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1.
Antimicrob Agents Chemother ; 65(11): e0058321, 2021 10 18.
Artigo em Inglês | MEDLINE | ID: mdl-34370580

RESUMO

Multiple drug discovery initiatives for tuberculosis are currently ongoing to identify and develop new potent drugs with novel targets in order to shorten treatment duration. One of the drug classes with a new mode of action is DprE1 inhibitors targeting an essential process in cell wall synthesis of Mycobacterium tuberculosis. In this investigation, three DprE1 inhibitors currently in clinical trials, TBA-7371, PBTZ169, and OPC-167832, were evaluated side-by-side as single agents in the C3HeB/FeJ mouse model presenting with caseous necrotic pulmonary lesions upon tuberculosis infection. The goal was to confirm the efficacy of the DprE1 inhibitors in a mouse tuberculosis model with advanced pulmonary pathology and perform comprehensive analysis of plasma, lung, and lesion-centric drug levels to establish pharmacokinetic-pharmacodynamic (PK-PD) parameters predicting efficacy at the site of infection. Results showed significant efficacy for all three DprE1 inhibitors in the C3HeB/FeJ mouse model after 2 months of treatment. Superior efficacy was observed for OPC-167832 even at low-dose levels, which can be attributed to its low MIC, favorable distribution, and sustained retention above the MIC throughout the dosing interval in caseous necrotic lesions, where the majority of bacteria reside in C3HeB/FeJ mice. These results support further progression of the three drug candidates through clinical development for tuberculosis treatment.


Assuntos
Mycobacterium tuberculosis , Tiazinas , Tuberculose , Animais , Camundongos , Camundongos Endogâmicos C3H , Piperazinas , Tuberculose/tratamento farmacológico
2.
Nat Commun ; 12(1): 2899, 2021 05 18.
Artigo em Inglês | MEDLINE | ID: mdl-34006838

RESUMO

There is urgent need for new drug regimens that more rapidly cure tuberculosis (TB). Existing TB drugs and regimens vary in treatment-shortening activity, but the molecular basis of these differences is unclear, and no existing assay directly quantifies the ability of a drug or regimen to shorten treatment. Here, we show that drugs historically classified as sterilizing and non-sterilizing have distinct impacts on a fundamental aspect of Mycobacterium tuberculosis physiology: ribosomal RNA (rRNA) synthesis. In culture, in mice, and in human studies, measurement of precursor rRNA reveals that sterilizing drugs and highly effective drug regimens profoundly suppress M. tuberculosis rRNA synthesis, whereas non-sterilizing drugs and weaker regimens do not. The rRNA synthesis ratio provides a readout of drug effect that is orthogonal to traditional measures of bacterial burden. We propose that this metric of drug activity may accelerate the development of shorter TB regimens.


Assuntos
Antituberculosos/administração & dosagem , Mycobacterium tuberculosis/efeitos dos fármacos , Precursores de RNA/metabolismo , RNA Ribossômico/metabolismo , Tuberculose/tratamento farmacológico , Animais , Modelos Animais de Doenças , Feminino , Humanos , Camundongos Endogâmicos BALB C , Mycobacterium tuberculosis/genética , Mycobacterium tuberculosis/fisiologia , Precursores de RNA/genética , RNA Bacteriano/genética , RNA Bacteriano/metabolismo , RNA Ribossômico/genética , Resultado do Tratamento , Tuberculose/diagnóstico , Tuberculose/microbiologia
3.
Vaccines (Basel) ; 9(4)2021 Apr 01.
Artigo em Inglês | MEDLINE | ID: mdl-33916180

RESUMO

The COVID-19 pandemic has generated intense interest in the rapid development and evaluation of vaccine candidates for this disease and other emerging diseases. Several novel methods for preparing vaccine candidates are currently undergoing clinical evaluation in response to the urgent need to prevent the spread of COVID-19. In many cases, these methods rely on new approaches for vaccine production and immune stimulation. We report on the use of a novel method (SolaVAX) for production of an inactivated vaccine candidate and the testing of that candidate in a hamster animal model for its ability to prevent infection upon challenge with SARS-CoV-2 virus. The studies employed in this work included an evaluation of the levels of neutralizing antibody produced post-vaccination, levels of specific antibody sub-types to RBD and spike protein that were generated, evaluation of viral shedding post-challenge, flow cytometric and single cell sequencing data on cellular fractions and histopathological evaluation of tissues post-challenge. The results from this preliminary evaluation provide insight into the immunological responses occurring as a result of vaccination with the proposed vaccine candidate and the impact that adjuvant formulations, specifically developed to promote Th1 type immune responses, have on vaccine efficacy and protection against infection following challenge with live SARS-CoV-2. This data may have utility in the development of effective vaccine candidates broadly. Furthermore, the results of this preliminary evaluation suggest that preparation of a whole virion vaccine for COVID-19 using this specific photochemical method may have potential utility in the preparation of one such vaccine candidate.

