Pathogenesis of diabetes-tuberculosis comorbidity, The

2014 Summer.Exposure to the bacterium, Mycobacterium tuberculosis, only leads to the active form of tuberculosis disease (TB) in 5-10% of infected individuals. The development of active TB, at any stage of infection, is often the result of a known TB risk factor, either intrinsic to the individual or acquired as a communicable or non-communicable disease. An association between diabetes and TB has long been recognized, but only recently was diabetes confirmed to increase the risk of developing active TB disease. The convergence of a growing diabetes epidemic on regions with endemic TB has positioned diabetes as an emerging global threat to TB control. Of particular importance is the rapidly growing incidence of type 2 diabetes, which accounts for up to 95% of the global diabetic population. Since the potential impact of this growing comorbidity has only been recently emphasized, little is known regarding the mechanisms of dysregulated immune function and metabolism by which diabetes predisposes to active TB disease. The current understanding of this comorbidity is further limited by the lack of appropriate animal models that replicate the pathogenesis of both human type 2 diabetes and TB. The guinea pig is a well-established model of TB that replicates human pathology and disease progression. This species was emphasized in this series of studies with the goal of better understanding the impact of type 2 diabetes on TB progression and the mechanisms that may change the host response to M. tuberculosis infection. In Chapter 2, we investigated the impact of hyperglycemia alone, induced as post-prandial hyperglycemia through daily administration of sucrose, on TB disease progression in non-diabetic guinea pigs. Guinea pigs receiving daily sucrose developed both higher bacterial burdens in pulmonary and extrapulmonary tissue and also more severe pathology by day 60 of infection. This exacerbated disease manifestation was accompanied by the accumulation of advanced glycation end-products, which are inflammatory by-products of chronic hyperglycemia with known involvement in the development of diabetes-related complications. Interestingly, by monitoring glucose and lipid metabolism in these guinea pigs, we learned that TB alone leads to severe metabolic disturbances, manifesting as hyperglycemia and accumulation of circulating total free fatty acids. From this study, we were able to conclude that not only does mild post-prandial hyperglycemia worsen the course of TB disease in guinea pigs, but also, infection with M. tuberculosis alone induces metabolic disease resembling diabetes, similar to what has been previously reported in human TB. These conclusions rationalize the investigation of novel adjunctive therapies to restore metabolic homeostasis, which may improve the host response to infection, limit bacterial growth, and increase the efficacy of frontline antimicrobial drugs. In Chapter 3, we developed a novel model of type 2 diabetes in the guinea pig to be used in future investigations of type 2 diabetes-TB comorbidity. Previously, the guinea pig as a diabetic model has been described only in the context of β-cell cytotoxicity with the drug, streptozotocin (STZ), but with variable efficacy. In this study, we initially optimized the dose response and STZ preparation to achieve an induction of hyperglycemia that was uniform with limited mortality. This hyperglycemic response was transient but could be stabilized through continued β-cell stress, in the form of a high fat, high sugar diet. Feeding of this modified diet led to impaired glucose tolerance and a compensatory β-cell response that could be abrogated with the use of a single optimized dose of STZ. This novel model of type 2 diabetes develops both insulin resistance and β-cell failure, which replicate the typical progression of type 2 diabetes in humans, all within a reasonable experimental timeframe. From this study, two models emerged, a type 2 diabetic guinea pig as well as a model of impaired glucose tolerance, or prediabetes, that would be used to investigate the mechanisms of diabetes-TB comorbidity. In Chapter 4, the newly developed guinea pig models were used to investigate the overall impact of type 2 diabetes and impaired glucose tolerance on TB progression and the host immune response to M. tuberculosis infection. Although impaired glucose tolerance alone had limited impact on TB progression with exacerbation of disease only at chronic end points, M. tuberculosis infected type 2 diabetic guinea pigs closely resembled the reported manifestations of human diabetes-TB comorbidity including more severe TB disease, higher bacterial burdens, and a robust innate and cell-mediated immune response. Despite evidence of strong Th1 cell-mediated immunity, which is known to be critical for limiting bacterial growth and disease progression, diabetic guinea pigs were unable to control bacterial growth and developed damaging neutrophilic inflammation. To better understand the immune mechanisms leading to uncontrolled bacterial growth and severe disease, in Chapter 5, we investigated the innate and adaptive immune response over the course of early infection in type 2 diabetic guinea pigs. Diabetic guinea pigs were slow to develop early lesions with delayed bacterial transport to the lung draining lymph node, and a corresponding delay in antigen-specific Th1 immunity. Early alterations in cytokine expression were identified that may explain the delayed development of cell-mediated immunity and allow for substantial growth of M. tuberculosis in the lung of infected diabetic guinea pigs. These data indicate that not only does type 2 diabetes increase the severity of TB but also that the chronic inflammatory process associated with TB itself may worsen diabetes. This has important implications worthy of further investigation revolving around the diagnostic criteria for diabetes when associated with TB, the impact of active TB on medical management of diabetes, and the investigation of novel therapeutic targets, both metabolic and immunological, to enhance the host immune response to infection and limit TB disease severity in diabetics

