Type-I-interferon-responsive microglia shape cortical development and behavior.

Microglia are brain-resident macrophages that shape neural circuit development and are implicated in neurodevelopmental diseases. Multiple microglial transcriptional states have been defined, but their functional significance is unclear. Here, we identify a type I interferon (IFN-I)-responsive microglial state in the developing somatosensory cortex (postnatal day 5) that is actively engulfing whole neurons. This population expands during cortical remodeling induced by partial whisker deprivation. Global or microglial-specific loss of the IFN-I receptor resulted in microglia with phagolysosomal dysfunction and an accumulation of neurons with nuclear DNA damage. IFN-I gain of function increased neuronal engulfment by microglia in both mouse and zebrafish and restricted the accumulation of DNA-damaged neurons. Finally, IFN-I deficiency resulted in excess cortical excitatory neurons and tactile hypersensitivity. These data define a role for neuron-engulfing microglia during a critical window of brain development and reveal homeostatic functions of a canonical antiviral signaling pathway in the brain.

Drivers determining tuberculosis disease screening yield in four European screening programmes: a comparative analysis.

BACKGROUND: The World Health Organization End TB Strategy emphasises screening for early diagnosis of tuberculosis (TB) in high-risk groups, including migrants. We analysed key drivers of TB yield differences in four large migrant TB screening programmes to inform TB control planning and feasibility of a European approach. METHODS: We pooled individual TB screening episode data from Italy, the Netherlands, Sweden and the UK, and analysed predictors and interactions for TB case yield using multivariable logistic regression models. RESULTS: Between 2005 and 2018 in 2 302 260 screening episodes among 2 107 016 migrants to four countries, the programmes identified 1658 TB cases (yield 72.0 (95% CI 68.6-75.6) per 100 000). In logistic regression analysis, we found associations between TB screening yield and age (≥55 years: OR 2.91 (95% CI 2.24-3.78)), being an asylum seeker (OR 3.19 (95% CI 1.03-9.83)) or on a settlement visa (OR 1.78 (95% CI 1.57-2.01)), close TB contact (OR 12.25 (95% CI 11.73-12.79)) and higher TB incidence in the country of origin. We demonstrated interactions between migrant typology and age, as well as country of origin. For asylum seekers, the elevated TB risk remained similar above country of origin incidence thresholds of 100 per 100 000. CONCLUSIONS: Key determinants of TB yield included close contact, increasing age, incidence in country of origin and specific migrant groups, including asylum seekers and refugees. For most migrants such as UK students and workers, TB yield significantly increased with levels of incidence in the country of origin. The high, country of origin-independent TB risk in asylum seekers above a 100 per 100 000 threshold could reflect higher transmission and re-activation risk of migration routes, with implications for selecting populations for TB screening.

Loss of Roundabout Guidance Receptor 2 (Robo2) in Podocytes Protects Adult Mice from Glomerular Injury by Maintaining Podocyte Foot Process Structure.

Roundabout guidance receptor 2 (ROBO2) plays an important role during early kidney development. ROBO2 is expressed in podocytes, inhibits nephrin-induced actin polymerization, down-regulates nonmuscle myosin IIA activity, and destabilizes kidney podocyte adhesion. However, the role of ROBO2 during kidney injury, particularly in mature podocytes, is not known. Herein, we report that loss of ROBO2 in podocytes [Robo2 conditional knockout (cKO) mouse] is protective from glomerular injuries. Ultrastructural analysis reveals that Robo2 cKO mice display less foot process effacement and better-preserved slit-diaphragm density compared with wild-type littermates injured by either protamine sulfate or nephrotoxic serum (NTS). The Robo2 cKO mice also develop less proteinuria after NTS injury. Further studies reveal that ROBO2 expression in podocytes is up-regulated after glomerular injury because its expression levels are higher in the glomeruli of NTS injured mice and passive Heymann membranous nephropathy rats. Moreover, the amount of ROBO2 in the glomeruli is also elevated in patients with membranous nephropathy. Finally, overexpression of ROBO2 in cultured mouse podocytes compromises cell adhesion. Taken together, these findings suggest that kidney injury increases glomerular ROBO2 expression that might compromise podocyte adhesion and, thus, loss of Robo2 in podocytes could protect from glomerular injury by enhancing podocyte adhesion that helps maintain foot process structure. Our findings also suggest that ROBO2 is a therapeutic target for podocyte injury and podocytopathy.