4.
Sci Rep ; 11(1): 9040, 2021 04 27.
Artigo em Inglês | MEDLINE | ID: mdl-33907221

RESUMO

The nontuberculous mycobacteria (NTM) Mycobacterium avium is a clinically significant pathogen that can cause a wide range of maladies, including tuberculosis-like pulmonary disease. An immunocompromised host status, either genetically or acutely acquired, presents a large risk for progressive NTM infections. Due to this quietly emerging health threat, we evaluated the ability of a recombinant fusion protein ID91 combined with GLA-SE [glucopyranosyl lipid adjuvant, a toll like receptor 4 agonist formulated in an oil-in-water stable nano-emulsion] to confer protection in both C57BL/6 (wild type) and Beige (immunocompromised) mouse models. We optimized an aerosol challenge model using a clinical NTM isolate: M. avium 2-151 smt, observed bacterial growth kinetics, colony morphology, drug sensitivity and histopathology, characterized the influx of pulmonary immune cells, and confirmed the immunogenicity of ID91 in both mouse models. To determine prophylactic vaccine efficacy against this M. avium isolate, mice were immunized with either ID91 + GLA-SE or bacillus Calmette-Guérin (BCG). Immunocompromised Beige mice displayed a delayed influx of innate and adaptive immune cells resulting in a sustained and increased bacterial burden in the lungs and spleen compared to C57BL/6 mice. Importantly, both ID91 + GLA-SE and BCG vaccines significantly reduced pulmonary bacterial burden in both mouse strains. This work is a proof-of-concept study of subunit vaccine-induced protection against NTM.


Assuntos
Vacina BCG/administração & dosagem , Modelos Animais de Doenças , Hospedeiro Imunocomprometido/imunologia , Mycobacterium avium/patogenicidade , Tuberculose/prevenção & controle , Vacinas de Subunidades/administração & dosagem , Animais , Vacina BCG/imunologia , Feminino , Camundongos , Camundongos Endogâmicos C57BL , Mycobacterium avium/metabolismo , Tuberculose/imunologia , Tuberculose/microbiologia , Vacinação , Vacinas de Subunidades/imunologia
5.
Sci Rep ; 10(1): 16257, 2020 10 01.
Artigo em Inglês | MEDLINE | ID: mdl-33004826

RESUMO

Tuberculosis (TB) is a chronic inflammatory disease that is often associated with alterations in systemic and cellular metabolism that resolves following successful antimicrobial drug treatment. We hypothesized that altered systemic glucose metabolism as a consequence of Mycobacterium tuberculosis (Mtb) infection, contributes to TB pathogenesis, and when normalized with anti-glycemic drugs would improve clinical outcomes. To test this hypothesis, guinea pigs were treated daily with the anti-diabetic drug metformin starting 4 weeks prior or concurrent with aerosol exposure to the H37Rv strain of Mtb. In the chronic stages of infection, Mtb infected metformin-treated animals had restored systemic insulin sensitivity but remained glucose intolerant as determined by oral glucose tolerance testing. Despite persistent glucose intolerance, metformin-treated guinea pigs had a 2.8-fold reduction in lung lesion burden and a 0.7 log decrease in CFUs. An alternative hypothesis that metformin treatment improved clinical disease by having a direct effect on immune cell energy metabolism was tested using extracellular flux analysis and flow cytometry. The proinflammatory immune response to Mtb infection in untreated guinea pigs was associated with a marked increase in energy metabolism (glycolysis and mitochondrial respiration) of peripheral blood mononuclear cells (PBMCs), which was normalized in metformin-treated guinea pigs. Moreover, both CD4+ and CD8+ T lymphocytes from Mtb infected, metformin treated animals maintained a more normal mitochondrial membrane potential while those isolated from untreated animals had persistent mitochondrial hyperpolarization. These data suggest that metformin promotes natural host resistance to Mtb infection by maintaining immune cell metabolic homeostasis and function during the chronic stages of active TB disease.


Assuntos
Hipoglicemiantes/uso terapêutico , Metformina/uso terapêutico , Tuberculose Pulmonar/tratamento farmacológico , Animais , Citocinas/metabolismo , Metabolismo Energético/efeitos dos fármacos , Glucose/metabolismo , Intolerância à Glucose/tratamento farmacológico , Cobaias , Resistência à Insulina , Pulmão/patologia , Masculino , Potencial da Membrana Mitocondrial/efeitos dos fármacos , Superóxidos/metabolismo , Linfócitos T/metabolismo , Tuberculose Pulmonar/metabolismo , Tuberculose Pulmonar/patologia
6.
Vaccine ; 38(45): 7156-7165, 2020 10 21.
Artigo em Inglês | MEDLINE | ID: mdl-32978002

