RESUMO
Escherichia coli strain 15 (ATCC 9723), which forms robust biofilms, was grown under optimal biofilm conditions in NaCl-free Luria-Bertani broth (LB*) or in LB* supplemented with one of the non-metabolizable analogues 2-deoxy-d-glucose (2DG), methyl α-d-mannopyranoside (αMM), or methyl α-d-glucopyranoside (αMG). Biofilm growth was inhibited by mannose analogue 2DG even at very low concentration in unbuffered medium, and the maximal inhibition was enhanced in the presence of either 100 mM KPO4 or 100 mM MOPS, pH 7.5; in buffered medium, concentrations of 0.02 % (1.2 mM) or more inhibited growth nearly completely. In contrast, mannose analogue αMM, which should not be able to enter the cells but has been reported to inhibit biofilm growth by binding to FimH, did not exhibit strong inhibition even at concentrations up to 1.8 % (108 mM). The glucose analogue αMG inhibited biofilm growth, but much less strongly than did 2DG. None of the analogues inhibited planktonic growth or caused a change in pH of the unbuffered medium. Similar inhibitory effects of the analogues were observed in minimal medium. The effects were not strain-specific, as 2DG and αMG also inhibited the weak biofilm growth of E. coli K12.
Assuntos
Antimetabólitos/farmacologia , Biofilmes/crescimento & desenvolvimento , Desoxiglucose/farmacologia , Escherichia coli/crescimento & desenvolvimento , AMP Cíclico/farmacologia , Escherichia coli/efeitos dos fármacos , Glucose-6-Fosfato/farmacologia , Metilglucosídeos/farmacologia , Metilmanosídeos/farmacologiaRESUMO
Genetic factors influence susceptibility to Paracoccidioidomycosis, a Latin American endemic mycosis. The pattern of susceptibility of congenic mouse strains infected with Paracoccidioides brasiliensis resembles the pattern of the Nramp1 gene. Thus, congenic murine bone-marrow-derived macrophage lines B10R (Nramp1rGly169) and B10S (null Nramp1 protein expression, Nramp1sAsp169) were infected with P. brasiliensis conidia and compared, under opsonic and nonopsonic conditions. Opsonization increased the percentage of phagocytosis by both cell lines. B10R macrophages exhibited a higher percentage of cells with associated conidia and higher number of conidia per macrophage than B10S. Heat-inactivation and EDTA treatment of serum used for opsonization, and treatment of macrophages with anti-complement receptor 3 (CR3) decreased phagocytosis by both cell lines. alpha-methyl-d-mannoside reduced phagocytosis by B10R macrophages, suggesting that the mannose receptor participates in phagocytosis by these cells. The CR3 expression was similar on both cell lines and B10R expressed more mannose receptors, but neither cell line expressed CR1. IFNgamma decreased the conversion of conidia to the yeast form of P. brasiliensis in B10R, but not in B10S macrophages.
Assuntos
Proteínas de Transporte de Cátions/imunologia , Complemento C3/imunologia , Lectinas Tipo C/imunologia , Macrófagos/imunologia , Lectinas de Ligação a Manose/imunologia , Paracoccidioides/imunologia , Receptores de Superfície Celular/imunologia , Animais , Antígeno CD11b/biossíntese , Antígeno CD11b/imunologia , Proteínas de Transporte de Cátions/genética , Linhagem Celular , Complemento C3/metabolismo , Predisposição Genética para Doença , Lectinas Tipo C/metabolismo , Antígeno de Macrófago 1/biossíntese , Antígeno de Macrófago 1/imunologia , Macrófagos/efeitos dos fármacos , Receptor de Manose , Lectinas de Ligação a Manose/metabolismo , Metilmanosídeos/farmacologia , Camundongos , Camundongos Congênicos , Paracoccidioides/genética , Paracoccidioidomicose/genética , Paracoccidioidomicose/imunologia , Paracoccidioidomicose/microbiologia , Fagocitose/efeitos dos fármacos , Fagocitose/genética , Fagocitose/fisiologia , Receptores de Superfície Celular/metabolismo , Receptores de Complemento 3b/antagonistas & inibidores , Receptores de Complemento 3b/imunologiaRESUMO
Identification of mycobacterial adhesins is needed to understand better the pathogenesis of tuberculosis and to develop new strategies to fight this infection. In this work, THP-1 monocytic cells were incubated with Mycobacterium tuberculosis culture filtrate proteins labelled with biotin and a dominant 19-kDa adhesin was found. This adhesin was characterized as the glycosylated and acylated 19-kDa antigen (Rv 3763). These findings were confirmed in assays with culture filtrate proteins and cell-wall fractions from a recombinant Mycobacterium smegmatis strain that overexpresses the 19-kDa antigen. Further, fluorescent microspheres coated with recombinant culture filtrate proteins adhere to cells in higher numbers than microspheres coated with native M. smegmatis proteins. The binding of the 19-kDa antigen to cells was inhibited with mannose receptor competitor sugars, Ca(2+) chelators and with a monoclonal antibody to the human mannose receptor. Phagocytosis assays showed high-level binding of bacilli to THP-1 cells that was inhibited with alpha-methyl-mannoside, mannan, EDTA and mAbs to the mannose receptor and to the 19-kDa M. tuberculosis antigen. Immunoprecipitation, cell-surface ELISA and immunostaining confirmed the expression of the mannose receptor by THP-1 cells. In conclusion, here we show that the macrophage mannose receptor, considered a pathogen pattern recognition receptor, may interact with mannose residues of mycobacterial glycoproteins that could promote the phagocytosis of mycobacteria.
