RESUMO
The activities of rifampin, nitazoxanide, PA-824, and sutezolid were tested against dormant Mycobacterium tuberculosis under conditions mimicking caseous granulomas (hypoxia at pH 7.3) in comparison with those of the combination rifampin-isoniazid-pyrazinamide-ethambutol (R-I-Z-E), which is used for human therapy. Mycobacterial viability was monitored by CFU and regrowth in MGIT 960. As shown by lack of regrowth in MGIT, rifampin-nitazoxanide-containing combinations, but not R-I-Z-E, killed dormant cells in 28 to 35 days. These observations might be important in designing new tuberculosis therapies.
Assuntos
Antituberculosos/farmacologia , Isoniazida/farmacologia , Mycobacterium tuberculosis/efeitos dos fármacos , Pirazinamida/farmacologia , Rifampina/farmacologia , Combinação de Medicamentos , Quimioterapia Combinada , Etambutol/farmacologia , Humanos , Concentração de Íons de Hidrogênio , Hipóxia , Testes de Sensibilidade Microbiana , Nitroimidazóis/farmacologia , Oxazolidinonas/farmacologia , Tuberculose/microbiologiaRESUMO
The activities of rifampin, rifapentine, bedaquiline, PA-824, clofazimine, nitazoxanide, isoniazid, amikacin, moxifloxacin, niclosamide, thioridazine, and pyrazinamide were tested against nonreplicating (dormant) Mycobacterium tuberculosis H37Rv under conditions of hypoxia at pHs 5.8 and 7.3, mimicking environments of cellular granulomas and caseous granulomas, respectively. At pH 5.8, several drugs killed dormant bacilli, with the best being rifampin and rifapentine. At pH 7.3, only rifampin and rifapentine efficiently killed dormant bacilli, while all other drugs showed little activity.
Assuntos
Antituberculosos/farmacologia , Mycobacterium tuberculosis/efeitos dos fármacos , Rifampina/análogos & derivados , Rifampina/farmacologia , Anaerobiose , Concentração de Íons de Hidrogênio , Ácidos Isonicotínicos/farmacologia , Testes de Sensibilidade Microbiana , Mycobacterium tuberculosis/crescimento & desenvolvimento , Nitroimidazóis/farmacologia , Fenazinas/farmacologia , Quinolinas/farmacologia , Tiazóis/farmacologiaRESUMO
BACKGROUND: The therapeutic HIV-1 Tat protein vaccine is in advanced clinical development. Tuberculosis, the main AIDS co-infection, is highly endemic in areas where AIDS prevention through vaccination is needed. However, safety and immunogenicity of Tat vaccination in the course of Mycobacterium tuberculosis (Mtb) infection is still unknown and it prevents the possibility to administer the vaccine to Mtb-infected individuals. We addressed the interplay and effects of Tat vaccination on Mtb infection in immunocompetent mice. METHODS: C57BL/6 mice were vaccinated or not with Bacillus Calmette-Guerin (BCG), the current tuberculosis vaccine, and after 5 weeks were infected with Mtb by intravenous route. The Tat protein was injected intradermally at 1, 2 and 4 weeks after Mtb challenge. Eight weeks after Mtb infection, all mice were sacrificed, and both the degree of pathology and immune responses to Mtb and Tat were evaluated. As additional control, some mice were either vaccinated or not with BCG, were not challenged with Mtb, but received the Tat protein. Statistical significances were evaluated by one-way or two-way ANOVA and Tukey's multiple comparisons post-test. RESULTS: In the lungs of Mtb-infected mice, Tat-vaccine did not favour Mtb replication and indeed reduced both area of cellular infiltration and protein levels of Interferon-γ, Chemokine (C-C motif) ligand-4 and Interleukin-1ß, pathological events triggered by Mtb-infection. Moreover, the protection against Mtb infection conferred by BCG remained good after Tat protein treatment. In spleen cells of Mtb-infected mice, Tat vaccination enhanced Mtb-specific Interferon-γ and Interleukin-17 responses, which may have a protective role. Of note, Mtb infection reduced, but did not suppress, the development of anti-Tat antibodies, required for Tat vaccine efficacy and the titer of anti-Tat IgG was potentiated by BCG vaccination in Mtb-free mice. In general, Tat treatment was well tolerated in both Mtb-infected and Mtb-free mice. CONCLUSIONS: Tat protein vaccine, administered in Mtb-infected mice with a protocol resembling that used in the clinical trials, was safe, immunogenic, limited the lung Mtb-associated immunopathology and did not abrogate the protective efficacy of BCG. These data provide preliminary evidence for a safe use of Tat vaccine in people vaccinated with BCG and/or suffering from tuberculosis.