Complement Targets Newborn Retinal Ganglion Cells for Phagocytic Elimination by Microglia.

Microglia play important roles in shaping the developing CNS, and at early stages they have been proposed to regulate progenitor proliferation, differentiation, and neuronal survival. However, these studies reveal contradictory outcomes, highlighting the complexity of these cell-cell interactions. Here, we investigate microglia function during embryonic mouse retina development, where only microglia, progenitors, and neurons are present. In both sexes, we determine that microglia primarily interact with retinal neurons and find that depletion of microglia via conditional KO of the Csf1 receptor results in increased density of retinal ganglion cells (RGCs). Pharmacological inhibition of microglia also results in an increase in RGCs, with no effect on retinal progenitor proliferation, RGC genesis, or apoptosis. We show that microglia in the embryonic retina are enriched for phagocytic markers and observe engulfment of nonapoptotic Brn3-labeled RGCs. We investigate the molecular pathways that can mediate cell engulfment by microglia and find selective downregulation of complement pathway components with microglia inhibition, and further show that C1q protein marks a subset of RGCs in the embryonic retina. KO of complement receptor 3 (CR3; Itgam), which is only expressed by microglia, results in increased RGC density, similar to what we observed after depletion or inhibition of microglia. Thus, our data suggest that microglia regulate neuron elimination in the embryonic mouse retina by complement-mediated phagocytosis of non-apoptotic newborn RGCs.SIGNIFICANCE STATEMENT Microglia are emerging as active and important participants in regulating neuron number in development, during adult neurogenesis, and following stem cell therapies. However, their role in these contexts and the mechanisms involved are not fully defined. Using a well-characterized in vivo system, we provide evidence that microglia regulate neuronal elimination by complement-mediated engulfment of nonapoptotic neurons. This work provides a significant advancement of the field by defining in vivo molecular mechanisms for microglia-mediated cell elimination. Our data add to a growing body of evidence that microglia are essential for proper nervous system development. In addition, we elucidate microglia function in the developing retina, which may shed light on microglia involvement in the context of retinal injury and disease.

Developmental roles of microglia: A window into mechanisms of disease.

Microglia are engineers of the central nervous system (CNS) both in health and disease. In addition to the canonical immunological roles of clearing damaging entities and limiting the spread of toxicity and death, microglia remodel the CNS throughout life. While they have been extensively studied in disease and injury, due to their highly variable functions, their precise role in these contexts still remains uncertain. Over the past decade, we have greatly expanded our understanding of microglial function, including their essential homeostatic roles during development. Here, we review these developmental roles, identify parallels in disease, and speculate whether developmental mechanisms re-emerge in disease and injury. Developmental Dynamics 248:98-117, 2019. © 2018 Wiley Periodicals, Inc.

Complement C3-Targeted Gene Therapy Restricts Onset and Progression of Neurodegeneration in Chronic Mouse Glaucoma.

