The Use of Murine Infection Models to Investigate the Protective Role of TNF in Central Nervous System Tuberculosis.

Tuberculosis of the central nervous system (CNS-TB) is the most severe form of extra-pulmonary tuberculosis that is often associated with high mortality. Secretion of tumor necrosis factor (TNF) has important protective and immune modulatory functions for immune responses during CNS-TB. Therefore, by combining the approaches of aerosol and intracerebral infection in mice, this chapter describes the methods to investigate the contribution of TNF in protective immunity against CNS-TB infection.

The immunological architecture of granulomatous inflammation in central nervous system tuberculosis.

Of all tuberculosis (TB) cases, 1% affects the central nervous system (CNS), with a mortality rate of up to 60%. Our aim is to fill the 'key gap' in TBM research by analyzing brain specimens in a unique historical cohort of 84 patients, focusing on granuloma formation. We describe three different types: non-necrotizing, necrotizing gummatous, and necrotizing abscess type granuloma. Our hypothesis is that these different types of granuloma are developmental stages of the same pathological process. All types were present in each patient and were mainly localized in the leptomeninges. Intra-parenchymal granulomas were less abundant than the leptomeningeal ones and mainly located close to the cerebrospinal fluid (subpial and subependymal). We found that most of the intraparenchymal granulomas are an extension of leptomeningeal lesions which is the opposite of the classical Rich focus theory. We present a 3D-model to facilitate further understanding of the topographic relation of granulomas with leptomeninges, brain parenchyma and blood vessels. We describe innate and adaptive immune responses during granuloma formation including the cytokine profiles. We emphasize the presence of leptomeningeal B-cell aggregates as tertiary lymphoid structures. Our study forms a basis for further research in neuroinflammation and infectious diseases of the CNS, especially TB.

Correlation between CNS Tuberculosis and the COVID-19 Pandemic: The Neurological and Therapeutic Insights.

The recent outbreak of Severe Acute Respiratory Syndrome-Coronavirus-2 (SARS-CoV-2) from Wuhan, China, was caused by a single-stranded RNA virus which has kept the entire world stranded. The outbreak was first diagnosed with respiratory illness, but recent findings of acute necrotizing hemorrhage of brain, brain encephalopathy, and the presence of the virus in the cerebrospinal fluid (CSF) have unveiled its neuroinvasivness. Various clinical features related to the central nervous system (CNS) and peripheral nervous system (PNS) due to COVID-19 infection are now identified. We demonstrate here an apparent similarity in neurological disorders of COVID-19 with CNS tuberculosis, which suggests that some anti-tubercular drugs may be used as therapeutic agents against COVID-19 infection.

Experimental animal models of central nervous system tuberculosis: A historical review.

Animal models are and will remain valuable tools in medical research because their use enables a deeper understanding of disease development, thus generating important knowledge for developing disease control strategies. Central nervous system tuberculosis (CNS TB) is the most devastating disease in humans. Moreover, as the variability of signs and symptoms delay a timely diagnosis, patients usually arrive at the hospital suffering from late stage disease. Therefore, it is impossible to obtain fresh human tissue for research before an autopsy. Because of these reasons, studies on human CNS TB are limited to case series, pharmacological response reports, and post mortem histopathological studies. Here, we review the contribution of the different animal models to understand the immunopathology of the disease and the host-parasitic relationship, as well as in the development of new strategies of vaccination and to test new drugs for the treatment of CNS TB.

Cerebral tuberculosis in a patient with systemic lupus erythematosus following cyclophosphamide treatment: a case report.

