RESUMO
Multiple peptide resistance factor (MprF) confers resistance to cationic antimicrobial peptides (AMPs) in several pathogens, thereby enabling evasion of the host immune response. The role of MprF in commensals remains, however, uncharacterized. To close this knowledge gap, we used a common gut commensal of animals, Lactiplantibacillus plantarum, and its natural host, the fruit fly Drosophila melanogaster, as an experimental model to investigate the role of MprF in commensal-host interactions. The L. plantarum ΔmprF mutant that we generated exhibited deficiency in the synthesis of lysyl-phosphatidylglycerol (Lys-PG), resulting in increased negative cell surface charge and increased susceptibility to AMPs. Susceptibility to AMPs had no effect on ΔmprF mutant's ability to colonize guts of uninfected flies. However, we observed significantly reduced abundance of the ΔmprF mutant after infection-induced inflammation in the guts of wild-type flies but not flies lacking AMPs. Additionally, we found that the ΔmprF mutant compared to wild-type L. plantarum induces a stronger intestinal immune response in flies due to the increased release of immunostimulatory peptidoglycan fragments, indicating an important role of MprF in promoting host tolerance to commensals. Our further analysis suggests that MprF-mediated lipoteichoic acid modifications are involved in host immunomodulation. Overall, our results demonstrate that MprF, besides its well-characterized role in pathogen immune evasion and virulence, is also an important commensal resilience factor.
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Resilience to short-term perturbations, like inflammation, is a fundamental feature of microbiota, yet the underlying mechanisms of microbiota resilience are incompletely understood. Here, we show that Lactiplantibacillus plantarum, a major Drosophila commensal, stably colonizes the fruit fly gut during infection and is resistant to Drosophila antimicrobial peptides (AMPs). By transposon screening, we identified L. plantarum mutants sensitive to AMPs. These mutants were impaired in peptidoglycan O-acetylation or teichoic acid D-alanylation, resulting in increased negative cell surface charge and higher affinity to cationic AMPs. AMP-sensitive mutants were cleared from the gut after infection and aging-induced gut inflammation in wild-type, but not in AMP-deficient flies, suggesting that resistance to host AMPs is essential for commensal resilience in an inflamed gut environment. Thus, our work reveals that in addition to the host immune tolerance to the microbiota, commensal-encoded resilience mechanisms are necessary to maintain the stable association between host and microbiota during inflammation.
Assuntos
Peptídeos Antimicrobianos , Drosophila , Animais , Peptídeos Catiônicos Antimicrobianos/genética , Envelhecimento , InflamaçãoRESUMO
Mouse guanylate-binding proteins (mGBPs) are recruited to various invasive pathogens, thereby conferring cell-autonomous immunity against these pathogens. However, whether and how human GBPs (hGBPs) target M. tuberculosis (Mtb) and L. monocytogenes (Lm) remains unclear. Here, we describe hGBPs association with intracellular Mtb and Lm, which was dependent on the ability of bacteria to induce disruption of phagosomal membranes. hGBP1 formed puncta structures which were recruited to ruptured endolysosomes. Furthermore, both GTP-binding and isoprenylation of hGBP1 were required for its puncta formation. hGBP1 was required for the recovery of endolysosomal integrity. In vitro lipid-binding assays demonstrated direct binding of hGBP1 to PI4P. Upon endolysosomal damage, hGBP1 was targeted to PI4P and PI(3,4)P2-positive endolysosomes in cells. Finally, live-cell imaging demonstrated that hGBP1 was recruited to damaged endolysosomes, and consequently mediated endolysosomal repair. In summary, we uncover a novel interferon-inducible mechanism in which hGBP1 contributes to the repair of damaged phagosomes/endolysosomes.
Assuntos
Proteínas de Ligação ao GTP , Fagossomos , Humanos , Animais , Camundongos , Proteínas de Ligação ao GTP/metabolismo , Fagossomos/metabolismo , Interferons/metabolismo , Endossomos/metabolismoRESUMO
BACKGROUND & AIMS: The homeostasis of the gastrointestinal epithelium relies on cell regeneration and differentiation into distinct lineages organized inside glands and crypts. Regeneration depends on Wnt/ß-catenin pathway activation, but to understand homeostasis and its dysregulation in disease, we need to identify the signaling microenvironment governing cell differentiation. By using gastric glands as a model, we have identified the signals inducing differentiation of surface mucus-, zymogen-, and gastric acid-producing cells. METHODS: We generated mucosoid cultures from the human stomach and exposed them to different growth factors to obtain cells with features of differentiated foveolar, chief, and parietal cells. We localized the source of the growth factors in the tissue of origin. RESULTS: We show that epidermal growth factor is the major fate determinant distinguishing the surface and inner part of human gastric glands. In combination with bone morphogenetic factor/Noggin signals, epidermal growth factor controls the differentiation of foveolar cells vs parietal or chief cells. We also show that epidermal growth factor is likely to underlie alteration of the gastric mucosa in the precancerous condition atrophic gastritis. CONCLUSIONS: Use of our recently established mucosoid cultures in combination with analysis of the tissue of origin provided a robust strategy to understand differentiation and patterning of human tissue and allowed us to draw a new, detailed map of the signaling microenvironment in the human gastric glands.