Dysregulation of the complement system is implicated in neurodegeneration, including human and animal glaucoma. Optic nerve and retinal damage in glaucoma is preceded by local complement upregulation and activation, but whether targeting this early innate immune response could have therapeutic benefit remains undefined. Because complement signals through three pathways that intersect at complement C3 activation, here we targeted this step to restore complement balance in the glaucomatous retina and to determine its contribution to degeneration onset and/or progression. To achieve this, we combined adeno-associated virus retinal gene therapy with the targeted C3 inhibitor CR2-Crry. We show that intravitreal injection of AAV2.CR2-Crry produced sustained Crry overexpression in the retina and reduced deposition of the activation product complement C3d on retinal ganglion cells and the inner retina of DBA/2J mice. This resulted in neuroprotection of retinal ganglion cell axons and somata despite continued intraocular pressure elevation, suggesting a direct restriction of neurodegeneration onset and progression and significant delay to terminal disease stages. Our study uncovers a damaging effect of complement C3 or downstream complement activation in glaucoma, and it establishes AAV2.CR2-Crry as a viable therapeutic strategy to target pathogenic C3-mediated complement activation in the glaucomatous retina.

Reduction in tuberculosis incidence in the UK from 2011 to 2015: a population-based study.

BACKGROUND: Following nearly two decades of increasing tuberculosis in the UK, TB incidence decreased by 32% from 2011 to 2015. Explaining this reduction is crucial to informing ongoing TB control efforts. METHODS: We stratified TB cases notified in the UK and TB cases averted in the UK through pre-entry screening (PES) between 2011 and 2015 by country of birth and time since arrival. We used population estimates and migration data to establish denominators, and calculated incidence rate ratios (IRRs) between 2011 and 2015. We calculated the contribution of changing migrant population sizes, PES and changes in TB rates to the reduction in TB notifications. RESULTS: TB IRRs fell in all non-EU migrant and UK-born populations between 2011 and 2015 (0.61; 95% CI 0.59 to 0.64 and 0.78; 0.73 to 0.83 respectively), with the greatest decrease in recent non-EU migrants (0.54; 0.48 to 0.61). 61.9% of the reduction in TB notifications was attributable to decreases in TB rates, 33.4% to a fall in the number of recent/mid-term non-EU migrants and 11.4% to PES. A small increase in notifications in EU-born migrants offset the reduction by 6.6%. CONCLUSIONS: Large decreases in TB rates in almost all populations accounted for the majority of the reduction in TB notifications, providing evidence of the impact of recent interventions to improve UK TB control. The particularly large decrease in TB rates in recent non-EU migrants provides evidence of the effectiveness of screening interventions that target this population. These findings will inform ongoing improvements to TB control.

Tuberculosis in the European Union and European Economic Area: a survey of national tuberculosis programmes.

How many European Union (EU) and European Economic Area (EEA) countries have national tuberculosis (TB) control plans/strategies, and what are the priority actions/populations and barriers to implementation?In order to answer this question, a survey of EU/EEA national TB programme leads was undertaken.The response rate was 100% (31 countries). 55% of countries reported having a national TB strategy, all of which were in implementation; five countries were preparing a strategy. 74% had a defined organisational TB control structure with central coordination and 19% had a costed programme budget; few organisational structures included patient/civil society representation. The most frequently mentioned priority TB control actions were: reaching vulnerable population groups (80%), screening for active TB in high-risk groups (63%), implementing electronic registries (60%), contact tracing and outbreak investigation (60%), and tackling multidrug-resistant TB (60%). Undocumented migrants were the most commonly (46%) identified priority population. Perceived obstacles to implementation included barriers related to care recipients (lack of TB knowledge, treatment seeking/adherence), care providers (including need for specialist training of nurses and doctors) and health system constraints (funding, communication between healthcare and social care systems).This survey has provided an insight into TB control programmes across the EU/EEA that will inform the development of a TB strategy toolkit for member states.

Tracking and responding to an outbreak of tuberculosis using MIRU-VNTR genotyping and whole genome sequencing as epidemiological tools.