Central nervous system (CNS) tuberculosis (TB) is a rare but catastrophic event in patients with systemic lupus erythematosus (SLE). Here we report a case of cerebral TB in a patient with lupus myocarditis and nephritis, following cyclophosphamide immunosuppression. To our knowledge this is the first reported case of cerebral TB in SLE in a non-endemic country. A 31-year-old female with SLE and a history of regular travel to Kenya presented to our centre with clinical features of acute heart failure. She was diagnosed with severe lupus myocarditis, and a renal biopsy also confirmed lupus nephritis. Prior to admission, she had also had a cough, fever and weight loss and was under investigation for suspected TB infection. She was treated with ivabradine, beta-blockers and diuretics together with methylprednisolone and cyclophosphamide immunosuppression. Subsequent sputum cultures confirmed TB and she was commenced on triple therapy. Despite this, she developed confusion, dizziness, blurred vision and fluctuating consciousness. Magnetic resonance imaging (MRI) and lumbar puncture revealed CNS TB infection resulting in meningitis. This was later complicated by obstructive hydrocephalus due to TB abscesses. A ventriculoperitoneal (VP) shunt was inserted and TB medications were given intravenously (IV) with dexamethasone. Following a prolonged hospital admission, the patient eventually recovered and rituximab treatment was used to control her SLE. TB infection has been associated with SLE flares. It is likely in this case that TB exacerbated a lupus flare and subsequent immunosuppression resulted in mycobacterial dissemination to the CNS. Systemic and CNS features of TB and SLE are difficult to distinguish and their contemporaneous management represents a diagnostic and therapeutic challenge.

Nitric oxide in cerebrospinal fluid of central nervous system tuberculosis: correlations with culture, antibody response, and cell count.

BACKGROUND/AIM: The role of nitric oxide (NO) has been established in infection over the years. NO functions by inhibiting the growth of intracellular pathogens. The present study was undertaken to ascertain the role of NO in central nervous system (CNS) infection by Mycobacterium tuberculosis. MATERIALS AND METHODS: A total of 781 chronic meningitis cerebrospinal fluid (CSF) samples suspected of CNS tuberculosis (TB) were categorized based on M. tuberculosis culture positivity, anti-TB antibody response, and CSF cell count and were analyzed for NO. RESULTS: We found that NO levels were positive in 10.88% of the CSF samples. Positivity for NO was 18%, 11.67%, 13.68%, 9.32%, and 9.66% in the cases with mycobacterial culture positivity, anti-TB antibody positivity, high cell count, low cell count, and zero cell count, respectively. Among the above cell count categories, NO levels were noticed to be elevated in high cell count samples with mononuclear cell predominance. CONCLUSION: This study suggests that NO might play some role in the later stages of tuberculous meningitis. This is the first study to our knowledge in which NO was evaluated in CSF in relation to immune response and the presence of a pathogen with such a large number of subjects.

TNF-dependent regulation and activation of innate immune cells are essential for host protection against cerebral tuberculosis.

BACKGROUND: Tuberculosis (TB) affects one third of the global population, and TB of the central nervous system (CNS-TB) is the most severe form of tuberculosis which often associates with high mortality. The pro-inflammatory cytokine tumour necrosis factor (TNF) plays a critical role in the initial and long-term host immune protection against Mycobacterium tuberculosis (M. tuberculosis) which involves the activation of innate immune cells and structure maintenance of granulomas. However, the contribution of TNF, in particular neuron-derived TNF, in the control of cerebral M. tuberculosis infection and its protective immune responses in the CNS were not clear. METHODS: We generated neuron-specific TNF-deficient (NsTNF(-/-)) mice and compared outcomes of disease against TNF(f/f) control and global TNF(-/-) mice. Mycobacterial burden in brains, lungs and spleens were compared, and cerebral pathology and cellular contributions analysed by microscopy and flow cytometry after M. tuberculosis infection. Activation of innate immune cells was measured by flow cytometry and cell function assessed by cytokine and chemokine quantification using enzyme-linked immunosorbent assay (ELISA). RESULTS: Intracerebral M. tuberculosis infection of TNF(-/-) mice rendered animals highly susceptible, accompanied by uncontrolled bacilli replication and eventual mortality. In contrast, NsTNF(-/-) mice were resistant to infection and presented with a phenotype similar to that in TNF(f/f) control mice. Impaired immunity in TNF(-/-) mice was associated with altered cytokine and chemokine synthesis in the brain and characterised by a reduced number of activated innate immune cells. Brain pathology reflected enhanced inflammation dominated by neutrophil influx. CONCLUSION: Our data show that neuron-derived TNF has a limited role in immune responses, but overall TNF production is necessary for protective immunity against CNS-TB.

Autoimmune encephalitis as differential diagnosis of infectious encephalitis.