RESUMO

Although vaccination with BCG prevents disseminated forms of childhood tuberculosis (TB), it does not protect against pulmonary infection or Mycobacterium tuberculosis (Mtb) transmission. In this study, we generated a complete deletion mutant of the Mtb Esx-5 type VII secretion system (Mtb Δesx-5). Mtb Δesx-5 was highly attenuated and safe in immunocompromised mice. When tested as a vaccine candidate to boost BCG-primed immunity, Mtb Δesx-5 improved protection against highly virulent Mtb strains in the murine and guinea pig models of TB. Enhanced protection provided by heterologous BCG-prime plus Mtb Δesx-5 boost regimen was associated with increased pulmonary influx of central memory T cells (TCM), follicular helper T cells (TFH) and activated monocytes. Conversely, lower numbers of T cells expressing exhaustion markers were observed in vaccinated animals. Our results suggest that boosting BCG-primed immunity with Mtb Δesx-5 is a potential approach to improve protective immunity against Mtb. Further insight into the mechanism of action of this novel prime-boost approach is warranted.


Assuntos
Mycobacterium bovis , Mycobacterium tuberculosis , Tuberculose , Sistemas de Secreção Tipo VII , Animais , Antígenos de Bactérias , Vacina BCG , Cobaias , Imunização Secundária , Camundongos , Mycobacterium tuberculosis/genética , Tuberculose/prevenção & controle , Vacinação
7.
Viruses ; 12(6)2020 06 17.
Artigo em Inglês | MEDLINE | ID: mdl-32560467

RESUMO

Dengue virus infection is associated with the upregulation of metabolic pathways within infected cells. This effect is common to infection by a broad array of viruses. These metabolic changes, including increased glucose metabolism, oxidative phosphorylation and autophagy, support the demands of viral genome replication and infectious particle formation. The mechanisms by which these changes occur are known to be, in part, directed by viral nonstructural proteins that contact and control cellular structures and metabolic enzymes. We investigated the roles of host proteins with overarching control of metabolic processes, the transcriptional regulators, cyclin-dependent kinase 8 (CDK8) and its paralog, CDK19, as mediators of virally induced metabolic changes. Here, we show that expression of CDK8, but not CDK19, is increased during dengue virus infection in Huh7 human hepatocellular carcinoma cells, although both are required for efficient viral replication. Chemical inhibition of CDK8 and CDK19 with Senexin A during infection blocks virus-induced expression of select metabolic and autophagic genes, hexokinase 2 (HK2) and microtubule-associated protein 1 light chain 3 (LC3), and reduces viral genome replication and infectious particle production. The results further define the dependence of virus replication on increased metabolic capacity in target cells and identify CDK8 and CDK19 as master regulators of key metabolic genes. The common inhibition of CDK8 and CDK19 offers a host-directed therapeutic intervention that is unlikely to be overcome by viral evolution.


Assuntos
Quinase 8 Dependente de Ciclina/metabolismo , Quinases Ciclina-Dependentes/metabolismo , Vírus da Dengue/crescimento & desenvolvimento , Metabolismo Energético/fisiologia , Replicação Viral/genética , Autofagia/fisiologia , Linhagem Celular Tumoral , Quinase 8 Dependente de Ciclina/antagonistas & inibidores , Quinases Ciclina-Dependentes/antagonistas & inibidores , Dengue/patologia , Vírus da Dengue/metabolismo , Técnicas de Silenciamento de Genes , Genoma Viral/genética , Glucose/metabolismo , Hexoquinase/biossíntese , Humanos , Masculino , Proteínas Associadas aos Microtúbulos/biossíntese , Pessoa de Meia-Idade , Fosforilação Oxidativa
8.
Sci Rep ; 10(1): 6047, 2020 04 08.
Artigo em Inglês | MEDLINE | ID: mdl-32269234

RESUMO

Efforts to develop effective and safe drugs for treatment of tuberculosis require preclinical evaluation in animal models. Alongside efficacy testing of novel therapies, effects on pulmonary pathology and disease progression are monitored by using histopathology images from these infected animals. To compare the severity of disease across treatment cohorts, pathologists have historically assigned a semi-quantitative histopathology score that may be subjective in terms of their training, experience, and personal bias. Manual histopathology therefore has limitations regarding reproducibility between studies and pathologists, potentially masking successful treatments. This report describes a pathologist-assistive software tool that reduces these user limitations, while providing a rapid, quantitative scoring system for digital histopathology image analysis. The software, called 'Lesion Image Recognition and Analysis' (LIRA), employs convolutional neural networks to classify seven different pathology features, including three different lesion types from pulmonary tissues of the C3HeB/FeJ tuberculosis mouse model. LIRA was developed to improve the efficiency of histopathology analysis for mouse tuberculosis infection models, this approach has also broader applications to other disease models and tissues. The full source code and documentation is available from https://Github.com/TB-imaging/LIRA.


