Nos2-/- mice infected with M. tuberculosis develop neurobehavioral changes and immunopathology mimicking human central nervous system tuberculosis.

BACKGROUND: Understanding the pathophysiology of central nervous system tuberculosis (CNS-TB) is hampered by the lack of a good pre-clinical model that mirrors the human CNS-TB infection. We developed a murine CNS-TB model that demonstrates neurobehavioral changes with similar immunopathology with human CNS-TB. METHODS: We injected two Mycobacterium tuberculosis (M.tb) strains, H37Rv and CDC1551, respectively, into two mouse strains, C3HeB/FeJ and Nos2-/- mice, either into the third ventricle or intravenous. We compared the neurological symptoms, histopathological changes and levels of adhesion molecules, chemokines, and inflammatory cytokines in the brain induced by the infections through different routes in different strains. RESULTS: Intra-cerebroventricular infection of Nos2-/- mice with M.tb led to development of neurological signs and more severe brain granulomas compared to C3HeB/FeJ mice. Compared with CDC1551 M.tb, H37Rv M.tb infection resulted in a higher neurobehavioral score and earlier mortality. Intra-cerebroventricular infection caused necrotic neutrophil-dominated pyogranulomas in the brain relative to intravenous infection which resulted in disseminated granulomas and mycobacteraemia. Histologically, intra-cerebroventricular infection of Nos2-/- mice with M.tb resembled human CNS-TB brain biopsy specimens. H37Rv intra-cerebroventricular infected mice demonstrated higher brain concentrations of inflammatory cytokines, chemokines and adhesion molecule ICAM-1 than H37Rv intravenous-infected mice. CONCLUSIONS: Intra-cerebroventricular infection of Nos2-/- mice with H37Rv creates a murine CNS-TB model that resembled human CNS-TB immunopathology, exhibiting the worst neurobehavioral score with a high and early mortality reflecting disease severity and its associated neurological morbidity. Our murine CNS-TB model serves as a pre-clinical platform to dissect host-pathogen interactions and evaluate therapeutic agents for CNS-TB.

A Novel In Vitro Mouse Model to Study Mycobacterium tuberculosis Dissemination Across Brain Vessels: A Combination Granuloma and Blood-Brain Barrier Mouse Model.

In vitro culture models of the blood-brain barrier (BBB) provide a useful platform to test the mechanisms of cellular infiltration and pathogen dissemination into the central nervous system (CNS). We present an in vitro mouse model of the BBB to test Mycobacterium tuberculosis (Mtb) dissemination across brain endothelial cells. One-third of the global population is infected with Mtb, and in 1%-2% of cases bacteria invade the CNS through a largely unknown process. The "Trojan horse" theory supports the role of a cellular carrier that engulfs bacteria and carries them to the brain without being recognized. We present for the first time a protocol for an in vitro BBB-granuloma model that supports the Trojan horse mechanism of Mtb dissemination into the CNS. Handling of bacterial cultures, in vivo and in vitro infections, isolation of primary astroglial and endothelial cells, and assembly of the in vitro BBB model is presented. These techniques can be used to analyze the interaction of adaptive and innate immune system cells with brain endothelial cells, cellular transmigration, BBB morphological and functional changes, and methods of bacterial dissemination. © 2020 Wiley Periodicals LLC. Basic Protocol 1: Isolation of primary mouse brain astrocytes and endothelial cells Basic Protocol 2: Isolation of primary mouse bone marrow-derived dendritic cells Support Protocol 1: Validation of dendritic cell purity by flow cytometry Basic Protocol 3: Isolation of primary mouse peripheral blood mononuclear cells Support Protocol 2: Isolation of primary mouse spleen cells Support Protocol 3: Purification and validation of CD4+ T cells from PBMCs and spleen cells Basic Protocol 4: Isolation of liver granuloma supernatant and determination of organ load Support Protocol 4: In vivo and in vitro infection with mycobacteria Basic Protocol 5: Assembly of the BBB co-culture model Basic Protocol 6: Assembly of the combined in vitro granuloma and BBB model.

Molecular and Histologic Diagnosis of Central Nervous System Infections.