PURPOSE OF REVIEW: This review describes the main types of autoimmune encephalitis with special emphasis on those associated with antibodies against neuronal cell surface or synaptic proteins, and the differential diagnosis with infectious encephalitis. RECENT FINDINGS: There is a continuous expansion of the number of cell surface or synaptic proteins that are targets of autoimmunity. The most recently identified include the metabotropic glutamate receptor 5 (mGluR5), dipeptidyl-peptidase-like protein-6 (DPPX), and γ-aminobutyric acid-A receptor (GABAAR). In these and previously known types of autoimmune encephalitis [N-methyl-D-aspartate receptor (NMDAR), α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptor (AMPAR), γ-aminobutyric acid-B receptor (GABABR), leucine-rich glioma inactivated protein 1 (LGI1), contactin-associated protein-like 2 (CASPR2)], the prodromal symptoms or types of presentations often suggest a viral encephalitis. We review here clues that help in the differential diagnosis with infectious encephalitis. Moreover, recent investigations indicate that viral encephalitis (e.g., herpes simplex) can trigger synaptic autoimmunity. In all these disorders, immunotherapy is usually effective. SUMMARY: Autoimmune encephalitis comprises an expanding group of potentially treatable disorders that should be included in the differential diagnosis of any type of encephalitis. VIDEO ABSTRACT: http://links.lww.com/CONR/A25,

Neurons are host cells for Mycobacterium tuberculosis.

Mycobacterium tuberculosis infection of the central nervous system is thought to be initiated once the bacilli have breached the blood brain barrier and are phagocytosed, primarily by microglial cells. In this study, the interactions of M. tuberculosis with neurons in vitro and in vivo were investigated. The data obtained demonstrate that neurons can act as host cells for M. tuberculosis. M. tuberculosis bacilli were internalized by murine neuronal cultured cells in a time-dependent manner after exposure, with superior uptake by HT22 cells compared to Neuro-2a cells (17.7% versus 9.8%). Internalization of M. tuberculosis bacilli by human SK-N-SH cultured neurons suggested the clinical relevance of the findings. Moreover, primary murine hippocampus-derived neuronal cultures could similarly internalize M. tuberculosis. Internalized M. tuberculosis bacilli represented a productive infection with retention of bacterial viability and replicative potential, increasing 2- to 4-fold within 48 h. M. tuberculosis bacillus infection of neurons was confirmed in vivo in the brains of C57BL/6 mice after intracerebral challenge. This study, therefore, demonstrates neurons as potential new target cells for M. tuberculosis within the central nervous system.

Vaccination with recombinant Mycobacterium tuberculosis PknD attenuates bacterial dissemination to the brain in guinea pigs.

BACKGROUND: We have previously identified Mycobacterium tuberculosis PknD to be an important virulence factor required for the pathogenesis of central nervous system (CNS) tuberculosis (TB). Specifically, PknD mediates bacillary invasion of the blood-brain barrier, which can be neutralized by specific antisera, suggesting its potential role as a therapeutic target against TB meningitis. METHODOLOGY/PRINCIPAL FINDINGS: We utilized an aerosol challenge guinea pig model of CNS TB and compared the protective efficacy of recombinant M. tuberculosis PknD subunit protein with that of M. bovis BCG against bacillary dissemination to the brain. BCG vaccination limited the pulmonary bacillary burden after aerosol challenge with virulent M. tuberculosis in guinea pigs and also reduced bacillary dissemination to the brain (P = 0.01). PknD vaccination also offered significant protection against bacterial dissemination to the brain, which was no different from BCG (P>0.24), even though PknD vaccinated animals had almost 100-fold higher pulmonary bacterial burdens. Higher levels of PknD-specific IgG were noted in animals immunized with PknD, but not in BCG-vaccinated or control animals. Furthermore, pre-incubation of M. tuberculosis with sera from PknD-vaccinated animals, but not with sera from BCG-vaccinated or control animals, significantly reduced bacterial invasion in a human blood-brain barrier model (P<0.01). CONCLUSION: Current recommendations for administering BCG at birth are based on protection gained against severe disease, such as TB meningitis, during infancy. We demonstrate that vaccination with recombinant M. tuberculosis PknD subunit offers a novel strategy to protect against TB meningitis, which is equivalent to BCG in a guinea pig model. Moreover, since BCG lacks the PknD sensor, BCG could also be boosted to develop a more effective vaccine against TB meningitis, a devastating disease that disproportionately affects young children.