Assuntos
HIV-1/metabolismo , Mycobacterium tuberculosis/patogenicidade , Produtos do Gene tat do Vírus da Imunodeficiência Humana/imunologia , Animais , Vacina BCG/imunologia , Carga Bacteriana , Células Cultivadas , Quimiocina CCL4/metabolismo , Ensaio de Imunoadsorção Enzimática , Feminino , Anticorpos Anti-HIV/sangue , HIV-1/imunologia , Interferon gama/metabolismo , Interleucina-17/metabolismo , Interleucina-1beta/metabolismo , Pulmão/microbiologia , Pulmão/patologia , Camundongos , Camundongos Endogâmicos C57BL , Proteínas Recombinantes/biossíntese , Proteínas Recombinantes/imunologia , Proteínas Recombinantes/isolamento & purificação , Baço/citologia , Baço/metabolismo , Baço/microbiologia , Vacinação , Produtos do Gene tat do Vírus da Imunodeficiência Humana/genética , Produtos do Gene tat do Vírus da Imunodeficiência Humana/metabolismoRESUMO
The antituberculosis (anti-TB) drug rifampin (RIF) binds to the beta subunit of the RNA polymerase (RpoB) of Mycobacterium tuberculosis, but the bactericidal responses triggered after target interaction are not known. To evaluate whether RIF induced an oxidative burst, lysates of RIF-treated M. tuberculosis were tested for determination of reactive oxygen species (ROS) by the electron paramagnetic resonance (EPR) technique using 1-hydroxy-3-carboxy-pyrrolidine (CPH) and 5,5-dimethyl-1-pyrrolidine-N-oxide (DMPO) as spin traps. M. tuberculosis killing by RIF stimulated an increase in the rate of formation of the CPH radical (CP·). Lysate pretreatment with the O2·(-) and ·OH scavengers superoxide dismutase (SOD) and thiourea (THIO), respectively, or with the metal chelator diethylene triamine pentaacetic acid (DTPA) inhibited CP· formation, arguing in favor of a metal-catalyzed ROS response. Formation of CP· did not increase following treatment of RIF-resistant strains with RIF, indicating that the ROS were induced after RpoB binding. To identify the ROS formed, lysates of RIF-treated bacilli were incubated with DMPO, a spin trap specific for ·OH and O2·(-), with or without pretreatment with SOD, catalase, THIO, or DTPA. Superoxide dismutase, catalase, and THIO decreased formation of the DMPO-OH adduct, and SOD plus DTPA completely suppressed it, suggesting that RIF activated metal-dependent O2·(-)-mediated mechanisms producing ·OH inside tubercle bacilli. The finding that the metal chelator DTPA reduced the bactericidal activity of RIF supported the possibility that ·OH was generated through these mechanisms and that it participated at least in part in M. tuberculosis killing by the drug.
