TH17 cells, interleukin-17 and interferon-γ in patients and households contacts of leprosy with multibacillary and paucibacillary forms before and after the start of chemotherapy treatment.

BACKGROUND: Studies on the immunology of leprosy are fundamental to the understanding of the various forms of clinical manifestation of the disease. In some diseases, lymphocytes TH17 and one of its key cytokines, interleukin-17 has been shown to be essential in developing an effective immune response. In leprosy, involvement of lymphocyte TH17 and interleukin-17 remains understudied. OBJECTIVES: This study is the first investigation to examine the association between TH17 cells, interleukin-17 and interferon- γ in patients and households contacts of leprosy. METHODS: To document the participation of TH17 cells and interleukin-17 in the immunology of leprosy, to observe the behavior of interferon-γ in relation to interleukin-17 and to verify the differences found between individuals paucibacillary, multibacillary and household contacts, we analyzed samples peripheral blood to identify TH-17 cells, interleukin-17 and IFN-γ; establishing relationships between all the groups. RESULTS: There was a significant difference in the results found in the comparison between the paucibacillary and multibacillary groups of patients (P < 0.001), as well with the household contacts (P < 0.005). The polychemotherapeutic treatment modified the profile of immune response in multibacillary patients compared to what was observed before the start of treatment. CONCLUSION: The principal finding was that TH17 lymphocytes and interleukin-17 actively participating in the immune response of Hansen's disease as well these cells can stimulate the cellular immunity.

Quantitative evaluation of Listeria monocytogenes in fresh and processed surubim fish (Pseudoplatystoma sp) / Avaliação quantitativa de Listeria monocytogenes em surubim (Pseudoplatystoma sp) fresco e processado

L. monocytogenes is a foodborne psychrotrophic bacterial pathogen of special importance for minimally processed foods. In this work, it was enumerated in samples of surubim fish by MPN technique. The population of L. monocytogenes was estimated as < 0.012 MPN/cm² in fresh and < 0.03 MPN/g in minimally processed fish.

L. monocytogenes é um patógeno psicrotrófico transmitido por alimentos, de importância especial para alimentos minimamente processados. Neste trabalho, a bactéria foi enumerada em amostras de peixe surubim utilizando-se a técnica do NMP. A população de L. monocytogenes foi estimada como < 0.012 NMP/cm² do peixe fresco e < 0.03 NMP/g do peixe minimamente processado.

Quantitative evaluation of Listeria monocytogenes in fresh and processed surubim fish (Pseudoplatystoma sp).

L. monocytogenes is a foodborne psychrotrophic bacterial pathogen of special importance for minimally processed foods. In this work, it was enumerated in samples of surubim fish by MPN technique. The population of L. monocytogenes was estimated as < 0.012 MPN/cm(2) in fresh and < 0.03 MPN/g in minimally processed fish.

Interference of methysergide, a specific 5-hydroxytryptamine receptor antagonist, with airway chronic allergic inflammation and remodelling in a murine model of asthma.

BACKGROUND: Airway remodelling encompasses the structural changes observed in asthmatic airways. Mast cells, through the release of histamine and 5-hydroxytryptamine (serotonin), are implicated in early asthmatic reactions, bronchoconstriction and mucosal oedema, and in the development of bronchial hyperresponsiveness. However, the association between serotonin and remodelling processes in murine model of airways inflammation remains to be elucidated. OBJECTIVE: As serotonin is released by murine mast cells upon antigen challenge, we tested the hypothesis of its involvement in the development of inflammatory and remodelling processes in a murine model of chronic airway inflammation following prolonged allergen challenge. Methods BALB/c mice were exposed to aerosolized ovalbumin for 20 min 2 days a week, for 4 consecutive weeks. Two hours before each challenge, they were treated with methysergide (intranasally, 40 microg/kg). Forty-eight hours after the last aerosol challenge, bronchoalveolar lavage (BAL) and lung tissue were collected for analysis. RESULTS: Methysergide inhibited the allergen-induced increase in airway eosinophilia, reduced T helper type 2 (Th2) cytokines in lung, spleen or thoracic lymph nodes, and specific IgE levels. The extravasation of plasma and fibronectin production in the lung, and collagen deposition in the lung were also inhibited after methysergide treatment. Although methysergide treatment induced an increase in IFN-gamma levels, experiments with neutralizing antibody suggest that this is not responsible for inhibition. In addition, instillation of serotonin to immunized mice induced eosinophil recruitment to BAL, Th2 cytokine production and fibronectin release in lung as well as collagen deposition. CONCLUSION: Serotonin may contribute to the development and maintenance of remodelling through the release of cytokines and of fibrogenic mediators. Serotonin should therefore be considered as relevant for the development and maintenance of airway remodelling.