High-resolution mapping of fluoroquinolones in TB rabbit lesions reveals specific distribution in immune cell types.

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

International Veterinary Epilepsy Task Force consensus proposal: Medical treatment of canine epilepsy in Europe

In Europe, the number of antiepileptic drugs (AEDs) licensed for dogs has grown considerably over the last years. Nevertheless, the same questions remain, which include, 1) when to start treatment, 2) which drug is best used initially, 3) which adjunctive AED can be advised if treatment with the initial drug is unsatisfactory, and 4) when treatment changes should be considered. In this consensus proposal, an overview is given on the aim of AED treatment, when to start long-term treatment in canine epilepsy and which veterinary AEDs are currently in use for dogs. The consensus proposal for drug treatment protocols, 1) is based on current published evidence-based literature, 2) considers the current legal framework of the cascade regulation for the prescription of veterinary drugs in Europe, and 3) reflects the authors’ experience. With this paper it is aimed to provide a consensus for the management of canine idiopathic epilepsy. Furthermore, for the management of structural epilepsy AEDs are inevitable in addition to treating the underlying cause, if possible

2015 ACVIM Small Animal Consensus Statement on Seizure Management in Dogs

This report represents a scientific and working clinical consensus statement on seizure management in dogs based on current literature and clinical expertise. The goal was to establish guidelines for a predetermined, concise, and logical sequential approach to chronic seizure management starting with seizure identification and diagnosis (not included in this report), reviewing decision‐making, treatment strategies, focusing on issues related to chronic antiepileptic drug treatment response and monitoring, and guidelines to enhance patient response and quality of life. Ultimately, we hope to provide a foundation for ongoing and future clinical epilepsy research in veterinary medicine

The Impact of Vitamin A Deficiency on Tuberculosis Progression

BACKGROUND: Although previous studies have shown that vitamin A deficiency is associated with incident tuberculosis (TB) disease, the direction of the association has not been established. We investigated the impact of vitamin A deficiency on TB disease progression. METHODS: We conducted a longitudinal cohort study nested within a randomized clinical trial among HIV-infected patients in Haiti. We compared serial vitamin A levels in individuals who developed TB disease to controls matched on age, gender, follow-up time, and time to antiretroviral therapy initiation. We also evaluated histopathology, bacterial load, and immune outcomes in TB infection in a guinea pig model of dietary vitamin A deficiency. RESULTS: Among 773 participants, 96 developed incident TB during follow-up, 62.5% (60) of whom had stored serum samples obtained 90-365 days before TB diagnosis. In age- and sex- adjusted and multivariate analyses, respectively, incident TB cases were 3.99 times (95% confidence interval [CI], 2.41 to 6.60) and 3.59 times (95% CI, 2.05 to 6.29) more likely to have been vitamin A deficient than matched controls. Vitamin A-deficient guinea pigs manifested more extensive pulmonary pathology, atypical granuloma morphology, and increased bacterial growth after experimental TB infection. Reintroduction of dietary vitamin A to deficient guinea pigs after established TB disease successfully abrogated severe disease manifestations and altered cellular immune profiles. CONCLUSIONS: Human and animal studies support the role of baseline vitamin A deficiency as a determinant of future TB disease progression