Background: We describe an outbreak that contributed to a near doubling of the incidence of tuberculosis in Southampton, UK. We examine the importance of 24 locus mycobacterial interspersed repetitive unit variable number tandem repeat (MIRU-VNTR) genotyping in its identification and management and the role of whole genome sequencing (WGS) in tracing the spread of the strain. Methods: Outbreak cases were defined as those diagnosed between January and December 2011 with indistinguishable 24 locus-MIRU-VNTR genotypes or, cases linked epidemiologically. A cluster questionnaire was administered by TB nurses to identify contacts and social settings. Results: Overall, 25 patients fulfilled the case definition. No cases with this MIRU-VNTR genotype had been detected in the UK previously. Connections were found between all cases through household contacts or social venues including a football club, Internet cafe and barber's shop. Public health actions included extended contact tracing, venue screening and TB awareness-raising. The outbreak resulted in a high rate of transmission and high incidence of clinical disease among contacts. Conclusions: This outbreak illustrates the value of combining active case-finding with prospective MIRU-VNTR genotyping to identify settings to undertake public health action. In addition WGS revealed that the VNTR-defined cluster was a single outbreak and that active TB transmission not reactivation was responsible for this outbreak in non-UK born individuals.

Recent TB transmission, clustering and predictors of large clusters in London, 2010-2012: results from first 3†years of universal MIRU-VNTR strain typing.

BACKGROUND: The incidence of TB has doubled in the last 20 years in London. A better understanding of risk groups for recent transmission is required to effectively target interventions. We investigated the molecular epidemiological characteristics of TB cases to estimate the proportion of cases due to recent transmission, and identify predictors for belonging to a cluster. METHODS: The study population included all culture-positive TB cases in London residents, notified between January 2010 and December 2012, strain typed using 24-loci multiple interspersed repetitive units-variable number tandem repeats. Multivariable logistic regression analysis was performed to assess the risk factors for clustering using sociodemographic and clinical characteristics of cases and for cluster size based on the characteristics of the first two cases. RESULTS: There were 10 147 cases of which 5728 (57%) were culture confirmed and 4790 isolates (84%) were typed. 2194 (46%) were clustered in 570 clusters, and the estimated proportion attributable to recent transmission was 34%. Clustered cases were more likely to be UK born, have pulmonary TB, a previous diagnosis, a history of substance abuse or alcohol abuse and imprisonment, be of white, Indian, black-African or Caribbean ethnicity. The time between notification of the first two cases was more likely to be <90 days in large clusters. CONCLUSIONS: Up to a third of TB cases in London may be due to recent transmission. Resources should be directed to the timely investigation of clusters involving cases with risk factors, particularly those with a short period between the first two cases, to interrupt onward transmission of TB.

Glial coverage in the optic nerve expands in proportion to optic axon loss in chronic mouse glaucoma.

Within the white matter, axonal loss by neurodegeneration is coupled to glial cell changes in gene expression, structure and function commonly termed gliosis. Recently, we described the highly variable expansion of gliosis alebosco@neuro.utah.edu in degenerative optic nerves from the DBA/2J mouse model of chronic, age-related glaucoma. Here, to estimate and compare the levels of axonal loss with the expansion of glial coverage and axonal degeneration in DBA/2J nerves, we combined semiautomatic axon counts with threshold-based segmentation of total glial/scar areas and degenerative axonal profiles in plastic cross-sections. In nerves ranging from mild to severe degeneration, we found that the progression of axonal dropout is coupled to an increase of gliotic area. We detected a strong correlation between axon loss and the aggregate coverage by glial cells and scar, whereas axon loss did not correlate with the small fraction of degenerating profiles. Nerves with low to medium levels of axon loss displayed moderate glial reactivity, consisting of hypertrophic astrocytes, activated microglia and normal distribution of oligodendrocytes, with minimal reorganization of the tissue architecture. In contrast, nerves with extensive axonal loss showed prevalent rearrangement of the nerve, with loss of axon fascicle territories and enlarged or almost continuous gliotic and scar domains, containing reactive astrocytes, oligodendrocytes and activated microglia. These findings support the value of optic nerve gliotic expansion as a quantitative estimate of optic neuropathy that correlates with axon loss, applicable to grade the severity of optic nerve damage in mouse chronic glaucoma.