The dual face of central nervous system tuberculosis: a new Janus Bifrons?

Tuberculosis (TB) is still a common infectious disease in developing countries, but it is also re-emerging in industrialized nations due to the HIV/AIDS pandemic. In addition to bacillary virulence, the host immune response plays a major role in the development of an active disease (either as a primary infection or reactivation) and in controlling the infection. Even though several mechanisms are involved in regulating the human immune response, biological environment seems to be determinant. In this context, the integrated neuro-immune-endocrine system strongly influences TB clinical outcome. One of the most important clinical aspects of TB is shown when the infection locates in the central nervous system (CNS), in which a very different set of immune responses is induced. Herein we review several aspects of the paradoxical immune response triggered during CNS-TB infection, and discuss the implications of this response in the cerebral infection outcome.

Drug-resistant tuberculosis in two children in Greece: report of the first extensively drug-resistant case.

Extensively drug-resistant (XDR) tuberculosis (TB) represents a serious and growing problem in both endemic and non-endemic countries. We describe a 2.5-year-old girl with XDR-pulmonary TB and an 18-month-old boy with pre-XDR-central nervous system TB. Patients received individualized treatment with second-line anti-TB agents based on genotypic and phenotypic drug susceptibility testing results. Both children achieved culture conversion 3 months and 1 month after treatment initiation, respectively. The child with XDR-pulmonary TB showed evidence of cure while treatment adverse events were managed without treatment interruption. The child with pre-XDR-central nervous system TB after 6-month hospitalization with multiple infectious complications had a dismal end due to hepatic insufficiency possibly related to anti-TB treatment. This is the first report of children with pre-XDR and XDR TB in Greece, emphasizing the public health dimensions and management complexity of XDR TB.

Tuberculous cerebellar abscess in immunocompetent individuals.

Tubercular abscess of the brain is a rare form of central nervous system tuberculosis. These lesions usually occur in the supratentorial compartment. They are associated with a state of immune deficiency. We report two immunocompetent individuals with tubercular abscess of the cerebellum and discuss the management of these lesions.

Lymphocyte apoptosis in children with central nervous system tuberculosis: a case control study.

BACKGROUND: Studies of the apoptosis mechanisms involved in the pathogenesis of tuberculosis have suggested that Mycobacterium tuberculosis can actively interfere with the apoptosis of infected cells. In vivo studies have been performed in adult populations but have not focused on this process in children. In the present study, we analyzed spontaneous T lymphocyte (PBT) apoptosis in the peripheral blood of children with central nervous system tuberculosis (CNS TB), before and after chemotherapy, and compared the results with healthy controls. METHODS: A case-control study was conducted from January 2002 to June 2009. It included 18 children with CNS TB and 17 healthy controls. Spontaneous apoptosis of PBTs, including CD4+, CD8+ and CD8+/CD28+ T cells, was evaluated after 24 and 72 h of culture in complete medium, using the Annexin V detection test. Analysis was conducted before and after chemotherapy, and expression of the apoptotic markers CD95 (Fas) and Fas ligand (FasL) was evaluated. RESULTS: Higher percentages of apoptotic T cells and CD4 lymphocytes were isolated from children with acute phase CNS TB than from children in the control group (p < 0.05). This difference significantly decreased after 60 days of specific treatment. In children with CNS TB, high levels of Fas ligand expression were detected in lymphocyte populations, associated with a high percentage of Fas positive cells, before and after treatment. In contrast to the CD4+ apoptosis profile, we did not find any significant difference in total CD8+ cell apoptosis between children with acute phase disease and the control group. However, the percentage of apoptotic CD8+/CD28+ T cells was significantly higher in the children with acute phase disease than in the healthy controls. CONCLUSIONS: Our findings indicate that CNS TB in pediatric patients increases the sensitivity of CD4 and CD8+/CD28+ T cells to apoptosis, suggesting a hypoergic status of this infection. This could play a key role in the immunopathogenesis of this complicated form of TB. Interestingly, specific chemotherapy is able to normalize both apoptosis sensitivity and T-cell activation.