Assuntos
Antituberculosos/farmacologia , Proteínas de Bactérias/metabolismo , Radical Hidroxila/metabolismo , Mycobacterium tuberculosis/efeitos dos fármacos , Rifampina/farmacologia , Superóxidos/metabolismo , Antituberculosos/metabolismo , Proteínas de Bactérias/química , Catalase/química , Óxidos N-Cíclicos , RNA Polimerases Dirigidas por DNA , Espectroscopia de Ressonância de Spin Eletrônica , Radical Hidroxila/química , Mycobacterium tuberculosis/química , Mycobacterium tuberculosis/metabolismo , Estresse Oxidativo , Ácido Pentético/química , Ligação Proteica , Rifampina/metabolismo , Marcadores de Spin , Superóxido Dismutase/química , Superóxidos/química , Tioureia/químicaRESUMO
The immunological mechanisms that modulate protection during Mycobacterium tuberculosis (Mtb) infection or vaccination are not fully understood. Secretion of IFN-γ and, to a lesser extent, of IL-17 by CD4(+) T cells plays a major role both in protection and immunopathology. Few Mtb Ags interacting with DCs affect priming, activation, and regulation of Ag-unrelated CD4(+) T-cell responses. Here we demonstrate that PstS1, a 38 kDa-lipoprotein of Mtb, promotes Ag-independent activation of memory T lymphocytes specific for Ag85B or Ag85A, two immunodominant protective Ags of Mtb. PstS1 expands CD4(+) and CD8(+) memory T cells, amplifies secretion of IFN-γ and IL-22 and induces IL-17 production by effector memory cells in an Ag-unrelated manner in vitro and in vivo. These effects were mediated through the stimulation of DCs, particularly of the CD8α(-) subtype, which respond to PstS1 by undergoing phenotypic maturation and by secreting IL-6, IL-1ß and, to a lower extent, IL-23. IL-6 secretion by PstS1-stimulated DCs was required for IFN-γ, and to a lesser extent for IL-22 responses by Ag85B-specific memory T cells. These results may open new perspectives for immunotherapeutic strategies to control Th1/Th17 immune responses in Mtb infections and in vaccinations against tuberculosis.
Assuntos
Transportadores de Cassetes de Ligação de ATP/metabolismo , Proteínas de Bactérias/metabolismo , Linfócitos T CD4-Positivos/imunologia , Interferon gama/metabolismo , Interleucina-17/metabolismo , Interleucinas/metabolismo , Aciltransferases/imunologia , Animais , Antígenos de Bactérias/imunologia , Proteínas de Bactérias/imunologia , Linfócitos T CD4-Positivos/metabolismo , Antígenos CD8/metabolismo , Linfócitos T CD8-Positivos/imunologia , Proliferação de Células , Células Dendríticas/imunologia , Memória Imunológica , Interferon gama/biossíntese , Interleucina-17/biossíntese , Interleucina-1beta/metabolismo , Interleucina-23/metabolismo , Interleucina-6/metabolismo , Interleucinas/biossíntese , Ativação Linfocitária/imunologia , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Camundongos Knockout , Mycobacterium tuberculosis/imunologia , Receptor 2 Toll-Like/deficiência , Receptor 2 Toll-Like/genética , Tuberculose/imunologia , Interleucina 22RESUMO
Tuberculosis is one of the deadliest infectious diseases worldwide. Mycobacterium tuberculosis has developed strategies not only to evade host immunity but also to manipulate it for its survival. We investigated whether Mycobacterium tuberculosis exploited the immunogenicity of Ag85B, one of its major secretory proteins, to redirect host antituberculosis immunity to its advantage. We found that administration of Ag85B protein to mice vaccinated with Bacillus Calmette-Guérin impaired the protection elicited by vaccination, causing a more severe infection when mice were challenged with Mycobacterium tuberculosis. Ag85B administration reduced Bacillus Calmette-Guérin-induced CD4 T-cell activation and IFN-γ, CCL-4, and IL-22 production in response to Mycobacterium tuberculosis-infected cells. On the other hand, it promoted robust Ag85B-responsive IFN-γ-producing CD4 T cells, expansion of a subset of IFN-γ/IL-10-producing CD4+FOXP3+Treg cells, differential activation of IL-17/IL-22 responses, and activation of regulatory and exhaustion pathways, including programmed death ligand 1 expression on macrophages. All this resulted in impaired intracellular Mycobacterium tuberculosis growth control by systemic immunity, both before and after the Mycobacterium tuberculosis challenge. Interestingly, Mycobacterium tuberculosis infection itself generated Ag85B-reactive inflammatory immune cells incapable of clearing Mycobacterium tuberculosis in both unvaccinated and Bacillus Calmette-Guérin-vaccinated mice. Our data suggest that Mycobacterium tuberculosis can exploit the strong immunogenicity of Ag85B to promote its own survival and spread. Since Ag85B is normally secreted by replicating bacteria and is commonly found in the lungs of the Mycobacterium tuberculosis-infected host, our findings may advance the understanding on the mechanisms of Mycobacterium tuberculosis pathogenesis and immune evasion.
