RESUMO
Pharyngitis and tonsillitis are the most common acute respiratory infections (ARIs) in children aged ≤5 years. The analysis of published data showed that some probiotics could decrease the frequency and number of days with ARIs. This study evaluated the safety and efficacy of Limosilactobacillus reuteri ATCC PTA 5289 and DSM 17938 to reduce the duration and severity of ARI symptoms. This randomised controlled trial included children aged from 6 months to 5 years, with pharyngitis or tonsillitis, who were randomised to receive a probiotic product containing L. reuteri ATCC PTA 5289 and L. reuteri DSM 17938 or placebo, as drops, ingested orally for 10 days as adjuvants to the use of non-steroidal anti-inflammatory drugs. The main outcomes were the duration and severity of ARI symptoms. The secondary outcomes were changes in salivary immunoglobulin A and inflammatory biomarkers. There was no fever on day 2 and subsequent days in the L. reuteri group (37.3 ±0.5 °C vs 38.6±0.3 °C, P<0.05). Beginning on day 3, the severity of sore throat (5±0.9 vs 8±1.2, P<0.05) was lower in the L. reuteri group. Significant differences in the days with runny nose, nasal congestion, days of non-programmed visits to the medical office or emergency department, levels in tumoral necrosis factor-alpha (TNF-alpha) and related costs of treatment were observed in the L. reuteri group. The frequency of adverse events was similar between the groups. Therefore, L. reuteri ATCC PTA 5289 combined with L. reuteri DSM 17938 is a safe and effective adjunct to reduce the symptoms of pharyngitis or tonsillitis in children.
Assuntos
Limosilactobacillus reuteri/fisiologia , Faringite/tratamento farmacológico , Probióticos/administração & dosagem , Tonsilite/tratamento farmacológico , Pré-Escolar , Feminino , Humanos , Imunoglobulina A/imunologia , Lactente , Masculino , Faringite/imunologia , Saliva/imunologia , Tonsilite/imunologia , Fator de Necrose Tumoral alfa/imunologiaRESUMO
An M13 phage random 12-mers peptide library was used to screen cathepsin L mimotopes of Fasciola hepatica and to evaluate their immunogenicity in sheep. Seven clones showed positive reactivity to a rabbit anti-cathepsin L1/L2 antiserum in ELISA, and their amino acid sequences deduced by DNA sequencing were tentatively mapped on the protein. Twenty sheep were randomly allocated into 4 groups of 5 animals each, for immunization with 1x10(14) phage particles of clones 1, 20, a mixture of 7 clones and PBS, without adjuvant at the beginning, and 4 weeks later. All groups were challenged with 300 metacercariae at week 6 and slaughtered 16 weeks later. The mean worm burdens after challenge were reduced by 47.61% and 33.91% in sheep vaccinated with clones 1 and 20, respectively; no effect was observed in animals inoculated with the clone mixture. Also, a significant reduction in worm size and burden was observed for those sheep immunized with clone 1. Animals receiving clone 20, showed a significant reduction in egg output. Immunization induced a reduction of egg viability ranging from 58.92 to 82.11%. Furthermore, vaccinated animals produced clone-specific antibodies which were boosted after challenge with metacercariae of F. hepatica.