Assuntos
Adesinas Bacterianas/imunologia , Lectinas Tipo C/imunologia , Lectinas de Ligação a Manose/imunologia , Monócitos/imunologia , Mycobacterium tuberculosis/imunologia , Fagocitose/imunologia , Receptores de Superfície Celular/imunologia , Tuberculose/microbiologia , Acetilglucosamina/farmacologia , Adesinas Bacterianas/metabolismo , Antígenos de Bactérias/imunologia , Aderência Bacteriana , Ensaio de Imunoadsorção Enzimática , Humanos , Immunoblotting , Imunoprecipitação , Lectinas Tipo C/metabolismo , Mananas/farmacologia , Receptor de Manose , Lectinas de Ligação a Manose/metabolismo , Metilmanosídeos/farmacologia , Monócitos/metabolismo , Monócitos/microbiologia , Mycobacterium tuberculosis/metabolismo , Ligação Proteica/imunologia , Receptores de Superfície Celular/metabolismoRESUMO
Previous studies have shown that Trichomonas vaginalis are capable of ingesting bacteria. This observation was confirmed in the present study and extended to Tritrichomonas foetus. Using a special strain of Escherichia coli grown under conditions that produced fimbriae presenting a lectin-like molecule recognizing mannose, we showed that the bacteria attached to and were ingested by trichomonads through a mechanism involving cell-to-cell recognition. Absence of the fimbriae or addition of alpha-methyl-D-mannoside to the interaction medium blocked attachment of the bacteria to the protozoa cell surface. Ingested bacteria were later digested within the cytoplasmic vacuole.
Assuntos
Aderência Bacteriana/fisiologia , Carboidratos/fisiologia , Escherichia coli/fisiologia , Trichomonas vaginalis/microbiologia , Tritrichomonas foetus/microbiologia , Animais , Aderência Bacteriana/efeitos dos fármacos , Escherichia coli/ultraestrutura , Fímbrias Bacterianas , Lectinas , Manose , Metilmanosídeos/farmacologia , Coloração Negativa , Trichomonas vaginalis/ultraestrutura , Tritrichomonas foetus/ultraestrutura , Leveduras/fisiologiaRESUMO
Lectins from Dioclea grandiflora (DG) and Canavalia brasiliensis (CB) were compared with Concanavalin A (ConA) for their ability to induce paw edema and peritoneal cell immigration in rats. ConA caused a slight edema with a peak at 1 h after injection, while DG or CB induced a pronounced and long-lasting edema that reached a maximum at about 6 h. Different antiinflammatory drugs partially inhibited the edema. alpha-D-glucose (GLU) partially blocked the edema caused by ConA and markedly inhibited that due to CB, but had no effect on the edema induced by DG. alpha-Methyl mannoside (alpha-MM) blocked the edema caused by DG and ConA, but did not affect that caused by CB. At doses much lower than those used to induce paw edema, the lectins promoted an intense accumulation of neutrophil and mononuclear cells in the rat peritoneal cavity. CB and DG were more potent than ConA, which also presented a different profile of cell immigration. GLU significantly inhibited leukocyte accumulation caused by all lectins. alpha-MM impaired ConA- and DG-induced cell immigration, but only partially inhibited CB. Thus, despite their physicochemical similarities with ConA, DG and CB have more powerful pro-inflammatory effects. This difference seems to be related to their sugar-binding properties. However, while ConA- and DG-induced effects were inhibited more by alpha-MM than by GLU, CB-induced effects were inhibited more by glucose.
Assuntos
Concanavalina A/toxicidade , Edema/induzido quimicamente , Lectinas/toxicidade , Leucócitos/fisiologia , Lectinas de Plantas , Animais , Anti-Inflamatórios/farmacologia , Movimento Celular , Glucose/farmacologia , Membro Posterior , Metilmanosídeos/farmacologia , Cavidade Peritoneal , Ratos , Ratos WistarRESUMO
The hemagglutinating activity of a crude extract and partially purified protein fractions from seeds of Dolichos lablab grown in the state of Bahia, Brazil, has been examined. The crude extract agglutinates rabbit erythrocytes non-specifically with respect to blood groups. The hemagglutination of rabbit erythrocytes induced by the crude extract of D. lablab was inhibited most effectively by N-acetyl-D-glucosamine (3 mM) and alpha-methyl-D-mannoside (3 mM) followed by D-glucosamine (6 mM), D-mannose (6 mM) and D-glucose (12 mM). The spectrum of sugar inhibition of the hemagglutinating activity of the Brazilian seeds combines the characteristics of seeds coming from Turkey and India as indicated by the preferential inhibitory effect exerted by N-acetyl-D-glucosamine and the inhibition by D-mannose and D-glucose. These data suggest that the Brazilian seeds may belong to a different variety of Dolichos lablab.