[Buspirone chlorhydrate in the treatment of cerebellar ataxia]. / Clorhidrato de buspirona en el tratamiento de la ataxia cerebelosa.

AIM: To evaluate the efficiency of buspirone chlorhydrate in a group of patients who all presented sporadic or family primary cerebellar ataxia. PATIENTS AND METHODS: In this open label study of addiction, the following eligibility criteria were used: 1) Clinical: primary cerebellar ataxia; 2) Radiological: nuclear magnetic resonance showing pure cerebellar cortical atrophy; 3) Age: over 20 years old. Any patient with a history of food deficiency, alcoholism, neoplasic, infectious, degenerative and vascular diseases was excluded from taking part in the study. Of the 20 patients examined initially, 18 met the eligibility criteria, and of these 11 reached the end of the study, although drop outs were not related to treatment. Four patients had been diagnosed as suffering from cerebellar cortical atrophy of the sporadic type and the remaining seven had a family type cerebellar cortical atrophy. All the patients were thoroughly evaluated and received scores both at the beginning and at the end of the study, according to the modified Massaquoi scale for clinical evaluation of cerebellar functioning, and the Hamilton Anxiety Scale. They were all administered buspirone chlorhydrate in doses that progressively increased by 5 mg/month over a total period of 12 months. The maximum dose was considered to be 1 mg/kg body weight, without exceeding 60 mg. The increase in dosage was stopped if it was not well tolerated by the patient

Clorhidrato de buspirona en el tratamiento de la ataxia cerebelosa / Buspirone chlorhydrate in the treatment of cerebellar ataxia

Objetivo. Evaluar la eficacia del clorhidrato de buspirona en un grupo de pacientes que se caracteriza por presentar una ataxia cerebelar primaria de tipo esporádico o familiar. Pacientes y métodos. Se trata de un estudio prospectivo de adición (open label study).Se utilizaron los siguientes criterios de inclusión: 1) Clínico: ataxia cerebelar primaria; 2) Radiológico: resonancia magnética que mostró una atrofia cortical cerebelar pura; 3) Edad: por encima de los 20 años. Se excluyeron del estudio todos los pacientes con historial de deficiencia alimentaria, alcoholismo, enfermedades neoplásicas, infecciosas, degenerativas y vasculares. De los 20 pacientes examinados inicialmente, 18 cumplieron los criterios de inclusión. De éstos, tan sólo 11 llegaron hasta el final del estudio, sin que los abandonos estuvieran relacionados con el tratamiento. Cuatro pacientes tenían un diagnóstico de atrofia cortical cerebelar de tipo esporádica, y siete, de atrofia cortical cerebelar de tipo familiar. Todos los pacientes se evaluaron de manera minuciosa y recibieron puntuaciones tanto al inicio como al final del estudio, según la escala de Massaquoi modificada para la evaluación clínica de la función cerebelar y según la escala de Hamilton para la ansiedad. A todos los pacientes se les administró clorhidrato de buspirona, con un aumento progresivo de la dosis de 5 mg/mes, durante un total de 12 meses. La dosis máxima considerada fue de 1 mg/kg de peso, sin sobrepasar los 60 mg. La progresión de la dosis se interrumpió cuando el paciente no la toleró (AU)

