Mycobacterium tuberculosis (Mtb) infection is initiated in the distal airways, but the bacteria ultimately disseminate to the lung interstitium. Although various cell types, including alveolar ...macrophages (AM), neutrophils, and permissive monocytes, are known to be infected with Mtb, the initially infected cells as well as those that mediate dissemination from the alveoli to the lung interstitium are unknown. In this study, using a murine infection model, we reveal that early, productive Mtb infection occurs almost exclusively within airway-resident AM. Thereafter Mtb-infected, but not uninfected, AM localize to the lung interstitium through mechanisms requiring an intact Mtb ESX-1 secretion system. Relocalization of infected AM precedes Mtb uptake by recruited monocyte-derived macrophages and neutrophils. This dissemination process is driven by non-hematopoietic host MyD88/interleukin-1 receptor inflammasome signaling. Thus, interleukin-1-mediated crosstalk between Mtb-infected AM and non-hematopoietic cells promotes pulmonary Mtb infection by enabling infected cells to disseminate from the alveoli to the lung interstitium.
Display omitted
•Early M. tuberculosis infection primarily targets lung alveolar macrophages (AM)•Mtb-infected AM selectively relocalize from the airways to the lung interstitium•AM interstitial localization precedes Mtb dissemination to other immune cell types•Relocalization of infected AM is dependent on Mtb ESX-1 and host IL-1R signaling
Using a mouse model, Cohen et al. demonstrate that early Mycobacterium tuberculosis infection predominantly targets alveolar macrophages (AM). Infected AM relocalize from the alveolar space to lung interstitium, preceding bacterial dissemination into migratory myeloid populations. Relocalization requires IL-1R and bacterial ESX-1, highlighting the host-pathogen interplay required to establish infection.
Progress in tuberculosis vaccine development is hampered by an incomplete understanding of the immune mechanisms that protect against infection with Mycobacterium tuberculosis (Mtb), the causative ...agent of tuberculosis. Although the M72/ASOE1 trial yielded encouraging results (54% efficacy in subjects with prior exposure to Mtb), a highly effective vaccine against adult tuberculosis remains elusive. We show that in a mouse model, establishment of a contained and persistent yet non-pathogenic infection with Mtb ("contained Mtb infection", CMTB) rapidly and durably reduces tuberculosis disease burden after re-exposure through aerosol challenge. Protection is associated with elevated activation of alveolar macrophages, the first cells that respond to inhaled Mtb, and accelerated recruitment of Mtb-specific T cells to the lung parenchyma. Systems approaches, as well as ex vivo functional assays and in vivo infection experiments, demonstrate that CMTB reconfigures tissue resident alveolar macrophages via low grade interferon-gamma exposure. These studies demonstrate that under certain circumstances, the continuous interaction of the immune system with Mtb is beneficial to the host by maintaining elevated innate immune responses.
Celotno besedilo
Dostopno za:
DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
Tuberculosis (TB) is a heterogeneous disease manifesting in a subset of individuals infected with aerosolized Mycobacterium tuberculosis (Mtb). Unlike human TB, murine infection results in uniformly ...high lung bacterial burdens and poorly organized granulomas. To develop a TB model that more closely resembles human disease, we infected mice with an ultra-low dose (ULD) of between 1–3 founding bacteria, reflecting a physiologic inoculum. ULD-infected mice exhibited highly heterogeneous bacterial burdens, well-circumscribed granulomas that shared features with human granulomas, and prolonged Mtb containment with unilateral pulmonary infection in some mice. We identified blood RNA signatures in mice infected with an ULD or a conventional Mtb dose (50–100 CFU) that correlated with lung bacterial burdens and predicted Mtb infection outcomes across species, including risk of progression to active TB in humans. Overall, these findings highlight the potential of the murine TB model and show that ULD infection recapitulates key features of human TB.
Display omitted
•Ultra-low dose (ULD) Mtb infection of mice results in heterogeneous bacterial burdens•ULD-infected mice often maintain unilateral pulmonary infection•ULD-infected mice develop single, well-organized, pulmonary granuloma•Blood RNA signatures reflecting murine Mtb burdens can predict human TB risk
Conventional murine Mtb infection fails to recapitulate several human TB features. Plumlee et al. show that ultra-low dose infection results in heterogeneous outcomes, including single, well-organized granulomas and unilateral lung containment. Blood transcriptional signatures derived to correlate with pulmonary bacterial burdens in Mtb-infected mice can predict human TB risk.
