.
Assuntos
Candida albicans/imunologia , Candidíase/imunologia , Concanavalina A/administração & dosagem , Fatores Imunológicos/administração & dosagem , Fígado/microbiologia , Alanina Transaminase/sangue , Animais , Candida albicans/crescimento & desenvolvimento , Candidíase/tratamento farmacológico , Candidíase/microbiologia , Candidíase/patologia , Contagem de Colônia Microbiana , Modelos Animais de Doenças , Histocitoquímica , Fígado/imunologia , Fígado/patologia , Macrófagos Peritoneais/imunologia , Camundongos , Fagocitose , Análise de Sobrevida , Fator de Necrose Tumoral alfa/biossínteseRESUMO
In this study, we investigated the effect of concanavalin-A (Con-A) on the activation of phagocytosis and killing of Candida albicans by peritoneal macrophages from suckling and adult mice. Pretreatment of adult mice with Con-A dose-dependently increased the percentage of macrophages phagocytosing C. albicans in vitro from 3.8 +/- 0.9 to 24.2 +/- 2.4 in the absence of serum opsonins. Addition of mannan (50 microg) and mannose (50 mM) to the incubation medium reduced phagocytosis from 21.5 +/- 1.3 to 4.7 +/- 1.9, suggesting that treatment with Con-A increased phagocytosis mediated by mannose receptors. Killing of C. albicans was also increased by increasing the dose of Con-A. Pretreatment of suckling mice with Con-A increased the macrophages' phagocytic and candidacidal activities by an amount similar to that observed in adult mice. Furthermore, suckling mice pretreated with Con-A survived an intraperitoneal inoculum of 5 x 10(7) C. albicans, whereas all control mice died within 24-48 h of infection. This suggested that increased phagocytosis and killing of C. albicans stimulated by the action of Con-A conferred early protection upon suckling mice experimentally infected with C. albicans.
Assuntos
Concanavalina A/farmacologia , Lectinas Tipo C , Ativação de Macrófagos/efeitos dos fármacos , Macrófagos Peritoneais/efeitos dos fármacos , Lectinas de Ligação a Manose , Fagocitose/efeitos dos fármacos , Animais , Animais Lactentes , Candida albicans/imunologia , Candida albicans/fisiologia , Técnicas de Cultura de Células , Lectinas/farmacologia , Macrófagos Peritoneais/imunologia , Macrófagos Peritoneais/microbiologia , Receptor de Manose , Fagocitose/imunologia , Receptores de Superfície Celular/imunologiaRESUMO
1. Ingestion of enteropathogenic Escherichia coli or Candida albicans by thioglycollate-elicited macrophages and polymorphonuclear leukocytes was investigated in vitro, 2. Goat antiserum against mannose receptors caused about 50% inhibition of E. coli phagocytosis and about 90% inhibition of C. albicans phagocytosis. 3. E. coli and C. albicans uptake was inhibited by about 60% and 98%, respectively, by plating the macrophages onto substrates coated with poly-L-lysine-mannan. Further addition of 50 mM mannose to the medium significantly increased the inhibition of phagocytosis of E. coli by macrophages from 60.7 +/- 1.5 to 79.8 +/- 13.1 and by polymorphonuclear cells from 58.9 +/- 3.7 to 88.7 +/- 4.9. 4. Preincubation of phagocytic cells with antiserum against substance A of human erythrocytes reduced E. coli ingestion by 95%, but this inhibition was not observed when the antiserum was incubated with N-acetylgalactosamine (50 mM) before being added to the phagocytes. The phagocytosis of C. albicans was not inhibited by anti-substance A antiserum. 5. The phagocytosis of E. coli was inhibited by about 25% by the addition of 7.8 micrograms/ml soluble mannan to the medium, and by about 50% by the addition of 50 mMN-acetylgalactosamine; when both substances were added to the medium, an additive inhibition of about 75% was observed. 6. These results indicate that mannose receptors on the surface of phagocytic cells mediate E. coli or Candida albicans uptake and that the binding of bacteria to N-acetylgalactosamine residues from the membrane of phagocytes is also involved in the phagocytosis of E. coli
Assuntos
Animais , Humanos , Candida albicans/imunologia , Escherichia coli/imunologia , Fagocitose/imunologia , Receptores Mitogênicos/imunologia , Acetilgalactosamina/farmacologia , Aderência Bacteriana/efeitos dos fármacos , Aderência Bacteriana/imunologia , Candida albicans/efeitos dos fármacos , Candida albicans/patogenicidade , Depressão Química , Eritrócitos/efeitos dos fármacos , Eritrócitos/imunologia , Escherichia coli/efeitos dos fármacos , Escherichia coli/patogenicidade , Soros Imunes/farmacologia , Macrófagos Peritoneais/efeitos dos fármacosRESUMO
In the present study we have documented the use of the reagent p-benzoquinone (PBQ) for the spectrophotometric determination of total protein in blood plasma. Since the products of reaction are stable for several hours at room temperature after the 20-min boiling step, the time at which absorbance is measured is not a critical factor. Common anticoagulants such as EDTYA, citrate, or heparin do not interfere with the PBQ method at concentrations used in clinical laboratories. The products of the reaction between PBQ and either plasma (specific absorbance 2.33 x 10-3 ñ 0.20 x 10-3 ug cm -2) or purified proteins (specific absorbance 2.61 x 10-3 ñ 0.31 x 10-3 ug cm-2) show an absorption band at 350 nm, which follows Beer's law, and therefore can be used for analytical purposes. The PBQ method has a lower limit of detection (4 ug/ml) than that of biuret method (45 yg/ml) for a final reaction mixture of 5.0 and 4.2 ml, respectively