Extracts of Ascaris suum egg and adult worm share similar immunosuppressive properties

Adult Ascaris suum body extract (Asc) prepared from male and female worms (with stored eggs) down-regulates the specific immune response of DBA/2 mice to ovalbumin (OA) and preferentially stimulates a Th2 response to its own components, which is responsible for the suppression of the OA-specific Th1 response. Here, we investigated the participation of soluble extracts prepared from male or female worms or from eggs (E-Asc) in these immunological events. Extracts from either sex (1 mg/animal) or E-Asc (0.35 or 1 mg protein/animal) suppressed the delayed-type hypersensitivity (DTH) reaction (60-85 percent), proliferative response (50-70 percent), IL-2 and IFN-gamma secretion (below detection threshold) and IgG1 antibody production (70-90 percent) of DBA/2 mice to OA. A dose of 0.1 mg E-Asc/animal did not change DTH or proliferation, but was as effective as 0.35 mg in suppressing IL-2 and IFN-gamma, and OA-specific IgG1 antibodies. Lymph node cells from DBA/2 mice injected with Asc (1 mg/animal) or a high dose of E-Asc (1 mg protein/animal) secreted IL-4 upon in vitro stimulation with concanavalin A. As previously demonstrated for Asc, the cytokine profile obtained with the E-Asc was dose dependent and changed towards Th1 when a low dose (0.1 mg protein/animal) was used. Taken together, these results suggest that adult worms of either sex and eggs induce the same type of T cell response and share similar immunosuppressive properties

Extracts of Ascaris suum egg and adult worm share similar immunosuppressive properties.

Adult Ascaris suum body extract (Asc) prepared from male and female worms (with stored eggs) down-regulates the specific immune response of DBA/2 mice to ovalbumin (OA) and preferentially stimulates a Th2 response to its own components, which is responsible for the suppression of the OA-specific Th1 response. Here, we investigated the participation of soluble extracts prepared from male or female worms or from eggs (E-Asc) in these immunological events. Extracts from either sex (1 mg/animal) or E-Asc (0.35 or 1 mg protein/animal) suppressed the delayed-type hypersensitivity (DTH) reaction (60-85%), proliferative response (50-70%), IL-2 and IFN-gamma secretion (below detection threshold) and IgG1 antibody production (70-90%) of DBA/2 mice to OA. A dose of 0.1 mg E-Asc/animal did not change DTH or proliferation, but was as effective as 0.35 mg in suppressing IL-2 and IFN-gamma, and OA-specific IgG1 antibodies. Lymph node cells from DBA/2 mice injected with Asc (1 mg/animal) or a high dose of E-Asc (1 mg protein/animal) secreted IL-4 upon in vitro stimulation with concanavalin A. As previously demonstrated for Asc, the cytokine profile obtained with the E-Asc was dose dependent and changed towards Th1 when a low dose (0.1 mg protein/animal) was used. Taken together, these results suggest that adult worms of either sex and eggs induce the same type of T cell response and share similar immunosuppressive properties.

Structural evidence that the methionyl aminopeptidase from Escherichia coli is a mononuclear metalloprotease.

The Co and Fe K-edge extended X-ray absorption fine structure (EXAFS) spectra of the methionyl aminopeptidase from Escherichia coli (EcMetAP) have been recorded in the presence of 1 and 2 equiv of either Co(II) or Fe(II) (i.e., [Co(II)_(EcMetAP)], [Co(II)Co(II)(EcMetAP)], [Fe(II)_(EcMetAP)], and [Fe(II)Fe(II)(EcMetAP)]). The Fourier transformed data of both [Co(II)_(EcMetAP)] and [Co(II)Co(II)(EcMetAP)] are dominated by a peak at ca. 2.05 A, which can be fit assuming 5 light atom (N,O) scatterers at 2.04 A. Attempts to include a Co-Co interaction (in the 2.4-4.0 A range) in the curve-fitting parameters were unsuccessful. Inclusion of multiple-scattering contributions from the outer-shell atoms of a histidine-imidazole ring resulted in reasonable Debye-Waller factors for these contributions and a slight reduction in the goodness-of-fit value (f '). These data suggest that a dinuclear Co(II) center does not exist in EcMetAP and that the first Co atom is located in the histidine-ligated side of the active site. The EXAFS data obtained for [Fe(II)_(EcMetAP)] and [Fe(II)Fe(II)(EcMetAP)] indicate that Fe(II) binds to EcMetAP in a similar site to Co(II). Since no X-ray crystallographic data are available for any Fe(II)-substituted EcMetAP enzyme, these data provide the first glimpse at the Fe(II) active site of MetAP enzymes. In addition, the EXAFS data for [Co(II)Co(II)(EcMetAP)] incubated with the antiangiogenesis drug fumagillin are also presented.