Growing evidence suggests the outcome of
infection is established rapidly after exposure, but how the current tuberculosis vaccine, bacillus Calmette-Guérin (BCG), impacts early immunity is poorly ...understood. In this study, we found that murine BCG immunization promotes a dramatic shift in infected cell types. Although alveolar macrophages are the major infected cell for the first 2 weeks in unimmunized animals, BCG promotes the accelerated recruitment and infection of lung-infiltrating phagocytes. Interestingly, this shift is dependent on CD4 T cells, yet does not require intrinsic recognition of Ag presented by infected alveolar macrophages.
-specific T cells are first activated in lung regions devoid of infected cells, and these events precede vaccine-induced reduction of the bacterial burden, which occurs only after the colocalization of T cells and infected cells. Understanding how BCG alters early immune responses to
provides new avenues to improve upon the immunity it confers.
Venoms of the sicariid spiders contain phospholipase D enzyme toxins that can cause severe dermonecrosis and even death in humans. These enzymes convert sphingolipid and lysolipid substrates to ...cyclic phosphates by activating a hydroxyl nucleophile present in both classes of lipid. The most medically relevant substrates are thought to be sphingomyelin and/or lysophosphatidylcholine. To better understand the substrate preference of these toxins, we used 31P NMR to compare the activity of three related but phylogenetically diverse sicariid toxins against a diverse panel of sphingolipid and lysolipid substrates. Two of the three showed significantly faster turnover of sphingolipids over lysolipids, and all three showed a strong preference for positively charged (choline and/or ethanolamine) over neutral (glycerol and serine) headgroups. Strikingly, however, the enzymes vary widely in their preference for choline, the headgroup of both sphingomyelin and lysophosphatidylcholine, versus ethanolamine. An enzyme from Sicarius terrosus showed a strong preference for ethanolamine over choline, whereas two paralogous enzymes from Loxosceles arizonica either preferred choline or showed no significant preference. Intrigued by the novel substrate preference of the Sicarius enzyme, we solved its crystal structure at 2.1 Å resolution. The evolution of variable substrate specificity may help explain the reduced dermonecrotic potential of some natural toxin variants, because mammalian sphingolipids use primarily choline as a positively charged headgroup; it may also be relevant for sicariid predatory behavior, because ethanolamine-containing sphingolipids are common in insect prey.
Background: Phospholipase D toxins from brown spider venoms can cause disease in humans.
Results: Different toxin family members show specificity for lipid substrates with choline or ethanolamine headgroups or can be ambiguous.
Conclusion: Spider phospholipase D toxins have evolved diverse substrate preferences.
Significance: The diverse substrate preference may be significant for predation and the mammalian toxicity of venom.
To investigate how host and pathogen diversity govern immunity against Mycobacterium tuberculosis (Mtb), we performed a large-scale screen of vaccine-mediated protection against aerosol Mtb infection ...using three inbred mouse strains C57BL/6 (B6), C3HeB/FeJ (C3H), Balb/c x 129/SvJ (C129F1) and three Mtb strains (H37Rv, CDC1551, SA161) representing two lineages and distinct virulence properties. We compared three protective modalities, all of which involve inoculation with live mycobacteria: Bacillus Calmette-Guérin (BCG), the only approved TB vaccine, delivered either subcutaneously or intravenously, and concomitant Mtb infection (CoMtb), a model of pre-existing immunity in which a low-level Mtb infection is established in the cervical lymph node following intradermal inoculation. We examined lung bacterial burdens at early (Day 28) and late (Day 98) time points after aerosol Mtb challenge and histopathology at Day 98. We observed substantial heterogeneity in the reduction of bacterial load afforded by these modalities at Day 28 across the combinations and noted a strong positive correlation between bacterial burden in unvaccinated mice and the degree of protection afforded by vaccination. Although we observed variation in the degree of reduction in bacterial burdens across the nine mouse/bacterium strain combinations, virtually all protective modalities performed similarly for a given strain-strain combination. We also noted dramatic variation in histopathology changes driven by both host and bacterial genetic backgrounds. Vaccination improved pathology scores for all infections except CDC1551. However, the most dramatic impact of vaccination on lesion development occurred for the C3H-SA161 combination, where vaccination entirely abrogated the development of the large necrotic lesions that arise in unvaccinated mice. In conclusion, we find that substantial TB heterogeneity can be recapitulated by introducing variability in both host and bacterial genetics, resulting in changes in vaccine-mediated protection as measured both by bacterial burden as well as histopathology. These differences can be harnessed in future studies to identify immune correlates of vaccine efficacy.