Infections of the central nervous system cause significant morbidity and mortality in immunocompetent and immunocompromised individuals. A wide variety of microorganisms can cause infections, including bacteria, mycobacteria, fungi, viruses, and parasites. Although less invasive testing is preferred, surgical biopsy may be necessary to collect diagnostic tissue. Histologic findings, including special stains and immunohistochemistry, can provide a morphologic diagnosis in many cases, which can be further classified by molecular testing. Correlation of molecular, culture, and other laboratory results with histologic findings is essential for an accurate diagnosis, and to minimize false positives from microbial contamination.

Primary Pituitary Tubercular Abscess: A Case Report.

Primary pituitary tubercular abscess is a very rare disease. It may present clinically with visual loss, headache, seizure, hormonal abnormalities or with cranial nerve palsies. MRI is the diagnostic modality and shows a cystic-solid mass in the sellar and suprasellar region, isointense on T1 and T2W images with heterogeneous areas and ring enhancement on contrast. Surgery remains the initial treatment and it is approached through the trans-sphenoidal/trans-nasal or transcranial route followed by anti-tubercular therapy. We report a case of primary pituitary tubercular abscess managed successfully with a brief review of its pathology. Keywords: abscess; pituitary gland; pyogenic; sella; tuberculosis.

Central nervous system tuberculosis reactivation following intravenous iron supplementation.

Patients with inflammatory bowel disease (IBD) are often treated with tumor necrosis factor (TNF)-alpha inhibitors and are therefore at higher risk for tuberculosis (TB) reactivation. IBD patients also frequently have iron deficiency which is often treated with intravenous iron supplementation. Iron plays an important role in mycobacterial infections, and reactivation of TB has been rarely reported after iron repletion. We present a case of a 63-year-old male with a history of Crohn's disease, on treatment with adalimumab for 2 years. The patient presented with malaise, mild lethargy, low-grade fever, and hyponatremia within a week after the first dose of intravenous iron. He was diagnosed with central nervous system TB (positive cerebrospinal fluid polymerase chain reaction and culture) and responded to treatment with a four-drug regimen. The timing of TB reactivation (within a week after intravenous iron administration) suggests that iron repletion contributed to the clinical reactivation of TB. Biological plausibility and prior similar clinical observations further support the causality of this association. Considering the frequency of iron deficiency in IBD, we believe that it is worthy to further explore the potential association between intravenous iron administration and the timing of TB reactivation in patients being treated with TNF-alpha inhibitors.

Tuberculosis of the central nervous system in cattle in Paraíba, Brazil / Tuberculose no sistema nervoso central de bovinos na Paraíba

This paper describes six cases of tuberculosis in the central nervous system (CNS) of cattle in the state of Paraíba in northeastern Brazil. We reviewed the autopsy reports of 851 bovine necropsies performed from 2003 to 2016. Seventy-three (8.6%) cattle were diagnosed with tuberculosis and six showed lesions in the CNS. Three cases affected cattle up to two-year-old and other three affected adults. Three cattle presented exclusively nervous signs, two had respiratory signs and weight loss and one did not present any clinical signs. At necropsy, five cattle had thickening of the leptomeninges of the cerebellum, pons, obex, spinal cord and cortex, mainly, in the region near the brain basilar Willis´ circle. Another animal, presented a single focal lesion in the cerebellum. Microscopically we observed moderate to severe granulomatous meningitis and encephalitis. Five cattle presented lesions in the lungs and mediastinal lymph nodes and three of them had disseminated lesions in other organs. In all cattle acid-fast bacilli were observed in the lesions and marked positive for immunohistochemistry with polyclonal antibody anti-Mycobacterium tuberculosis. It is concluded that bovine tuberculosis of central nervous system occurs sporadically in Paraíba, in cattle of different ages, most of them with disseminate lesions in other organs. The location of the lesions suggests that the agent invaded the brain by hematogenous route through the circle of Willis.(AU)