Interactions of Cdc4p, a myosin light chain, with IQ-domain containing proteins in Schizosaccharomyces pombe.

The fission yeast Schizosaccharomyces pombe undergoes cell division through a medially placed actomyosin-based contractile ring. One of the key components of this ring is the F-actin based motor protein myosin II. The myosin II heavy chain Myo2p has two light-chain-binding domains, IQl and IQ2, which bind the essential light chain, Cdc4p, and the regulatory light chain, Rlc1p. Previously, we have reported the characterization of cells expressing Myo2p lacking the IQ2 domain that facilitates Myo2p interaction with Rlc1p. In this study, we have created and characterized S. pombe strains carrying precise deletions of IQ1 and the entire neck region encompassing the IQ1 and IQ2 domains. Surprisingly, we found that the entire neck region of Myo2p is dispensable for Myo2p function. Cells deleted for IQ1, IQ2 and the entire neck region of Myo2p do not display any obvious cytoskeletal abnormalities. Immunofluorescence studies indicated that Cdc4p localizes at the ring in early and late mitotic cells in a strain in which interactions of Cdc4p with both the myosin II heavy chains (Myo2p and Myp2p) are abolished. Unlike mutations in Rlc1p that are suppressed by a simultaneous deletion of its binding site on Myo2p, mutations in the essential light chain Cdc4p are not suppressed by deletion of its binding sites on Myo2p, suggesting that Cdc4p may have additional partners essential for cytokinesis. Consistent with this, we provide evidence that two other IQ-domain containing actomyosin ring proteins, Rng2p (an IQGAP-related protein) and Myo51p (a type V myosin heavy chain), physically interact with Cdc4p. We concluded that Cdc4p, a novel myosin light chain, interacts with multiple actomyosin ring components to effect cytokinesis.

[Synthesis and structural study of substituted arylideneimidazolidines and arylidenebenzothiazines]. / Synthesis and structural study of substituted arylideneimidazolidines and arylidenebenzothiazines.

Synthesis and physico-chemical properties of six 5-arylidène-3-benzyl-1-methyl-2-thioxoimidazolidin-4-ones and three 2-arylidene-6-nitro-2H-1,4-benzothiazin-3(4H)-ones have been described. These new compounds were synthetised by Knoevenagel condensation reaction from aromatic aldehydes. The N-alkylation reaction of arylidenebenzothiazines by methyl iodide give the N-methylarylidenebenzothiazines.

Divalent metal binding properties of the methionyl aminopeptidase from Escherichia coli.