Assuntos
Processamento de Imagem Assistida por Computador/métodos , Pulmão/diagnóstico por imagem , Mycobacterium tuberculosis/fisiologia , Tuberculose Pulmonar/diagnóstico por imagem , Algoritmos , Animais , Modelos Animais de Doenças , Humanos , Pulmão/patologia , Camundongos , Camundongos Endogâmicos C3H , Redes Neurais de Computação , Software , Tuberculose Pulmonar/patologia
9.
Curr HIV Res ; 18(1): 19-28, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-31870268

RESUMO

The main advantage of animal models of infectious diseases over in vitro studies is the gain in the understanding of the complex dynamics between the immune system and the pathogen. While small animal models have practical advantages over large animal models, it is crucial to be aware of their limitations. Although the small animal model at least needs to be susceptible to the pathogen under study to obtain meaningful data, key elements of pathogenesis should also be reflected when compared to humans. Well-designed small animal models for HIV, hepatitis viruses and tuberculosis require, additionally, a thorough understanding of the similarities and differences in the immune responses between humans and small animals and should incorporate that knowledge into the goals of the study. To discuss these considerations, the NIAID hosted a workshop on 'Small Animal Models for HIV, Hepatitis B, and Tuberculosis' on May 30, 2019. Highlights of the workshop are outlined below.


Assuntos
Modelos Animais de Doenças , Infecções por HIV/patologia , HIV-1/imunologia , Vírus da Hepatite B/imunologia , Hepatite B/patologia , Mycobacterium tuberculosis/imunologia , Tuberculose/patologia , Animais , Coinfecção/microbiologia , Cobaias , Infecções por HIV/imunologia , Hepatite B/imunologia , Humanos , Macaca mulatta , Marmota , Camundongos , National Institute of Allergy and Infectious Diseases (U.S.) , Coelhos , Tuberculose/imunologia , Estados Unidos
10.
Elife ; 72018 11 14.
Artigo em Inglês | MEDLINE | ID: mdl-30427309

RESUMO

Understanding the distribution patterns of antibiotics at the site of infection is paramount to selecting adequate drug regimens and developing new antibiotics. Tuberculosis (TB) lung lesions are made of various immune cell types, some of which harbor persistent forms of the pathogen, Mycobacterium tuberculosis. By combining high resolution MALDI MSI with histology staining and quantitative image analysis in rabbits with active TB, we have mapped the distribution of a fluoroquinolone at high resolution, and identified the immune-pathological factors driving its heterogeneous penetration within TB lesions, in relation to where bacteria reside. We find that macrophage content, distance from lesion border and extent of necrosis drive the uneven fluoroquinolone penetration. Preferential uptake in macrophages and foamy macrophages, where persistent bacilli reside, compared to other immune cells present in TB granulomas, was recapitulated in vitro using primary human cells. A nonlinear modeling approach was developed to help predict the observed drug behavior in TB lesions. This work constitutes a methodological advance for the co-localization of drugs and infectious agents at high spatial resolution in diseased tissues, which can be applied to other diseases with complex immunopathology.


Assuntos
Antituberculosos/farmacocinética , Fluoroquinolonas/farmacocinética , Tuberculose/tratamento farmacológico , Tuberculose/patologia , Animais , Antituberculosos/administração & dosagem , Antituberculosos/análise , Modelos Animais de Doenças , Fluoroquinolonas/administração & dosagem , Fluoroquinolonas/análise , Histocitoquímica , Macrófagos/química , Macrófagos/microbiologia , Mycobacterium tuberculosis/crescimento & desenvolvimento , Coelhos , Espectrometria de Massas por Ionização e Dessorção a Laser Assistida por Matriz
11.
PLoS One ; 13(10): e0205882, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-30312355

RESUMO

Historically, vaccination with Coxiella burnetii whole cell vaccines has induced hypersensitivity reactions in humans and animals that have had prior exposure to the pathogen as a result of infection or vaccination. Intradermal skin testing is routinely used to evaluate exposure in humans, and guinea pig hypersensitivity models have been developed to characterize the potential for reactogenicity in vaccine candidates. Here we describe a refinement of the guinea pig model using an alternate vaccine for positive controls. An initial comparative study used viable C. burnetii to compare the routes of sensitizing exposure of guinea pigs (intranasal vs intraperitoneal), evaluation of two time points for antigen challenge (21 and 42 days) and an assessment of two routes (intradermal and subcutaneous) of challenge using the ruminant vaccine Coxevac as the antigenic control. Animals sensitized by intraperitoneal exposure exhibited slightly larger gross reactions than did those sensitized by intranasal exposure, and reactions were more pronounced when skin challenge was performed at 42 days compared to 21 days post-sensitization. The intradermal route proved to be the optimal route of reactogenicity challenge. Histopathological changes at injection sites were similar to those previously reported and a scoring system was developed to compare reactions between groups receiving vaccine by intradermal versus subcutaneous routes. Based on the comparative study, a standardized protocol for assessment of vaccine reactogenicity in intranasally-sensitized animals was tested in a larger confirmatory study. Results suggest that screens utilizing a group size of n = 3 would achieve 90% power for detecting exposure-related reactogenic responses of the magnitude induced by Coxevac using either of two outcome measures.