Descrevem-se seis casos de tuberculose no sistema nervoso central (SNC) em bovinos, no semiárido da Paraíba. Foram revisados os laudos de um total de 851 necropsias de bovinos realizadas no período de 2003 a 2016. Destes, 73 (8,6%) foram diagnosticados com tuberculose e seis apresentavam lesões no SNC. Três casos ocorreram em bovinos de até dois anos de idade e três em bovinos adultos. Três bovinos apresentaram exclusivamente sinais nervosos, dois tinham sinais respiratórios e perda de peso e um não apresentava nenhum sinal clínico. Macroscopicamente, em cinco bovinos, havia espessamento das leptomeninges do cerebelo, medula espinhal, ponte, obex, colículos e córtex, principalmente, na região basilar encefálica próxima ao polígono de Willis. Em apenas um bovino houve a presença de tubérculo único no cerebelo. Microscopicamente observou-se moderada a acentuada meningite e encefalite granulomatosa. Cinco bovinos apresentaram lesões pulmonares e nos gânglios mediastínicos e três deles tinham lesões disseminadas em outros órgãos. Em todos os bovinos foram encontrados bacilos ácido-álcool resistentes intralesionais e todos tiveram marcação positiva na técnica de imuno-histoquímica com anticorpo policlonal anti-Mycobacterium tuberculosis. Conclui-se que a tuberculose do sistema nervoso central de bovinos ocorre de forma esporádica na Paraíba, principalmente em bovinos com lesões disseminadas em outros órgãos. Sugere-se que a disseminação do agente ocorre pela via hematógena, possivelmente através do polígono de Willis.(AU)

Human tuberculosis brain promotes neuronal apoptosis but not in astrocytes with high expression of vascular endothelial growth factor.

The present study aimed to investigate the involvement of the angiogenic marker vascular endothelia growth factor (VEGF) and apoptotic markers of Bcl-2 and Bax in the neurons and astrocytes in the brain infected by Mycobacterium tuberculosis. The immunohistochemistry staining was performed to analyze the expression of the VEGF, Bcl-2 and Bax in the astrocytes and neurons. The expression of VEGF was high in neurons and astrocytes in both the infected brain and control tissues with no difference of angiogenic activity (p = 0.40). Higher Bcl-2 expression was seen in astrocytes of infected brain tissues compared to the control tissues (p = 0.004) promoted a higher anti-apoptotic activity in astrocytes. The neurons expressed strong Bax expression in the infected brain tissues compared to the control tissues (p < 0.001), which indicated more apoptosis in neurons. Thus, neuronal death and survival of infected astrocytes together with high expression of VEGF might be associated with formation of brain tuberculosis. In conclusion, neurons could be more vulnerable than astrocytes in human tuberculosis brain with high expression of VEGF.

Tuberculosis of the central nervous system in children.

BACKGROUND: Central nervous system tuberculosis (CNS TB) in children is still a socioeconomic problem in developing countries. It has varied manifestations, symptoms are nonspecific, diagnosis can be challenging, and treatment may be difficult. It is often missed or overlooked. Among the various pathological entities, tuberculous meningitis is the most common and devastating manifestation. The resultant vasculitis, infarction, and hydrocephalus can be life-threatening. It can have grave cognitive, intellectual, and endocrine sequelae if not treated in time resulting in handicap, especially in resource constraint countries. Early diagnosis and treatment of tuberculous meningitis is the single most important factor determining outcome. Tuberculous hydrocephalus needs to be recognized early, and cerebrospinal fluid diversion procedure needs to be performed in adequate time to prevent morbidity or mortality in some cases. Tuberculous pachymeningitis and arachnoiditis are rare in children. Tuberculous abscess can mimic pyogenic abscess and requires high index of suspicion. Calvarial tuberculosis is seen in children and responds well to antituberculous chemotherapy. Drug-resistant tuberculosis is a formidable problem, and alternate chemotherapy should be promptly instituted. AIM: The pathogenesis, clinical features, diagnosis, and management of central nervous system tuberculosis in children are summarized. CONCLUSION: Heightened clinical suspicion, early diagnosis, appropriate antituberculous treatment, and surgery in relevant situation are essential for a gratifying outcome and preventing complications.

Complex regulation of neutrophil-derived MMP-9 secretion in central nervous system tuberculosis.