The metal-binding properties of the methionyl aminopeptidase from Escherichia coli (MetAP) were investigated. Measurements of catalytic activity as a function of added Co(II) and Fe(II) revealed that maximal enzymatic activity is observed after the addition of only 1 equiv of divalent metal ion. Based on these studies, metal binding constants for the first metal binding event were found to be 0.3 +/- 0.2 microM and 0.2 +/- 0.2 microM for Co(II)- and Fe(II)-substituted MetAP, respectively. Binding of excess metal ions (>50 equiv) resulted in the loss of approximately 50% of the catalytic activity. Electronic absorption spectral titration of a 1 mM sample of MetAP with Co(II) provided a binding constant of 2.5 +/- 0.5 mM for the second metal binding site. Furthermore, the electronic absorption spectra of Co(II)-loaded MetAP indicated that both metal ions reside in a pentacoordinate geometry. Consistent with the absorption data, electron paramagnetic resonance (EPR) spectra of [CoCo(MetAP)] also indicated that the Co(II) geometries are not highly constrained, suggesting that each Co(II) ion in MetAP resides in a pentacoordinate geometry. EPR studies on [CoCo(MetAP)] also revealed that at pH 7.5 there is no significant spin-coupling between the two Co(II) ions, though a small proportion ( approximately 5%) of the sample exhibited detectable spin-spin interactions at pH values > 9.6. EPR studies on [Fe(III)_(MetAP)] and [Fe(III)Fe(III)(MetAP)] also suggested no spin-coupling between the two metal ions. (1)H nuclear magnetic resonance (NMR) spectra of [Co(II)_(MetAP)] in both H(2)O and D(2)O buffer indicated that the first metal binding site contains the only active-site histidine residue, His171. Mechanistic implications of the observed binding properties of divalent metal ions to the MetAP from E. coli are discussed.

Dermatophytosis caused by Trichophyton raubitschekii. Report of the first case in São Paulo, Brazil.

The authors report the first case of dermatophytosis caused by Trichophyton raubitschekii in a patient from the State of São Paulo with Tinea corporis lesions localized on the buttocks. Culture on Sabouraud-agar with cycloheximide permitted the isolation and identification of the fungus, and the diagnosis was confirmed by Dr. Lynne Sigler, University of Alberta, Canada. Systemic treatment with fluconazole, 150 mg/week for 4 weeks, in combination with topical treatment with isoconazole initially yielded favorable results, with recurrence of the lesions after the medication was discontinued. This is the fifth case of this dermatophytosis published in the Brazilian medical literature.

The methionyl aminopeptidase from Escherichia coli can function as an iron(II) enzyme.

The identity of the physiologically relevant metal ions for the methionyl aminopeptidase (MetAP) from Escherichia coli was investigated and is suggested to be Fe(II). The metal content of whole cells in the absence and presence of expression of the type I MetAP from E. coli was determined by inductively coupled plasma (ICP) emission analysis. The observed change in whole cell concentrations of cobalt, cadmium, copper, nickel, strontium, titanium, and vanadium upon expression of MetAP was negligible. On the other hand, significant increases in the cellular metal ion concentrations of chromium, zinc, manganese, and iron were observed with the increase in iron concentration being 4.4 and 6.2 times greater than that of manganese and zinc, respectively. Activity assays of freshly lysed BL21(DE3) cells containing the pMetAAP plasmid revealed detectable levels (>2 units/mg) of MetAP activity. Control experiments with BL21(DE3) without the MetAP plasmid showed no detectable enzymatic activity. Since MetAP is active upon expression, these data strongly suggest that cobalt is not the in vivo metal ion for the MetAP from E. coli. The MetAP from E. coli as purified was found to be catalytically inactive (

Mechanistic studies on the aminopeptidase from Aeromonas proteolytica: a two-metal ion mechanism for peptide hydrolysis.