Assuntos
Vacinas Bacterianas/imunologia , Coxiella burnetii , Febre Q/prevenção & controle , Administração Intranasal , Animais , Antígenos/química , Modelos Animais de Doenças , Ensaio de Imunoadsorção Enzimática , Feminino , Cobaias , Injeções Intradérmicas , Injeções Subcutâneas , Pele , Resultado do Tratamento , Vacinação
12.
J Exp Med ; 215(8): 1975-1986, 2018 08 06.
Artigo em Inglês | MEDLINE | ID: mdl-30018074

RESUMO

In the 1970s, inclusion of pyrazinamide (PZA) in the drug regimen of tuberculosis (TB) patients for the first 2 mo achieved a drastic reduction of therapy duration. Until now, however, the mechanisms underlying PZA's unique contribution to efficacy have remained controversial, and animal efficacy data vary across species. To understand how PZA kills bacterial populations present in critical lung lesion compartments, we first characterized a rabbit model of active TB, showing striking similarities in lesion types and fates to nonhuman primate models deemed the most appropriate surrogates of human TB. We next employed this model with lesion-centric molecular and bacteriology readouts to demonstrate that PZA exhibits potent activity against Mycobacterium tuberculosis residing in difficult-to-sterilize necrotic lesions. Our data also indicate that PZA is slow acting, suggesting that PZA administration beyond the first 2 mo may accelerate the cure. In conclusion, we provide a pharmacodynamic explanation for PZA's treatment-shortening effect and deliver new tools to dissect the contribution of immune response versus drug at the lesion level.


Assuntos
Antituberculosos/uso terapêutico , Pirazinamida/uso terapêutico , Tuberculose/tratamento farmacológico , Tuberculose/imunologia , Animais , Antituberculosos/farmacologia , Modelos Animais de Doenças , Feminino , Humanos , Viabilidade Microbiana/efeitos dos fármacos , Mycobacterium tuberculosis/efeitos dos fármacos , Mycobacterium tuberculosis/fisiologia , Necrose , Neutrófilos/efeitos dos fármacos , Neutrófilos/metabolismo , Pirazinamida/farmacologia , Coelhos , Tuberculose/microbiologia , Tuberculose/patologia
13.
Vaccines (Basel) ; 6(2)2018 May 24.
Artigo em Inglês | MEDLINE | ID: mdl-29795025

RESUMO

It is estimated that one third of the world's population is infected with Mycobacterium tuberculosis (Mtb). This astounding statistic, in combination with costly and lengthy treatment regimens make the development of therapeutic vaccines paramount for controlling the global burden of tuberculosis. Unlike prophylactic vaccination, therapeutic immunization relies on the natural pulmonary infection with Mtb as the mucosal prime that directs boost responses back to the lung. The purpose of this work was to determine the protection and safety profile over time following therapeutic administration of our lead Mtb vaccine candidate, ID93 with a synthetic TLR4 agonist (glucopyranosyl lipid adjuvant in a stable emulsion (GLA-SE)), in combination with rifampicin, isoniazid, and pyrazinamide (RHZ) drug treatment. We assessed the host inflammatory immune responses and lung pathology 7⁻22 weeks post infection, and determined the therapeutic efficacy of combined treatment by enumeration of the bacterial load and survival in the SWR/J mouse model. We show that drug treatment alone, or with immunotherapy, tempered the inflammatory responses measured in brochoalveolar lavage fluid and plasma compared to untreated cohorts. RHZ combined with therapeutic immunizations significantly enhanced TH1-type cytokine responses in the lung over time, corresponding to decreased pulmonary pathology evidenced by a significant decrease in the percentage of lung lesions and destructive lung inflammation. These data suggest that bacterial burden assessment alone may miss important correlates of lung architecture that directly contribute to therapeutic vaccine efficacy in the preclinical mouse model. We also confirmed our previous finding that in combination with antibiotics therapeutic immunizations provide an additive survival advantage. Moreover, therapeutic immunizations with ID93/GLA-SE induced differential T cell immune responses over the course of infection that correlated with periods of enhanced bacterial control over that of drug treatment alone. Here we advance the immunotherapy model and investigate reliable correlates of protection and Mtb control.