Assuntos
Aciltransferases , Antígenos de Bactérias , Vacina BCG , Proteínas de Bactérias , Camundongos Endogâmicos C57BL , Mycobacterium tuberculosis , Tuberculose , Animais , Antígenos de Bactérias/imunologia , Mycobacterium tuberculosis/imunologia , Aciltransferases/imunologia , Vacina BCG/imunologia , Camundongos , Tuberculose/imunologia , Tuberculose/microbiologia , Proteínas de Bactérias/imunologia , Feminino , Linfócitos T CD4-Positivos/imunologia , Citocinas/metabolismo , Viabilidade MicrobianaRESUMO
Mycobacterium tuberculosis is exposed to hypoxia and acidity within granulomatous lesions. In this study, an acidic culture model of M. tuberculosis was used to test drug activity against aerobic 5-day-old (A5) and hypoxic 5-, 12-, and 19-day-old (H5, H12, and H19, respectively) bacilli after 7, 14, and 21 days of exposure. In A cultures, CFU and pH rapidly increased, while in H cultures growth stopped and pH increased slightly. Ten drugs were tested: rifampin (R), isoniazid (I), pyrazinamide (Z), ethambutol (E), moxifloxacin (MX), amikacin (AK), metronidazole (MZ), nitazoxanide (NZ), niclosamide (NC), and PA-824 (PA). Rifampin was the most active against A5, H5, H12, and H19 bacilli. Moxifloxacin and AK efficiently killed A5 and H5 cells, I was active mostly against A5 cells, Z was most active against H12 and H19 cells, and E showed low activity. Among nitrocompounds, NZ, NC, and PA were effective against A5, H5, H12, and H19 cells, while MZ was active against H12 and H19 cells. To kill all A and H cells, A5- and H5-active agents R, MX, and AK were used in combination with MZ, NZ, NC, or PA, in comparison with R-I-Z-E, currently used for human therapy. Mycobacterial viability was determined by CFU and a sensitive test in broth (day to positivity, MGIT 960 system). As shown by lack of regrowth in MGIT, the most potent combination was R-MX-AK-PA, which killed all A5, H5, H12, and H19 cells in 14 days. These observations demonstrate the sterilizing effect of drug combinations against cells of different M. tuberculosis stages grown in aerobic and hypoxic acidic conditions.
Assuntos
Aerobiose/efeitos dos fármacos , Anaerobiose/efeitos dos fármacos , Antituberculosos/farmacologia , Combinação de Medicamentos , Mycobacterium tuberculosis/efeitos dos fármacos , Aerobiose/fisiologia , Anaerobiose/fisiologia , Contagem de Colônia Microbiana , Meios de Cultura , Concentração de Íons de Hidrogênio , Testes de Sensibilidade Microbiana , Viabilidade Microbiana/efeitos dos fármacos , Mycobacterium tuberculosis/crescimento & desenvolvimentoRESUMO
Caveolae have been indicated as a center of cytoskeleton regulation for Src kinase/Rho GTPase signaling. In addition, Src recruitment on intact cortical actin cytoskeleton appears to be required for bFGF/FGFR signal activation. Recently, we established a relationship between caveolin-1 (Cav-1) expression and cell migration in human malignant melanoma, constitutively activated by a bFGF autoregulatory loop. This work intends to investigate whether caveolae's asset, through bFGF/FGFR/c-Src/Rho signaling, could be related to melanoma cell anchorage. Accordingly, we revealed the existence of a FGFR/Src kinase pathway in Cav-1 enriched detergent-resistant membranes (DRMs) of Me665/1 metastatic melanoma cells, as confirmed by FGFR silencing. Moreover, we determined the expression and phosphorylation levels of Cav-1/Src/Erk signal pathway as a function of FGFR activation and cell density. A sucrose density gradient ultracentrifugation was employed to monitor Cav-1 membrane association and buoyancy in Me665/1 cells treated for actin fragmentation or for altered phosphorylation signals. As a result, melanoma cells show remarkable resistance to Cav-1 disassembly, together with persisting cell signal activity, being Src and Cav-1 crucial modulators of Rho GTPases. In conclusion, our study primarily highlights, in a metastatic melanoma cell line expressing caveolin, the circumstances whereby caveola structural and functional endurance enables the FGFR/Src/Rho GTPases pathway to keep on cell progression.