Assuntos
Padronização Corporal/efeitos dos fármacos , Proteína Morfogenética Óssea 4/farmacologia , Diferenciação Celular/efeitos dos fármacos , Fator de Crescimento Epidérmico/farmacologia , Células Epiteliais/efeitos dos fármacos , Mucosa Gástrica/efeitos dos fármacos , Proteínas de Transporte/farmacologia , Linhagem da Célula , Células Cultivadas , Microambiente Celular , Celulas Principais Gástricas/efeitos dos fármacos , Celulas Principais Gástricas/metabolismo , Celulas Principais Gástricas/ultraestrutura , Células Epiteliais/metabolismo , Células Epiteliais/ultraestrutura , Mucosa Gástrica/metabolismo , Mucosa Gástrica/ultraestrutura , Gastrite Atrófica/metabolismo , Gastrite Atrófica/patologia , Regulação da Expressão Gênica no Desenvolvimento , Humanos , Organoides , Células Parietais Gástricas/efeitos dos fármacos , Células Parietais Gástricas/metabolismo , Células Parietais Gástricas/ultraestrutura , Via de Sinalização WntRESUMO
Retraction: Emoto, M., Emoto, Y., Yoshizawa, I., Kita, E., Shimizu, T., Hurwitz, R., Brinkmann, V. and Kaufmann, S.H.E. (2010), α-GalCer ameliorates listeriosis by accelerating infiltration of Gr-1+ cells into the liver. Eur. J. Immunol., 40: 1328-1341. DOI: https://doi.org/10.1002/eji.200939594 The above article, published online on 16 February 2010 in Wiley Online Library (wileyonlinelibrary.com), has been retracted by agreement between the Chairman of the Executive Committee of the European Journal of Immunology and Wiley-VCH Verlag GmbH & Co. KGaA. The retraction has been agreed following an investigation carried out by Gunma University (http://www.gunma-u.ac.jp/wp-content/uploads/2017/10/chosakekka29.pdf). The investigation was unable to determine the validity of the images for which Professor Emoto, the article's corresponding author, was responsible. As a result, the journal has made the decision to retract the article.
RESUMO
Mosquito blood cells are immune cells that help control infection by vector-borne pathogens. Despite their importance, little is known about mosquito blood cell biology beyond morphological and functional criteria used for their classification. Here, we combined the power of single-cell RNA sequencing, high-content imaging flow cytometry, and single-molecule RNA hybridization to analyze a subset of blood cells of the malaria mosquito Anopheles gambiae By demonstrating that blood cells express nearly half of the mosquito transcriptome, our dataset represents an unprecedented view into their transcriptional program. Analyses of differentially expressed genes identified transcriptional signatures of two cell types and provide insights into the current classification of these cells. We further demonstrate the active transfer of a cellular marker between blood cells that may confound their identification. We propose that cell-to-cell exchange may contribute to cellular diversity and functional plasticity seen across biological systems.
Assuntos
Anopheles/genética , Células Sanguíneas/classificação , Plasticidade Celular/genética , Malária/transmissão , Mosquitos Vetores/genética , Animais , Animais Geneticamente Modificados , Anopheles/imunologia , Células Sanguíneas/imunologia , Comunicação Celular/genética , Conjuntos de Dados como Assunto , Feminino , Genômica/métodos , Mosquitos Vetores/imunologia , RNA/genética , Análise de Sequência de RNA/métodos , Análise de Célula Única/métodos , TranscriptomaRESUMO
Systemic lupus erythematosus (SLE) is an autoimmune disease that has high morbidity and can result in multi-organ damage. SLE is characterized by dysregulated activation of T- and B-lymphocytes and the production of autoantibodies directed against nuclear components. The endonuclease deoxyribonuclease 1 (DNase1) is abundant in blood and a subset of SLE patients have mutations in DNASE1. Furthermore, a report showed that Dnase1-deficient mice develop an SLE-like disease, but these mice also carry a deletion of the gene adjacent to Dnase1, which encodes the chaperone TRAP1/HSP75. We generated a murine strain deficient in Dnase1 with an intact Trap1 gene to examine if a lack of DNase1 is responsible for the development of a spontaneous SLE-like disease. We show that the Dnase1-deficient mice do indeed develop an SLE-like phenotype with elevated autoantibody production by 9 months and kidney damage by 12 months. Notably, this model recapitulates the female bias seen in human SLE patients since female Dnase1-deficient mice produced the highest concentrations of autoantibodies and had more severe kidney damage than males. Since there is currently no cure for SLE the protective role of DNase1 as demonstrated in our study remains of great therapeutic interest.