BACKGROUND: Central nervous system tuberculosis (CNS-TB) may be fatal even with treatment. Neutrophils are the key mediators of TB immunopathology, and raised CSF matrix metalloproteinase-9 (MMP-9) which correlates to neutrophil count in CNS-TB is associated with neurological deficit and death. The mechanisms by which neutrophils drive TB-associated CNS matrix destruction are not clearly defined. METHODS: Human brain biopsies with histologically proven CNS-TB were stained for neutrophils, neutrophil elastase, and MMP-9. Neutrophil MMP-9 secretion and gene expression were analyzed using Luminex and real-time PCR. Type IV collagen degradation was evaluated using confocal microscopy and quantitative fluorescent assays. Intracellular signaling pathways were investigated by immunoblotting and chemical inhibitors. RESULTS: MMP-9-expressing neutrophils were present in tuberculous granulomas in CNS-TB and neutrophil-derived MMP-9 secretion was upregulated by Mycobacterium tuberculosis (M.tb). Concurrent direct stimulation by M.tb and activation via monocyte-dependent networks had an additive effect on neutrophil MMP-9 secretion. Destruction of type IV collagen, a key component of the blood-brain barrier, was inhibited by neutralizing neutrophil MMP-9. Monocyte-neutrophil networks driving MMP-9 secretion in TB were regulated by MAP-kinase and Akt-PI3 kinase pathways and the transcription factor NF-kB. TNFα neutralization suppressed MMP-9 secretion to baseline while dexamethasone did not. CONCLUSIONS: Multiple signaling paths regulate neutrophil-derived MMP-9 secretion, which is increased in CNS-TB. These paths may be better targets for host-directed therapies than steroids currently used in CNS-TB.

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.

An unusual case of spinal cord restricted mycobacteriosis in a European mink.

Granulomatous myelitis due to infection with Mycobacterium avium was diagnosed in a 4-year-old male neutered European mink (Mustela lutreola). The causative agent was detected by an acid-fast stain and further characterized by polymerase chain reaction and DNA sequencing of the PCR product. A thorough histological evaluation of the remaining organs revealed no granulomatous lesions or detectable acid-fast organisms. Although minks are generally highly susceptible for mycobacteria, localised infections, especially of the central nervous system, are unusual and may represent an atypical chronic form of the disease.

The contribution of a murine CNS-TB model for the understanding of the host-pathogen interactions in the formation of granulomas.

Central nervous system (CNS) tuberculosis (TB) is the most severe form of TB, characterized morphologically by brain granulomas and tuberculous meningitis (TBM). Experimental strategies for the study of the host-pathogen interaction through the analysis of granulomas and its intrinsic molecular mechanisms could provide new insights into the neuropathology of TB. To verify whether cerebellar mycobacterial infection induces the main features of the disease in human CNS and better understand the physiological mechanisms underlying the disease, we injected bacillus Calmette-Guerin (BCG) into the mouse cerebellum. BCG-induced CNS-TB is characterized by the formation of granulomas and TBM, a build up of bacterial loads in these lesions, and microglial recruitment into the lesion sites. In addition, there is an enhanced expression of signaling molecules such as nuclear factor-κB (NF-κB) and there is a presence of inducible nitric oxide synthase (iNOS) in the lesions and surrounding areas. This murine model of cerebellar CNS-TB was characterized by cellular and biochemical immune responses typically found in the human disease. This model could expand our knowledge about granulomas in TB infection of the cerebellum, and help characterize the physiological mechanisms involved with the progression of this serious illness that is responsible for killing millions people every year.

CNS tuberculosis: a review and illustration from an autopsy case.

An estimated one-third of the world's population (2 billion people) is infected with the tubercle bacilli (TB), which is estimated to cause 6% of all deaths worldwide. Despite there being a decline in the incidence of tuberculosis seen in Europe, there are still some countries in the rest of the world where the estimated number of new cases is very high. When a person presents with persistent fever with or without neurological symptoms, the diagnosis of TB cannot be excluded. We present a case report of a 26-year-old male patient, who died of CNS tuberculosis. Such case studies will help keep neuroscience nurses alert to potential medical issues in multiethnic patient populations.

[Multiple intracranial lesions: a clinicalpathologic study of 62 cases].