As early endosomes mature, the SAND-1/CCZ-1 complex acts as a guanine nucleotide exchange factor (GEF) for RAB-7 to promote the activity of its effector, HOPS, which facilitates late ...endosome-lysosome fusion and the consumption of AP-3-containing vesicles. We show that CCZ-1 and the HOPS complex are essential for the biogenesis of gut granules, cell type-specific, lysosome-related organelles (LROs) that coexist with conventional lysosomes in Caenorhabditis elegans intestinal cells. The HOPS subunit VPS-18 promotes the trafficking of gut granule proteins away from lysosomes and functions downstream of or in parallel to the AP-3 adaptor. CCZ-1 also acts independently of AP-3, and ccz-1 mutants mistraffic gut granule proteins. Our results indicate that SAND-1 does not participate in the formation of gut granules. In the absence of RAB-7 activity, gut granules are generated; however, their size and protein composition are subtly altered. These observations suggest that CCZ-1 acts in partnership with a protein other than SAND-1 as a GEF for an alternate Rab to promote gut granule biogenesis. Point mutations in GLO-1, a Rab32/38-related protein, predicted to increase spontaneous guanine nucleotide exchange, specifically suppress the loss of gut granules by ccz-1 and glo-3 mutants. GLO-3 is known to be required for gut granule formation and has homology to SAND-1/Mon1-related proteins, suggesting that CCZ-1 functions with GLO-3 upstream of the GLO-1 Rab, possibly as a GLO-1 GEF. These results support LRO formation occurring via processes similar to conventional lysosome biogenesis, albeit with key molecular differences.
Abstract
During Mycobacterium tuberculosis (Mtb) infection, bacteria are inhaled into the lung where they first infect resident, alveolar macrophages (AM). The majority of the infection remains in AM ...until D14, when other cells, including recruited monocyte-derived macrophages (MDM) begin to become infected. By the peak of infection (D28) MDM represent the major infected cell type. However, even at this time point, a small population of infected AM remain. This led us to ask how these cell types control Mtb. Using microscopy, we observed that AM harbor more bacteria than MDM on a per cell basis at multiple time points, including after the initiation of the T cell response. Using RNA-seq, we identified multiple differentially expressed pathways between the two cell types. While infected MDM upregulate proinflammatory signaling pathways associated with Mtb control, infected AM are enriched for proliferation and fatty acid metabolism pathways. We performed validation studies using dyes to track cell division and mitochondrial metabolism and observed that AM were more proliferative and had sustained engagement of mitochondrial metabolism relative to MDM. In parallel, we analyzed the bacterial transcriptome and found that Mtb in MDM have a signature associated with late hypoxia. These bacterial responses also suggested that Mtb may differ in its drug tolerance in a cell type specific manner, which we are currently investigating. Together, this work is in agreement with other studies demonstrating differences in the response to Mtb by tissue resident and recruited macrophages and suggests that these differences may lead to distinct transcriptional changes in the bacteria, which could have important implications for therapeutic interventions.
Abstract
Mycobacterium tuberculosis (Mtb) infection is initiated in the distal airways, a site patrolled by alveolar macrophages (AM), and infection outcomes are governed by early immune events that ...remain poorly understood. We show that Mtb replicates almost exclusively within airway-resident AM during the first week after aerosol exposure. Although AM are typically thought to reside within the alveolar lumen, confocal imaging demonstrated that Mtb-infected AM establish a novel niche within the lung interstitium, where they undergo proliferation and physically associate with recruited monocyte-derived cells (MC). Localization of AM to the interstitium precedes subsequent Mtb uptake by MC and neutrophils and is driven by non-hematopoietic MyD88/IL-1R inflammasome signaling and the Mtb ESX-1 secretion system. The interstitial localization of infected AM occurs in the absence of recruited monocytes and neutrophils, suggesting that this pulmonary niche may result from AM-intrinsic egress from the alveolar space rather than inflammation-driven alveolar consolidation. Comparisons of the transcriptomes of infected AM localized to the airway or interstitium by RNA-sequencing revealed unique transcriptional profiles in the two populations, suggesting that these two AM subsets may be functionally distinct. For example, Mtb-infected AM in the interstitium dramatically upregulated interferon-response genes, lending support to the idea that these cells are in unique lung environments. Thus, crosstalk between Mtb-infected AM and non-hematopoietic cells establishes pulmonary Mtb infection by promoting the translocation of infected cells from the alveoli to lung interstitium, facilitating dissemination to other myeloid subsets.
PDF