Assuntos
Caveolina 1/metabolismo , Melanoma/metabolismo , Receptores de Fatores de Crescimento de Fibroblastos/metabolismo , Proteínas rho de Ligação ao GTP/metabolismo , Quinases da Família src/metabolismo , Actinas/metabolismo , Caveolina 1/genética , Contagem de Células , Linhagem Celular Tumoral , Membrana Celular/genética , Membrana Celular/metabolismo , Movimento Celular/fisiologia , Citoesqueleto/genética , Citoesqueleto/metabolismo , Humanos , Sistema de Sinalização das MAP Quinases , Melanoma/genética , Melanoma/patologia , Fosforilação , Receptores de Fatores de Crescimento de Fibroblastos/genética , Transdução de Sinais , Proteínas rho de Ligação ao GTP/genética , Quinases da Família src/genéticaAssuntos
Migrantes , Tuberculose/epidemiologia , África , Antituberculosos/uso terapêutico , Europa (Continente) , Tuberculose Extensivamente Resistente a Medicamentos/epidemiologia , Geografia , Humanos , Itália/epidemiologia , Testes de Sensibilidade Microbiana , Mycobacterium tuberculosis , Federação Russa , Tuberculose Resistente a Múltiplos Medicamentos/epidemiologia , Organização Mundial da SaúdeRESUMO
The M. tuberculosis phosphate-binding transporter lipoproteins PstS1 and PstS3 were good immunogens inducing CD8(+) T-cell activation and both Th1 and Th17 immunity in mice. However, this antigen-specific immunity, even when amplified by administration of the protein with the adjuvant LTK63 or by the DNA priming/protein boosting regimen, was not able to contain M. tuberculosis replication in the lungs of infected mice. The lack of protection might be ascribed with the scarce/absent capacity of PstS1/PstS3 antigens to modulate the IFN-γ response elicited by M. tuberculosis infection during which, however, PstS1-specific IL-17 secreting cells were generated in both unvaccinated and BCG-vaccinated mice. In spite of a lack of protection by PstS1/PstS3 immunizations, our results do show that PstS1 is able to induce IL-17 response upon M. tuberculosis infection which is of interest in the study of anti-M. tuberculosis immunity and as potential immunomodulator in combined vaccines.
Assuntos
Proteínas de Bactérias/imunologia , Mycobacterium tuberculosis/imunologia , Células Th1/imunologia , Células Th17/imunologia , Tuberculose/imunologia , Tuberculose/prevenção & controle , Aciltransferases/imunologia , Animais , Antígenos de Bactérias/imunologia , Vacina BCG/imunologia , Carga Bacteriana/imunologia , Linfócitos T CD8-Positivos/imunologia , Linfócitos T CD8-Positivos/metabolismo , Epitopos , Feminino , Imunização , Memória Imunológica/imunologia , Interferon gama/biossíntese , Interleucina-17/biossíntese , Interleucina-17/imunologia , Camundongos , Camundongos Endogâmicos C57BL , Vacinas contra a Tuberculose/imunologiaRESUMO
Previous studies on Escherichia coli demonstrated that sub-minimum inhibitory concentration (MIC) of fluoroquinolones induced the SOS response, increasing drug tolerance. We characterized the transcriptional response to moxifloxacin in Mycobacterium tuberculosis. Reference strain H37Rv was treated with moxifloxacin and gene expression studied by qRT-PCR. Five SOS regulon genes, recA, lexA, dnaE2, Rv3074 and Rv3776, were induced in a dose- and time-dependent manner. A range of moxifloxacin concentrations induced recA, with a peak observed at 2 × MIC (0.25 µg/mL) after 16 h. Another seven SOS responses and three DNA repair genes were significantly induced by moxifloxacin. Induction of recA by moxifloxacin was higher in log-phase than in early- and stationary-phase cells, and absent in dormant bacilli. Furthermore, in an H37Rv fluoroquinolone-resistant mutant carrying the D94G mutation in the gyrA gene, the SOS response was induced at drug concentrations higher than the mutant MIC value. The 2 × MIC of moxifloxacin determined no significant changes in gene expression in a panel of 32 genes, except for up-regulation of the relK toxin and of Rv3290c and Rv2517c, two persistence-related genes. Overall, our data show that activation of the SOS response by moxifloxacin, a likely link to increased mutation rate and persister formation, is time, dose, physiological state and, possibly, MIC dependent.