Assuntos
Catepsinas/imunologia , Cisteína Endopeptidases/imunologia , Fasciolíase/veterinária , Biblioteca de Peptídeos , Doenças dos Ovinos/imunologia , Sequência de Aminoácidos , Animais , Catepsina L , Catepsinas/química , Cisteína Endopeptidases/química , Fasciola hepatica , Fasciolíase/prevenção & controle , Fezes/parasitologia , Dados de Sequência Molecular , Contagem de Ovos de Parasitas , Ovinos , Doenças dos Ovinos/parasitologia , Vacinas/imunologiaRESUMO
The amino acid sequences and X-ray structures of homodimeric triosephosphate isomerase from the pathogenic parasites Trypanosoma brucei (TbTIM) and Trypanosoma cruzi (TcTIM) are markedly similar. In the two TIMs, the side chain of the only interface cysteine (Cys14) of one subunit docks into loop 3 of the other subunit. This portion of the interface is also markedly similar in the two enzymes. Nonetheless, Cys14 of TcTIM is nearly 2 orders of magnitude more susceptible to the thiol reagent methylmethane thiosulfonate (MMTS) than Cys14 of TbTIM. The causes of this difference were explored by measuring the second-order rate constant of inactivation by MMTS (k(2)) under various conditions. At pH 7.4, k(2) in TcTIM is 70 times higher than in TbTIM. The difference decreases to 30 when the amino acid sequence of loop 3 and adjoining residues of TbTIM are conferred to TcTIM (triple mutant). The pK(a) values of the thiol group of the interface cysteine of TcTIM and the triple mutant were 0.7 pH unit lower than in TbTIM. Because this difference could account for the different sensitivity of the enzymes to thiol reagents, we determined the k(2) of inactivation at equal levels of ionization of their interface cysteines. Under these conditions, the difference in k(2) between TcTIM and TbTIM became 8-fold, whereas that of the triple mutant to TbTIM was 1.5 times. The substrate analogue phosphoglycolate did not modify the pK(a) of the thiol group of the interface, albeit it diminished the rate of its derivatization by MMTS. In the presence of phosphoglycolate, under conditions in which the interface cysteines of the enzymes had equal levels of protonation, the difference in k(2) of TcTIM and TbTIM became smaller, whereas k(2) of the triple mutant was almost equal to that of TbTIM. Thus, from measurements of the reactivity of the interface cysteine in various conditions, it was possible to obtain information on the factors that control the dynamics of a portion of the dimer interface.
Assuntos
Cisteína/metabolismo , Triose-Fosfato Isomerase/metabolismo , Trypanosoma brucei brucei/enzimologia , Trypanosoma cruzi/enzimologia , Sequência de Aminoácidos , Substituição de Aminoácidos/genética , Animais , Dimerização , Ativação Enzimática/efeitos dos fármacos , Ativação Enzimática/genética , Inibidores Enzimáticos/farmacologia , Ácido Glutâmico/genética , Glutamina/genética , Glicolatos/farmacologia , Concentração de Íons de Hidrogênio , Leishmania mexicana/enzimologia , Metanossulfonato de Metila/farmacologia , Dados de Sequência Molecular , Mutagênese Sítio-Dirigida , Fragmentos de Peptídeos/genética , Fragmentos de Peptídeos/metabolismo , Prolina/genética , Estrutura Terciária de Proteína , Triose-Fosfato Isomerase/antagonistas & inibidores , Triose-Fosfato Isomerase/genética , Trypanosoma brucei brucei/genética , Trypanosoma cruzi/genéticaRESUMO
BACKGROUND: The presence of RNA in the cell nucleus is well known. However, a high resolution in situ hybridization evidence for the presence of RNA in some nuclear particles is still lacking. The aim of this work is to localize RNA in subnuclear particles using a novel ultrastructural in situ hybridization procedure. In this study, biotinylated genomic mouse DNA as a probe to localize total RNA in the nuclei of mouse hepatocytes was used. METHODS: The procedure is based on paraformaldehyde fixation and embedding in lowicryl resin. Thin sections are mounted in formvar-coated gold grids. Hybridization is performed on non-denatured thin sections. DNA-RNA hybrids are detected with streptavidin-10 nm gold particles complex. By controlling the time of nick-translation during incorporation of biotin into the probe, labeling in the fibrillar portions of the nucleoplasm is obtained. More digested probes generate more labeling in the granular components. Nucleoli were similarly labeled. RESULTS: As expected, no label was observed in the compact chromatin clumps. These results indicate that granular components as perichromatin granules in the nucleus contain more processed RNA than fibrillar portions. As a comparison, viral DNA sequences on denatured RNase-treated thin sections of adenovirus-2 (Ad-2)-infected human cells were detected. As previously reported, at late stages DNA was observed in the viral particles and surrounding nucleoplasm, where Ad-2 DNA is synthesized. CONCLUSIONS: The present procedure allows the study of intranuclear RNA distribution and will be useful for the analysis of RNA processing in several types of cells.