The aminopeptidase from Aeromonas proteolytica (AAP) is uncompetitively inhibited by fluoride ion at pH 8.0 with an inhibition constant (Ki) of 30 mM. Thus, fluoride inactivates AAP only after substrate binding, and only a single fluoride ion binds to AAP. On the other hand, chloride ion does not inhibit AAP up to concentrations of 2 M at pH 8.0. The pH dependence of fluoride inhibition of AAP was measured over the pH range 6.0-9.5. Between pH values of 6.0 and 9.0, fluoride ion acts as a pure uncompetitive inhibitor of AAP, and the Ki increases from 1.2 to 370 mM. From a plot of pKi vs pH, a pKa value of 7.0 +/- 0.3 was extracted which corresponds to a single deprotonation process. At pH values higher than 9.0, the fluoride inhibition pattern changes to competitive. This change in inhibition pattern was attributed to a change in ionic strength or perhaps pH of the solution since fluoride ion was also found to become a competitive inhibitor of AAP at pH 8.0 in the presence of 2 M NaCl. These data, taken together with previous kinetic studies of mono- and dinuclear hydrolases with fluoride ion, suggest that a Zn(II)-bound water/hydroxide exists at the dimetal active site of AAP with a pKa of 7.0 and that this water/hydroxide acts as the active site nucleophile. The hydrolysis of L-leucine-p-nitroanilide was measured spectrophotometrically in triplicate between 25 and 85 degrees C at eight substrate concentrations ranging from 5 to 800 microM. From these data, Km values were derived at each temperature studied and were found to increase exponentially with increasing temperature. Moreover, the calculated Vmax values were also found to increase over this temperature range, mimicking the Km values. An Arrhenius plot was constructed from k(cat) values and was found to be linear over the temperature range 25-85 degrees C, indicating that the rate-limiting step in AAP peptide hydrolysis is product formation and does not change as a function of temperature. From the slope of the line, the activation energy (Ea) was calculated to be 36.5 kJ/mol. The enthalpy and entropy of activation at 25 degrees C calculated over the temperature range 298-358 K were found to be 34.0 kJ/mol and -94.2 J/(mol x K), respectively. The free energy of activation at 25 degrees C was found to be 62.1 kJ/mol. Combination of the available X-ray crystallographic data with the present kinetic and thermodynamic results, as well as the previously reported kinetic and spectroscopic data, has allowed a detailed catalytic mechanism for AAP to be proposed.

Calor versus dissociaçåo acida de imunocomplexos para a detecçåo da antigemia P24 / Heat versus acid dissociation of immune complexes for the detection of p24 antigemia

INTRODUÇÄO - A antigemia P24 é amplamente utilizada como marcador substitutivo de progresso da infecçåo pelo HIV e da eficácia da terapêutica. A dissociaçåo ácida de imunocomplexos, por aumentar a sensibilidade dos testes para a detecçåo do P24, tornou-se a técnica padråo. OBJETIVO - Comparar a dissociaçåo pelo calor (HD) com a dissociaçåo ácida (AD) dos imunocomplexos nos testes de captura para antígeno P24. MÉTODOS - Soros de pacientes infectados pelo HIV previamente conhecidos por serem P24 negativos por AD, foram ensaiadas paralelamente depois da AD e HD, usando um tampåo de Glicina seguido por neutralizaçåo com um tampåo tris. A HD foi realizada diluindo-se o soro 1:3 em água destilada seguido pela imersåo dos tubos em banho de água fervente por 5 minutos. Os títulos de P24 foram quantificados pela comparaçåo da densidade óptica do soro com uma curva padråo de espécimes positivos. RESULTADOS - Das 39 amostras testadas, todas permaneceram negativas por AD e 3(7,7 por cento; 95 CI= 0-16 por cento) tornaram-se positivas depois do HD. CONCLUSÄO - A HD de imunocomplexos é de execuçåo mais barata, simples e rápida do que a AD, aumentando ainda a sensibilidade das técnicas de detecçåo do antígeno P24

Mycoflora and aflatoxigeni species of aspergillus spp isolated from stored maize

Pesquisou-se a microbiota fúngica e esécies aflatoxigênicas do gênero Asperegillus em 90 amostras milho provenientes de silos de armazenamento. Foram isolados 8 gêneros fúgicos: aspergillus, penicillium, fusarium, rhizopus, acrenonium, cladosporium, neurospora e paecilomyces. Do gênero aspergillus, a espécie aspergillus flavus foi a mais frequente, isolada de amostras com umidade entre 13 e 18 por cento. Todos os isolados desta espécie foram toxigênicos e produziram aflatoxinas B1, B2, G1 e G2. As outras espécies do gênero foram: A. chriselius, A. versicolor, A. sydowi, A. fumigatus e A. niger, e ocorrem somente nas amostras com teores de umidade entre 12 e 13 por cento