RESUMO
There is a strong relationship between metabolic state and susceptibility to Mycobacterium tuberculosis (MTB) infection, with energy metabolism setting the basis for an exaggerated immuno-inflammatory response, which concurs with MTB pathogenesis. Herein, we show that controlled caloric restriction (CR), not leading to malnutrition, protects susceptible DBA/2 mice against pulmonary MTB infection by reducing bacterial load, lung immunopathology, and generation of foam cells, an MTB reservoir in lung granulomas. Mechanistically, CR induced a metabolic shift toward glycolysis, and decreased both fatty acid oxidation and mTOR activity associated with induction of autophagy in immune cells. An integrated multi-omics approach revealed a specific CR-induced metabolomic, transcriptomic, and proteomic signature leading to reduced lung damage and protective remodeling of lung interstitial tightness able to limit MTB spreading. Our data propose CR as a feasible immunometabolic manipulation to control MTB infection, and this approach offers an unexpected strategy to boost immunity against MTB.
Assuntos
Tuberculose/prevenção & controle , Animais , Restrição Calórica , Feminino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Endogâmicos DBA , Mycobacterium tuberculosis/imunologia , Mycobacterium tuberculosis/metabolismo , Tuberculose/imunologia , Tuberculose/metabolismoRESUMO
Aerobic (5-day-old cultures) and nonreplicating (dormant) Mycobacterium tuberculosis (5-, 12-, and 19-day-old cultures) bacteria were treated with rifampin (R), moxifloxacin (MX), metronidazole (MZ), amikacin (AK), or capreomycin (CP) for 7, 14, and 21 days. R-MX-MZ-AK and R-MX-MZ-CP killed both aerobic and dormant bacilli in 21 days, as shown by lack of regrowth in solid and liquid media. R-MX-MZ-AK and R-MX-MZ-CP also caused a strong decrease of nonreplicating bacilli in 7 days in a cell-based dormancy model.
Assuntos
Antituberculosos/administração & dosagem , Tuberculose Latente/tratamento farmacológico , Tuberculose Latente/microbiologia , Mycobacterium tuberculosis/efeitos dos fármacos , Mycobacterium tuberculosis/crescimento & desenvolvimento , Células 3T3 , Adipócitos/efeitos dos fármacos , Adipócitos/microbiologia , Amicacina/administração & dosagem , Animais , Compostos Aza/administração & dosagem , Capreomicina/administração & dosagem , Contagem de Colônia Microbiana , Combinação de Medicamentos , Farmacorresistência Bacteriana , Fluoroquinolonas , Humanos , Técnicas In Vitro , Metronidazol/administração & dosagem , Camundongos , Testes de Sensibilidade Microbiana , Modelos Biológicos , Moxifloxacina , Quinolinas/administração & dosagem , Rifampina/administração & dosagemRESUMO
Most microbes invading through mucosal surfaces cause disease and therefore strategies to induce mucosal immune responses are strongly needed. Vitamin A metabolites, such as retinoic acid (RA), play crucial roles in programming T and B cells to home to mucosal compartments, therefore we evaluated the capacity of RA to elicit mucosal immune responses against tuberculosis (TB) after parenteral vaccination. We found that mice immunized through subcutaneous injections with the TB subunit vaccine (CAF01+H56) in presence of RA show enhanced mucosal H56-specific IgA responses and enhanced Ag-specific CD4+ T lymphocytes homing to the lung as compared with control mice. Immunization with CAF01+H56 in presence of RA resulted in lower bacterial loads in the lungs of mice 14 days after challenge with virulent Mycobacterium tuberculosis (Mtb) as compared to mice immunized in the absence of RA or vaccinated with BCG. Higher amounts of IFNγ and IL-17 pro-inflammatory cytokines were found in lung homogenates of mice immunized with CAF01+H56 and RA 24 h after Mtb infection. However, 6 weeks after infection the protection was comparable in vaccinated mice with or without RA even though treatment with RA during immunization is able to better contain the inflammatory response by the host. Furthermore, at later stage of the infection a higher percentage of Mtb specific CD4+PD1+ T lymphocytes were found in the lungs of mice immunized with CAF01+H56 and RA. These data show that an enhanced mucosal immune response is generated during parenteral vaccination in presence of RA. Furthermore, RA treatment contained the bacterial growth at an early stage of the infection and limited the inflammatory response in the lung at later time points.