Mycobacterium tuberculosis upregulates microglial matrix metalloproteinase-1 and -3 expression and secretion via NF-kappaB- and Activator Protein-1-dependent monocyte networks.

Inflammatory tissue destruction is central to pathology in CNS tuberculosis (TB). We hypothesized that microglial-derived matrix metalloproteinases (MMPs) have a key role in driving such damage. Analysis of all of the MMPs demonstrated that conditioned medium from Mycobacterium tuberculosis-infected human monocytes (CoMTb) stimulated greater MMP-1, -3, and -9 gene expression in human microglial cells than direct infection. In patients with CNS TB, MMP-1/-3 immunoreactivity was demonstrated in the center of brain granulomas. Concurrently, CoMTb decreased expression of the inhibitors, tissue inhibitor of metalloproteinase-2, -3, and -4. MMP-1/-3 secretion was significantly inhibited by dexamethasone, which reduces mortality in CNS TB. Surface-enhanced laser desorption ionization time-of-flight analysis of CoMTb showed that TNF-alpha and IL-1beta are necessary but not sufficient for upregulating MMP-1 secretion and act synergistically to drive MMP-3 secretion. Chemical inhibition and promoter-reporter analyses showed that NF-kappaB and AP-1 c-Jun/FosB heterodimers regulate CoMTb-induced MMP-1/-3 secretion. Furthermore, NF-kappaB p65 and AP-1 c-Jun subunits were upregulated in biopsy granulomas from patients with cerebral TB. In summary, functionally unopposed, network-dependent microglial MMP-1/-3 gene expression and secretion regulated by NF-kappaB and AP-1 subunits were demonstrated in vitro and, for the first time, in CNS TB patients. Dexamethasone suppression of MMP-1/-3 gene expression provides a novel mechanism explaining the benefit of steroid therapy in these patients.

Murine model to study the invasion and survival of Mycobacterium tuberculosis in the central nervous system.

BACKGROUND: Tuberculosis of the central nervous system (CNS) is a serious, often fatal disease primarily affecting young children. It develops after hematogenous dissemination and subsequent invasion of the CNS by Mycobacterium tuberculosis. The microbial determinants involved in CNS disease are poorly characterized. METHODS: Hematogenously disseminated M. tuberculosis infection was simulated in BALB/c mice by intravenous challenge. Bacteria were recovered using standard culture techniques. Host immune response to M. tuberculosis infection was assessed by histopathological and cytokine profile analysis. By means of a pooled infection with genotypically defined M. tuberculosis mutants, bacterial genes required for invasion or survival were determined in the CNS and lung tissue. RESULTS: M. tuberculosis were detected in whole mouse brains as early as 1 day after intravenous infection and at all time points assessed thereafter. No significant immune response was elicited in the infected brain tissue, compared with extensive inflammation in the infected lung tissue at the same time point. We identified mutants for 5 M. tuberculosis genes (Rv0311, Rv0805, Rv0931c, Rv0986, and MT3280) with CNS-specific phenotypes, absent in lung tissue. CONCLUSIONS: We have identified CNS-specific M. tuberculosis genes involved in the pathogenesis of tuberculosis. Further characterization of these genes will help in understanding the microbial pathogenesis of CNS tuberculosis.

Rapid diagnosis of CNS tuberculosis by a T-cell interferon-gamma release assay on cerebrospinal fluid mononuclear cells.

Central nervous system tuberculosis remains a clinical diagnostic challenge. The ex vivo Mycobacterium tuberculosis-specific enzyme-linked immunospot assay (ELISPOT) is a novel assay for the rapid detection of M. tuberculosis-specific T-lymphocytes in the peripheral blood. However, when performed on peripheral blood, this assay cannot distinguish between active tuberculosis or latent tuberculosis infection. On the assumption that M. tuberculosis-specific T-lymphocytes migrate to sites of infection, we were able to demonstrate high levels of M. tuberculosis-specific cells by ELISPOT in the cerebrospinal fluid of a patient with tuberculous meningitis and intracerebral tuberculoma four weeks before cerebrospinal fluid culture became positive for M. tuberculosis by culture.