14.
PLoS One ; 12(7): e0180925, 2017.
Artigo em Inglês | MEDLINE | ID: mdl-28749949

RESUMO

There is an urgent need to develop new drug treatment strategies to control the global spread of drug-sensitive and multidrug-resistant Mycobacterium tuberculosis (M. tuberculosis). The ß-lactam class of antibiotics is among the safest and most widely prescribed antibiotics, but they are not effective against M. tuberculosis due to intrinsic resistance. This study shows that 2-aminoimidazole (2-AI)-based small molecules potentiate ß-lactam antibiotics against M. tuberculosis. Active 2-AI compounds significantly reduced the minimal inhibitory and bactericidal concentrations of ß-lactams by increasing M. tuberculosis cell envelope permeability and decreasing protein secretion including ß-lactamase. Metabolic labeling and transcriptional profiling experiments revealed that 2-AI compounds impair mycolic acid biosynthesis, export and linkage to the mycobacterial envelope, counteracting an important defense mechanism reducing permeability to external agents. Additionally, other important constituents of the M. tuberculosis outer membrane including sulfolipid-1 and polyacyltrehalose were also less abundant in 2-AI treated bacilli. As a consequence of 2-AI treatment, M. tuberculosis displayed increased sensitivity to SDS, increased permeability to nucleic acid staining dyes, and rapid binding of cell wall targeting antibiotics. Transcriptional profiling analysis further confirmed that 2-AI induces transcriptional regulators associated with cell envelope stress. 2-AI based small molecules potentiate the antimicrobial activity of ß-lactams by a mechanism that is distinct from specific inhibitors of ß-lactamase activity and therefore may have value as an adjunctive anti-TB treatment.


Assuntos
Anti-Infecciosos/farmacologia , Permeabilidade da Membrana Celular/efeitos dos fármacos , Imidazóis/farmacologia , Mycobacterium tuberculosis/citologia , Mycobacterium tuberculosis/enzimologia , beta-Lactamases/metabolismo , beta-Lactamas/farmacologia , Carbenicilina/farmacologia , Corantes/química , Lipídeos/análise , Testes de Sensibilidade Microbiana , Mycobacterium tuberculosis/efeitos dos fármacos , Mycobacterium tuberculosis/crescimento & desenvolvimento , Ácidos Nucleicos/metabolismo , Penicilina V/farmacologia , Dodecilsulfato de Sódio/farmacologia , Coloração e Rotulagem , Transcrição Genética/efeitos dos fármacos , Vancomicina/farmacologia
15.
Dis Model Mech ; 10(2): 151-162, 2017 02 01.
Artigo em Inglês | MEDLINE | ID: mdl-28093504

RESUMO

Type 2 diabetes is a leading cause of morbidity and mortality among noncommunicable diseases, and additional animal models that more closely replicate the pathogenesis of human type 2 diabetes are needed. The goal of this study was to develop a model of type 2 diabetes in guinea pigs, in which diet-induced glucose intolerance precedes ß-cell cytotoxicity, two processes that are crucial to the development of human type 2 diabetes. Guinea pigs developed impaired glucose tolerance after 8 weeks of feeding on a high-fat, high-carbohydrate diet, as determined by oral glucose challenge. Diet-induced glucose intolerance was accompanied by ß-cell hyperplasia, compensatory hyperinsulinemia, and dyslipidemia with hepatocellular steatosis. Streptozotocin (STZ) treatment alone was ineffective at inducing diabetic hyperglycemia in guinea pigs, which failed to develop sustained glucose intolerance or fasting hyperglycemia and returned to euglycemia within 21 days after treatment. However, when high-fat, high-carbohydrate diet-fed guinea pigs were treated with STZ, glucose intolerance and fasting hyperglycemia persisted beyond 21 days post-STZ treatment. Guinea pigs with diet-induced glucose intolerance subsequently treated with STZ demonstrated an insulin-secretory capacity consistent with insulin-independent diabetes. This insulin-independent state was confirmed by response to oral antihyperglycemic drugs, metformin and glipizide, which resolved glucose intolerance and extended survival compared with guinea pigs with uncontrolled diabetes. In this study, we have developed a model of sequential glucose intolerance and ß-cell loss, through high-fat, high-carbohydrate diet and extensive optimization of STZ treatment in the guinea pig, which closely resembles human type 2 diabetes. This model will prove useful in the study of insulin-independent diabetes pathogenesis with or without comorbidities, where the guinea pig serves as a relevant model species.


Assuntos
Diabetes Mellitus Experimental/complicações , Diabetes Mellitus Tipo 2/complicações , Intolerância à Glucose/complicações , Administração Oral , Antagonistas de Receptores Adrenérgicos alfa 2/farmacologia , Animais , Morte Celular/efeitos dos fármacos , Diabetes Mellitus Experimental/tratamento farmacológico , Diabetes Mellitus Experimental/patologia , Diabetes Mellitus Tipo 2/tratamento farmacológico , Diabetes Mellitus Tipo 2/patologia , Dieta , Carboidratos da Dieta , Modelos Animais de Doenças , Dislipidemias/complicações , Dislipidemias/tratamento farmacológico , Dislipidemias/patologia , Comportamento Alimentar , Feminino , Intolerância à Glucose/tratamento farmacológico , Intolerância à Glucose/patologia , Cobaias , Hiperglicemia/complicações , Hiperglicemia/tratamento farmacológico , Hiperglicemia/patologia , Hiperinsulinismo/complicações , Hiperinsulinismo/tratamento farmacológico , Hiperplasia , Hipoglicemiantes/administração & dosagem , Hipoglicemiantes/farmacologia , Hipoglicemiantes/uso terapêutico , Insulina/farmacologia , Insulina/uso terapêutico , Células Secretoras de Insulina/efeitos dos fármacos , Células Secretoras de Insulina/metabolismo , Células Secretoras de Insulina/patologia , Estreptozocina , Análise de Sobrevida , Ganho de Peso/efeitos dos fármacos
16.
Vector Borne Zoonotic Dis ; 16(6): 400-7, 2016 06.
Artigo em Inglês | MEDLINE | ID: mdl-27111674