Assuntos
Imunidade nas Mucosas , Tretinoína/administração & dosagem , Vacinas contra a Tuberculose/administração & dosagem , Tuberculose Pulmonar/prevenção & controle , Vacinas de Subunidades Antigênicas/administração & dosagem , Alérgenos/imunologia , Animais , Anticorpos Antibacterianos/imunologia , Células Produtoras de Anticorpos/imunologia , Linfócitos T CD4-Positivos/imunologia , Citocinas/imunologia , Feminino , Imunoglobulina A/imunologia , Imunoglobulina G/imunologia , Pulmão/imunologia , Linfócitos/imunologia , Camundongos , Mycobacterium tuberculosis/crescimento & desenvolvimento , Mycobacterium tuberculosis/imunologia , Ovalbumina/imunologia , Tuberculose Pulmonar/imunologia , VacinaçãoRESUMO
Probiotics, defined as "living microorganisms that, whether ingested in useful amount, may have beneficial effects on human body", are widely used in various products for human use, such as dietary supplements, medical devices and pharmaceutical products. The European Directive on medical devices (MDs) (DDM 93/42), also includes those MDs containing live microorganisms, particularly probiotics, that may have various destinations of use, including that of assisting the therapy of several human pathologies. In this brief note we analyzed the use of probiotics in MDs and how probiotics administration could represent one of the new frontiers of scientific research on the prevention and treatment of various diseases. We'll analyze the literature on probiotics based MDs, to review their major targets in the therapy of some of the most common human pathologies: bacterial vaginosis and vaginitis, atopic dermatitis, infantile colic, obesity, type 2 diabetes, and pharyngotonsillitis.
Assuntos
Probióticos/administração & dosagem , Vaginose Bacteriana/terapia , Administração Oral , Adolescente , Adulto , Criança , Pré-Escolar , Cólica/terapia , Dermatite Atópica , Diabetes Mellitus Tipo 2/terapia , Método Duplo-Cego , Equipamentos e Provisões/microbiologia , Feminino , Humanos , Lactente , Recém-Nascido , Lactobacillus , Masculino , Obesidade/prevenção & controle , Obesidade/terapia , Faringite/terapia , Prebióticos/administração & dosagem , Prebióticos/efeitos adversos , Probióticos/efeitos adversos , Ensaios Clínicos Controlados Aleatórios como Assunto , Supositórios , Simbióticos/administração & dosagem , Simbióticos/efeitos adversos , Tonsilite/terapia , Vaginite/microbiologia , Vaginite/terapia , Vaginose Bacteriana/microbiologiaRESUMO
Cholesterol-enriched functional portions of plasma membranes, such as caveolae and rafts, were isolated from lungs of wild-type (WT) and caveolin-1 knockout (Cav-1 KO) mice within detergent resistant membranes (DRMs). To gain insight into their molecular composition we performed proteomic and lipid analysis on WT and Cav-1 KO-DRMs that showed predicted variations of proteomic profiles and negligible differences in lipid composition, while Langmuir monolayer technique and small and wide-angle X-ray scattering (SAXS-WAXS) were here originally introduced to study DRMs biophysical association state. Langmuir analysis of Cav-1 containing DRMs displayed an isotherm with a clear-cut feature, suggesting the coexistence of the liquid-ordered (Lo) phase typical of the raft structure, namely "cholesterol-rich Lo phase," with a phase fully missing in Cav-1 KO that we named "caveolin-induced Lo phase." Furthermore, while the sole lipid component of both WT and KO-DRMs showed qualitatively similar isotherm configuration, the reinsertion of recombinant Cav-1 into WT-DRMs lipids restored the WT-DRM pattern. X-ray diffraction results confirmed that Cav-1 causes the formation of a "caveolin-induced Lo phase," as suggested by Langmuir experiments, allowing us to speculate about a possible structural model. These results show that the unique molecular link between Cav-1 and cholesterol can spur functional order in a lipid bilayer strictly derived from biological sources.