OBJECTIVE: To study the clinicalpathologic features of intracranial multiple lesions. METHODS: The clinical, radiologic and pathologic features of intracranial multiple lesions in 62 cases during the period from 2005 to 2009 in Xuanwu Hospital were retrospectively reviewed. RESULTS: There were 32 males and 30 females in 62 cases. The mean age of seize onset and duration of disease were 37.4-year-old and 11.6 months, respectively. The lesions could affect cerebral hemisphere, basal ganglia, brain stem, cerebellum and other parts, most lesions were located above the tentorium. Pathological diagnosis as follows: 13 patients with glioma; metastatic tumors in 13 cases; 12 cases of central nervous system infection; immune-mediated inflammatory demyelinating disease in 8 cases; 5 cases of primary lymphoma of central nervous system; primary angiitis of the central nervous system 3 cases; mitochondrial encephalopathy 2 cases; vein thrombosis in 2 cases; Rosai-Dorfman disease in 2 cases; 2 case of radiation encephalopathy. Among them, mitochondrial encephalopathy and vein thrombosis lesions located in the cortex; metastatic tumor and blood-borne infection mainly involving junction of grey and white matter; glioma, radiation encephalopathy and demyelinating disease include white matter lesions; vascular inflammation showed cortical and subcortical white matter lesions. CONCLUSIONS: A variety of tumor and non-neoplastic diseases can be expressed in intracranial multiple lesions, which gliomas, metastatic tumor and central nervous system infections are more common. In order to improve the diagnosis of intracranial multiple lesions, active work in the brian biopsy, study the clinical, imaging and pathological findings must be closely.

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.

Tuberculous encephalopathy without meningitis: pathology and brain MRI findings.

Tuberculous encephalopathy (TBE) is an established disease entity of diffuse cerebral damage occurring with tuberculosis and an underlying immune pathogenesis. However, the presence of this disease entity remains controversial. We report a 15-year-old boy with seizures and a progressive decline of cognitive function. Brain MRI showed diffuse, hyperintense lesions in the white matter on a T2-weighted image, with gadolinium enhancement on a T1-weighted image. Brain biopsy revealed demyelination and granuloma in the white matter. Ziehl-Neelsen staining showed acid-fast bacilli in the granulomas. Antituberculous medication with concomitant steroid treatment resulted in radiological resolution in addition to clinical improvement. Clinicopathological evidence in this case provides additional convincing evidence of TBE as a disease entity distinct from tuberculous meningitis.

Acanthamoeba meningoencephalitis in an immunocompetent patient: an autopsy case report.

Chronic granulomatous CNS infections may be caused by tuberculosis, fungi and rarely by free-living amoeba, especially in immunocompromised individuals. We report a rare, fatal case of granulomatous amoebic encephalitis in an immunocompetent patient mimicking CNS tuberculosis, and review the imageological features and diagnostic tests.

Extrapulmonary tuberculosis in Kabul, Afghanistan: a hospital-based retrospective review.

OBJECTIVES: The purpose of this study is to amplify the knowledge base of the epidemiology, symptoms, and signs of extrapulmonary tuberculosis (EPTB) in Afghanistan. METHODS: This is a retrospective review of EPTB diagnosed at CURE International Hospital and CURE Family Health Center (FHC) in Kabul, Afghanistan during a recent 20-month period. RESULTS: One hundred eighteen cases were identified from patients presenting to the hospital and FHC. This group represents the spectrum of EPTB seen at a single referral center in Kabul. The ratio of females to males was 2.03:1. Lymph node tuberculosis comprised the greatest number of EPTB cases (37.3%, n=44). The central nervous system was the next most frequent site of EPTB involvement (20.3%, n=24), followed in descending order by skeletal, pleural, abdominal, cutaneous, genitourinary, pericardial, miliary, and breast tuberculosis. CONCLUSIONS: The 2:1 ratio of female to male EPTB cases coincides with the unusual epidemiologic pattern seen in smear-positive pulmonary TB in Afghanistan. As the first epidemiological report of EPTB from Afghanistan, this study illustrates the varied presentations of EPTB that should be known by healthcare workers throughout the country.