International Veterinary Epilepsy Task Force recommendations for systematic sampling and processing of brains from epileptic dogs and cats

Traditionally, histological investigations of the epileptic brain are required to identify epileptogenic brain lesions, to evaluate the impact of seizure activity, to search for mechanisms of drug-resistance and to look for comorbidities. For many instances, however, neuropathological studies fail to add substantial data on patients with complete clinical work-up. This may be due to sparse training in epilepsy pathology and or due to lack of neuropathological guidelines for companion animals. The protocols introduced herein shall facilitate systematic sampling and processing of epileptic brains and therefore increase the efficacy, reliability and reproducibility of morphological studies in animals suffering from seizures. Brain dissection protocols of two neuropathological centres with research focus in epilepsy have been optimised with regards to their diagnostic yield and accuracy, their practicability and their feasibility concerning clinical research requirements. The recommended guidelines allow for easy, standardised and ubiquitous collection of brain regions, relevant for seizure generation. Tissues harvested the prescribed way will increase the diagnostic efficacy and provide reliable material for scientific investigations

A preclinical model of TB meningitis to determine drug penetration and activity at the sites of disease.

Tuberculosis meningitis (TBM) is essentially treated with the first-line regimen used against pulmonary tuberculosis, with a prolonged continuation phase. However, clinical outcomes are poor in comparison, for reasons that are only partially understood, highlighting the need for improved preclinical tools to measure drug distribution and activity at the site of disease. A predictive animal model of TBM would also be of great value to prioritize promising drug regimens to be tested in clinical trials, given the healthy state of the development pipeline for the first time in decades. Here, we report the optimization of a rabbit model of TBM disease induced via inoculation of Mycobacterium tuberculosis into the cisterna magna, recapitulating features typical of clinical TBM: neurological deterioration within months post-infection, acid-fast bacilli in necrotic lesions in the brain and spinal cord, and elevated lactate levels in cerebrospinal fluid (CSF). None of the infected rabbits recovered or controlled the disease. We used young adult rabbits, the size of which allows for spatial drug quantitation in critical compartments of the central nervous system that cannot be collected in clinical studies. To illustrate the translational value of the model, we report the penetration of linezolid from plasma into the CSF, meninges, anatomically distinct brain areas, cervical spine, and lumbar spine. Across animals, we measured the bacterial burden concomitant with neurological deterioration, offering a useful readout for drug efficacy studies. The model thus forms the basis for building a preclinical platform to identify improved regimens and inform clinical trial design

International Veterinary Epilepsy Task Force Consensus Proposal: Outcome of therapeutic interventions in canine and feline epilepsy

Common criteria for the diagnosis of drug resistance and the assessment of outcome are needed urgently as a prerequisite for standardized evaluation and reporting of individual therapeutic responses in canine epilepsy. Thus, we provide a proposal for the definition of drug resistance and partial therapeutic success in canine patients with epilepsy. This consensus statement also suggests a list of factors and aspects of outcome, which should be considered in addition to the impact on seizures. Moreover, these expert recommendations discuss criteria which determine the validity and informative value of a therapeutic trial in an individual patient and also suggest the application of individual outcome criteria. Agreement on common guidelines does not only render a basis for future optimization of individual patient management, but is also a presupposition for the design and implementation of clinical studies with highly standardized inclusion and exclusion criteria. Respective standardization will improve the comparability of findings from different studies and renders an improved basis for multicenter studies. Therefore, this proposal provides an in-depth discussion of the implications of outcome criteria for clinical studies. In particular ethical aspects and the different options for study design and application of individual patient-centered outcome criteria are considered