RESUMO

Bluetongue virus (BTV) is an orbivirus transmitted by biting midges (Culicoides spp.) that can result in moderate to high morbidity and mortality primarily in sheep and white-tailed deer. Although only 5 serotypes of BTV are considered endemic to the United States, as many as 11 incursive serotypes have been detected in livestock and wildlife in the past 16 years. Introductions of serotypes, with unknown virulence and disease risk, are constant threats to US agriculture. One potential incursive serotype of particular concern is the European strain of BTV-8, which was introduced into Northern Europe in 2006 and caused unprecedented livestock disease and mortality during the 2006-2007 vector seasons. To assess disease risk of BTV-8 in a common white-faced American sheep breed, eight Polled Dorset yearlings were experimentally infected and monitored for clinical signs. Viremia and viral tissue distribution were detected and quantified by real-time qRT-PCR. Overall, clinical disease was moderate with no mortality. Viremia reached as high as 9.7 log10 particles/mL and persisted at 5 logs or higher through the end of the study (28 days). Virus distribution in tissues was extensive with the highest mean titers at the peak of viremia (day 8) in the kidney (8.38 log10 particles/mg) and pancreas (8.37 log10 particles/mg). Virus persisted in tissues of some sheep at 8 logs or higher by day 28. Results of this study suggest that should BTV-8 emerge in the United States, clinical disease in this common sheep breed would likely be similar in form, duration, and severity to what is typically observed in severe outbreaks of endemic serotypes, not the extraordinary disease levels seen in Northern Europe. In addition, a majority of exposed sheep would be expected to survive and act as significant BTV-8 reservoirs with high titer viremias for subsequent transmission to other livestock and wildlife populations.


Assuntos
Vírus Bluetongue/classificação , Bluetongue/virologia , Animais , Bluetongue/epidemiologia , Bluetongue/genética , Bluetongue/patologia , Vírus Bluetongue/genética , Europa (Continente)/epidemiologia , Feminino , Predisposição Genética para Doença , Fatores de Risco , Ovinos , Estados Unidos/epidemiologia , Viremia , Replicação Viral
17.
Semin Immunopathol ; 38(2): 167-83, 2016 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-26510950

RESUMO

Infection by the intracellular bacterial pathogen Mycobacterium tuberculosis (Mtb) is a major cause of morbidity and mortality worldwide. Slow progress has been made in lessening the impact of tuberculosis (TB) on human health, especially in parts of the world where Mtb is endemic. Due to the complexity of TB disease, there is still an urgent need to improve diagnosis, prevention, and treatment strategies to control global spread of disease. Active research targeting avenues to prevent infection or transmission through vaccination, to diagnose asymptomatic carriers of Mtb, and to improve antimicrobial drug treatment responses is ongoing. However, this research is hampered by a relatively poor understanding of the pathogenesis of early infection and the factors that contribute to host susceptibility, protection, and the development of active disease. There is increasing interest in the development of adjunctive therapy that will aid the host in responding to Mtb infection appropriately thereby improving the effectiveness of current and future drug treatments. In this review, we summarize what is known about the host response to Mtb infection in humans and animal models and highlight potential therapeutic targets involved in TB granuloma formation and resolution. Strategies designed to shift the balance of TB granuloma formation toward protective rather than destructive processes are discussed based on our current knowledge. These therapeutic strategies are based on the assumption that granuloma formation, although thought to prevent the spread of the tubercle bacillus within and between individuals contributes to manifestations of active TB disease in human patients when left unchecked. This effect of granuloma formation favors the spread of infection and impairs antimicrobial drug treatment. By gaining a better understanding of the mechanisms by which Mtb infection contributes to irreversible tissue damage, down regulates protective immune responses, and delays tissue healing, new treatment strategies can be rationally designed. Granuloma-targeted therapy is advantageous because it allows for the repurpose of existing drugs used to treat other communicable and non-communicable diseases as adjunctive therapies combined with existing and future anti-TB drugs. Thus, the development of adjunctive, granuloma-targeted therapy, like other host-directed therapies, may benefit from the availability of approved drugs to aid in treatment and prevention of TB. In this review, we have attempted to summarize the results of published studies in the context of new innovative approaches to host-directed therapy that need to be more thoroughly explored in pre-clinical animal studies and in human clinical trials.