Disrupted SOX10 function causes spongiform neurodegeneration in gray tremor mice.

Mice homozygous for the gray tremor (gt) mutation have a pleiotropic phenotype that includes pigmentation defects, megacolon, whole body tremors, sporadic seizures, hypo- and dys-myelination of the central nervous system (CNS) and peripheral nervous system, vacuolation of the CNS, and early death. Vacuolation similar to that caused by prions was originally reported to be transmissible, but subsequent studies showed the inherited disease was not infectious. The gt mutation mapped to distal mouse chromosome 15, to the same region as Sox10, which encodes a transcription factor with essential roles in neural crest survival and differentiation. As dominant mutations in mouse or human SOX10 cause white spotting and intestinal aganglionosis, we screened the Sox10 coding region for mutations in gt/gt DNA. An adenosine to guanine transversion was identified in exon 2 that changes a highly conserved glutamic acid residue in the SOX10 DNA binding domain to glycine. This mutant allele was not seen in wildtype mice, including the related GT/Le strain, and failed to complement a Sox10 null allele. Gene expression analysis revealed significant down-regulation of genes involved in myelin lipid biosynthesis pathways in gt/gt brains. Knockout mice for some of these genes develop CNS vacuolation and/or myelination defects, suggesting that their down-regulation may contribute to these phenotypes in gt mutants and could underlie the neurological phenotypes associated with peripheral demyelinating neuropathy-central dysmyelinating leukodystrophy-Waardenburg syndrome-Hirschsprung disease, caused by mutations in human SOX10.

Raising standards in UK TB control: introducing cohort review.

Cohort review has been used internationally to support tuberculosis (TB) control. We describe its first use in the UK by a London TB service. Improvements were noted in case management and contact tracing, weaknesses identified and important service changes put in place. Key areas of impact were directly observed therapy (DOT) provision (a greater proportion of cases offered DOT, and in response to low uptake resources diverted to create posts responsible for patient-centred DOT delivery), and contact tracing (more contacts per case screened and assessed). Cohort review enables whole system review and improvement. It has subsequently been adopted across the UK.

Ethambutol and visual assessment in England: current practice and recommendations.

BACKGROUND: Standard treatment for tuberculosis (TB) in children and adults includes an initial two-month course of ethambutol, a drug that in rare cases can cause optic neuropathy and irreversible vision loss. There is a lack of clear guidance on what vision assessments are needed before and during treatment with ethambutol, with the Royal College of Ophthalmologists, National Institute for Health and Care Excellence, British National Formulary and British Thoracic Society offering different guidance. We aimed to assess how vision is routinely tested in patients treated with ethambutol in TB services across England. METHODS: An online survey developed by Public Health England was sent to all TB services in England in 2018 to assess current practice and inform the development of best practice recommendations for visual assessment of patients treated with ethambutol for TB. RESULTS: Sixty-six TB professionals from across England responded, a response rate of 54%. The results showed variations in practice, including when to omit ethambutol from treatment, the timing and frequency of visual assessment, the type of visual assessment, referral processes and management of visual changes. CONCLUSION: This national survey highlights the need for clear guidelines on the testing of vision for patients taking ethambutol at recommended doses, before and during treatment. We suggest a pragmatic approach to visual assessment to reduce variation in practice, proposing a stepwise pathway for patients on standard TB treatment for local adaptation.

Type I interferon responsive microglia shape cortical development and behavior.