Assuntos
Caveolina 1/metabolismo , Colesterol/metabolismo , Proteômica/métodos , Animais , Cavéolas/metabolismo , Humanos , Difração de Raios XAssuntos
Emigrantes e Imigrantes/estatística & dados numéricos , Tuberculose Resistente a Múltiplos Medicamentos/diagnóstico , Tuberculose Resistente a Múltiplos Medicamentos/etnologia , Tuberculose Resistente a Múltiplos Medicamentos/epidemiologia , Adulto , Idoso , Controle de Doenças Transmissíveis , Feminino , Humanos , Itália , Masculino , Pessoa de Meia-Idade , Prevalência , Características de ResidênciaRESUMO
Current tuberculosis (TB) treatment requires 6 months of combination therapy with isoniazid (INH), rifampin (RIF), pyrazinamide (PZA), and ethambutol for active TB and 9 months of INH or 3 months of rifapentine (RFP) + INH for latent TB. The lungs of patients with active and latent TB contain heterogeneous mixtures of cellular and caseous granulomas harboring Mycobacterium tuberculosis bacilli ranging from actively replicating (AR) to nonreplicating (NR), phenotypically drug-resistant stages. Several in vitro models to obtain NR cells were reported, including exposure to hypoxia, nutrient starvation, acid + nitric oxide, and stationary phase. Overall, these models showed that RIF, RFP, PA-824 (PA), metronidazole (MZ), bedaquiline (BQ), and fluoroquinolones were the most active drugs against NR M. tuberculosis. In hypoxia at pH 5.8, some combinations killed AR plus NR cells, as shown by lack of regrowth in liquid media, whereas in hypoxia at pH 7.3 (the pH of the caseum), only RIF and RFP efficiently killed NR bacilli while several other drugs showed little effect. In conventional mouse models, combinations containing RFP, BQ, PA, PZA, moxifloxacin, sutezolid, linezolid, and clofazimine sterilized animals in ≤2 months, as shown by lack of viable bacilli in lung homogenates after 3 months without therapy. Drugs were less effective in C3HeB/FeJ mice forming caseous granulomas. Overall, in vitro observations and in vivo studies suggest that the search for new TB drugs could be addressed to low lipophilic molecules (e.g., new rpoB inhibitors with clogP < 3) killing NR M. tuberculosis in hypoxia at neutral pH and reaching high rates of unbound drug in the caseum.
Assuntos
Antituberculosos/uso terapêutico , Tuberculose Latente/tratamento farmacológico , Mycobacterium tuberculosis/efeitos dos fármacos , Tuberculose/tratamento farmacológico , Tuberculose/microbiologia , Animais , Antituberculosos/farmacologia , Contagem de Colônia Microbiana , Humanos , Isoniazida/uso terapêutico , Tuberculose Latente/microbiologia , Camundongos , Mycobacterium tuberculosis/fisiologia , Pirazinamida/uso terapêutico , Rifampina/análogos & derivados , Rifampina/uso terapêutico , Tuberculose Pulmonar/tratamento farmacológico , Tuberculose Pulmonar/microbiologiaRESUMO
The physiology of dormant Mycobacterium tuberculosis was studied in detail by examining the gene expression of 51 genes using quantitative Reverse-Transcription Polymerase Chain Reaction. A forty-day period of dormancy in the Wayne culture model depicted four major transcription patterns. Some sigma factors and many metabolic genes were constant, whereas genes belonging to the dormancy regulon were activated on day 9. In particular, alpha-crystallin mRNA showed more than a 1,000-fold increase compared to replicating bacilli. Genes belonging to the enduring hypoxic response were up-regulated at day 16, notably, transcription factors sigma B and E. Early genes typical of log-phase bacilli, esat-6 and fbpB, were uniformly down-regulated during dormancy. Late stages of dormancy showed a drop in gene expression likely due to a lack of substrates in anaerobic respiration as demonstrated by the transcriptional activation observed following nitrates addition. Among genes involved in nitrate metabolism, narG was strongly up-regulated by nitrates addition. Dormant bacilli responded very rapidly when exposed to oxygen and fresh medium, showing a transcriptional activation of many genes, including resuscitation-promoting factors, within one hour. Our observations extend the current knowledge on dormant M. tuberculosis gene expression and its response to nutrients and to aerobic and anaerobic respiration.