Assuntos
Antituberculosos/uso terapêutico , Granuloma/tratamento farmacológico , Interações Hospedeiro-Patógeno , Mycobacterium tuberculosis/imunologia , Tuberculose/tratamento farmacológico , Tuberculose/imunologia , Inibidores da Angiogênese/farmacologia , Inibidores da Angiogênese/uso terapêutico , Animais , Antituberculosos/farmacologia , Farmacorresistência Bacteriana , Metabolismo Energético/efeitos dos fármacos , Matriz Extracelular/imunologia , Matriz Extracelular/metabolismo , Matriz Extracelular/patologia , Granuloma/etiologia , Granuloma/metabolismo , Granuloma/patologia , Interações Hospedeiro-Patógeno/imunologia , Humanos , Sistema Imunitário/imunologia , Sistema Imunitário/metabolismo , Sistema Imunitário/microbiologia , Sistema Imunitário/patologia , Mycobacterium tuberculosis/efeitos dos fármacos , Mycobacterium tuberculosis/genética , Mycobacterium tuberculosis/metabolismo , Neovascularização Patológica/tratamento farmacológico , Neovascularização Patológica/genética , Neovascularização Patológica/metabolismo , Transdução de Sinais/efeitos dos fármacos , Tuberculose/complicações , Tuberculose/microbiologia
18.
Clin Vaccine Immunol ; 23(2): 137-47, 2016 02.
Artigo em Inglês | MEDLINE | ID: mdl-26656121

RESUMO

Mycobacterium tuberculosis HN878 represents a virulent clinical strain from the W-Beijing family, which has been tested in small animal models in order to study its virulence and its induction of host immune responses following infection. This isolate causes death and extensive lung pathology in infected C57BL/6 mice, whereas lab-adapted strains, such as M. tuberculosis H37Rv, do not. The use of this clinically relevant isolate of M. tuberculosis increases the possibilities of assessing the long-lived efficacy of tuberculosis vaccines in a relatively inexpensive small animal model. This model will also allow for the use of knockout mouse strains to critically examine key immunological factors responsible for long-lived, vaccine-induced immunity in addition to vaccine-mediated prevention of pulmonary immunopathology. In this study, we show that the ID93/glucopyranosyl lipid adjuvant (GLA)-stable emulsion (SE) tuberculosis vaccine candidate, currently in human clinical trials, is able to elicit protection against M. tuberculosis HN878 by reducing the bacterial burden in the lung and spleen and by preventing the extensive lung pathology induced by this pathogen in C57BL/6 mice.


Assuntos
Mycobacterium tuberculosis/imunologia , Células Th1/imunologia , Vacinas contra a Tuberculose/administração & dosagem , Vacinas contra a Tuberculose/imunologia , Tuberculose/imunologia , Tuberculose/prevenção & controle , Adjuvantes Imunológicos , Animais , Carga Bacteriana , Ensaios Clínicos como Assunto , Modelos Animais de Doenças , Humanos , Pulmão/imunologia , Pulmão/microbiologia , Pulmão/patologia , Camundongos Endogâmicos C57BL , Camundongos Knockout , Mycobacterium tuberculosis/isolamento & purificação , Baço/microbiologia , Tuberculose/microbiologia , Vacinas Sintéticas/administração & dosagem , Vacinas Sintéticas/imunologia
19.
Bioorg Med Chem Lett ; 25(21): 4820-4823, 2015 Nov 01.
Artigo em Inglês | MEDLINE | ID: mdl-26187705

RESUMO

The formation of advanced glycation end-products (AGE) as a result of the action of reducing sugars on host macromolecules plays a role in increased morbidity of diabetic patients. There are currently no clinically available therapeutics for the prevention or eradication of AGEs. Following our previous identification of 2-aminoimidazole (2-AI) based AGE inhibitors and breakers, we now report the use of a rapid, scalable, two-step procedure to access a second generation of 2-AI based anti-AGE compounds from commercially available amino acids. Several second generation compounds exhibit increased AGE inhibition and breaking activty compared to the first generation compounds and to the known AGE inhibitor aminoguanidine.


Assuntos
Produtos Finais de Glicação Avançada/antagonistas & inibidores , Imidazóis/farmacologia , Produtos Finais de Glicação Avançada/metabolismo , Imidazóis/síntese química , Imidazóis/química , Estrutura Molecular , Relação Estrutura-Atividade
20.
Tetrahedron Lett ; 56(23): 3406-3409, 2015.
Artigo em Inglês | MEDLINE | ID: mdl-26146419

RESUMO

Advanced glycation end-products (AGEs), unregulated modifications to host macromolecules that occur as a result of metabolic dysregulation, play a role in many diabetes related complications, inflammation and aging, and may lead to increased cardiovascular risk. Small molecules that have the ability to inhibit AGE formation, and even break preformed AGEs have enormous therapeutic potential in the treatment of these disease states. We report the screening of a series of 2-aminoimidazloles for anti-AGE activity, and the identification of a bis-2-aminoimidazole lead compound that possesses superior AGE inhibition and breaking activity compared to the known AGE inhibitor aminoguanidine.

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