Microglia are brain resident phagocytes that can engulf synaptic components and extracellular matrix as well as whole neurons. However, whether there are unique molecular mechanisms that regulate these distinct phagocytic states is unknown. Here we define a molecularly distinct microglial subset whose function is to engulf neurons in the developing brain. We transcriptomically identified a cluster of Type I interferon (IFN-I) responsive microglia that expanded 20-fold in the postnatal day 5 somatosensory cortex after partial whisker deprivation, a stressor that accelerates neural circuit remodeling. In situ, IFN-I responsive microglia were highly phagocytic and actively engulfed whole neurons. Conditional deletion of IFN-I signaling (Ifnar1fl/fl) in microglia but not neurons resulted in dysmorphic microglia with stalled phagocytosis and an accumulation of neurons with double strand DNA breaks, a marker of cell stress. Conversely, exogenous IFN-I was sufficient to drive neuronal engulfment by microglia and restrict the accumulation of damaged neurons. IFN-I deficient mice had excess excitatory neurons in the developing somatosensory cortex as well as tactile hypersensitivity to whisker stimulation. These data define a molecular mechanism through which microglia engulf neurons during a critical window of brain development. More broadly, they reveal key homeostatic roles of a canonical antiviral signaling pathway in brain development.

Effectiveness of nationwide programmatic testing and treatment for latent tuberculosis infection in migrants in England: a retrospective, population-based cohort study.

BACKGROUND: In low-incidence countries, tuberculosis mainly affects migrants, mostly resulting from reactivation of latent tuberculosis infection (LTBI) acquired in high-incidence countries before migration. A nationwide primary care-based LTBI testing and treatment programme for migrants from high-incidence countries was therefore established in high tuberculosis incidence areas in England. We aimed to assess the effectiveness of this programme. METHODS: We did a retrospective, population-based cohort study of migrants who registered in primary care between Jan 1, 2011, and Dec 31, 2018, in 55 high-burden areas with programmatic LTBI testing and treatment. Eligible individuals were aged 16-35 years, born in a high-incidence country, and had entered England in the past 5 years. Individuals who tested interferon-γ release assay (IGRA)-negative were advised about symptoms of tuberculosis, whereas those who tested IGRA-positive were clinically assessed to rule out active tuberculosis and offered preventive therapy. The primary outcome was incident tuberculosis notified to the national Enhanced Tuberculosis Surveillance system. FINDINGS: Our cohort comprised 368 097 eligible individuals who had registered in primary care, of whom 37 268 (10·1%) were tested by the programme. 1446 incident cases of tuberculosis were identified: 166 cases in individuals who had IGRA testing (incidence 204 cases [95% CI 176-238] per 100 000 person-years) and 1280 in individuals without IGRA testing (82 cases [77-86] per 100 000 person-years). Overall, in our primary analysis including all diagnosed tuberculosis cases, a time-varying association was identified between LTBI testing and treatment and lower risk of incident tuberculosis (hazard ratio [HR] 0·76 [95% CI 0·63-0·91]) when compared with no testing. In stratified analysis by follow-up period, the intervention was associated with higher risk of tuberculosis diagnosis during the first 6 months of follow-up (9·93 [7·63-12·9) and a lower risk after 6 months (0·57 [0·41-0·79]). IGRA-positive individuals had higher risk of tuberculosis diagnosis than IGRA-negative individuals (31·9 [20·4-49·8]). Of 37 268 migrants who were tested, 6640 (17·8%) were IGRA-positive, of whom 1740 (26·2%) started preventive treatment. LTBI treatment lowered the risk of tuberculosis: of 135 incident cases in the IGRA-positive cohort, seven cases were diagnosed in the treated group (1·87 cases [95% CI 0·89-3·93] per 1000 person-years) and 128 cases were diagnosed in the untreated group (10·9 cases [9·16-12·9] per 1000 person-years; HR 0·14 [95% CI 0·06-0·32]). INTERPRETATION: A low proportion of eligible migrants were tested by the programme and a small proportion of those testing positive started treatment. Despite this, programmatic LTBI testing and treatment of individuals migrating to a low-incidence region is effective at diagnosing active tuberculosis earlier and lowers the long-term risk of progression to tuberculosis. Increasing programme participation and treatment rates for those testing positive could substantially impact national tuberculosis incidence. FUNDING: National Institute for Health Research Health Protection Research Unit in Respiratory Infections.