Assuntos
Desoxirribonuclease I/deficiência , Estudos de Associação Genética , Predisposição Genética para Doença , Lúpus Eritematoso Sistêmico/etiologia , Animais , Autoanticorpos/imunologia , Linfócitos B/imunologia , Linfócitos B/metabolismo , Biomarcadores , Biópsia , Modelos Animais de Doenças , Feminino , Estudos de Associação Genética/métodos , Proteínas de Choque Térmico HSP90/genética , Proteínas de Choque Térmico HSP90/metabolismo , Lúpus Eritematoso Sistêmico/metabolismo , Lúpus Eritematoso Sistêmico/patologia , Nefrite Lúpica/etiologia , Nefrite Lúpica/metabolismo , Nefrite Lúpica/patologia , Masculino , Camundongos , Camundongos Knockout , Fatores SexuaisRESUMO
The aryl hydrocarbon receptor (AhR) is a highly conserved ligand-dependent transcription factor that senses environmental toxins and endogenous ligands, thereby inducing detoxifying enzymes and modulating immune cell differentiation and responses. We hypothesized that AhR evolved to sense not only environmental pollutants but also microbial insults. We characterized bacterial pigmented virulence factors, namely the phenazines from Pseudomonas aeruginosa and the naphthoquinone phthiocol from Mycobacterium tuberculosis, as ligands of AhR. Upon ligand binding, AhR activation leads to virulence factor degradation and regulated cytokine and chemokine production. The relevance of AhR to host defence is underlined by heightened susceptibility of AhR-deficient mice to both P. aeruginosa and M. tuberculosis. Thus, we demonstrate that AhR senses distinct bacterial virulence factors and controls antibacterial responses, supporting a previously unidentified role for AhR as an intracellular pattern recognition receptor, and identify bacterial pigments as a new class of pathogen-associated molecular patterns.
Assuntos
Fatores de Transcrição Hélice-Alça-Hélice Básicos/metabolismo , Mycobacterium tuberculosis/imunologia , Pigmentos Biológicos/metabolismo , Pseudomonas aeruginosa/imunologia , Receptores de Hidrocarboneto Arílico/metabolismo , Receptores de Reconhecimento de Padrão/metabolismo , Animais , Antibacterianos/metabolismo , Células da Medula Óssea/citologia , Citocinas/imunologia , Citocinas/metabolismo , Retroalimentação Fisiológica , Humanos , Ligantes , Ativação de Macrófagos , Camundongos , Mycobacterium tuberculosis/crescimento & desenvolvimento , Mycobacterium tuberculosis/metabolismo , Fenazinas/metabolismo , Pigmentos Biológicos/química , Infecções por Pseudomonas/metabolismo , Pseudomonas aeruginosa/metabolismo , Piocianina/metabolismo , Fatores de Virulência/química , Fatores de Virulência/metabolismoRESUMO
The human gastric pathogen Helicobacter pylori occurs in two basic variants, either exhibiting a functional cagPAI-encoded type-4-secretion-system (T4SS) or not. Only a few cagPAI-positive strains have been successfully adapted for long-term infection of mice, including the pre-mouse Sydney strain 1 (PMSS1). Here we confirm that PMSS1 induces gastric inflammation and neutrophil infiltration in mice, progressing to intestinal metaplasia. Complete genome analysis of PMSS1 revealed 1,423 coding sequences, encompassing the cagPAI gene cluster and, unusually, the location of the cytotoxin-associated gene A (cagA) approximately 15 kb downstream of the island. PMSS1 harbours three genetically exchangeable loci that are occupied by the hopQ coding sequences. HopQ represents a critical co-factor required for the translocation of CagA into the host cell and activation of NF-κB via the T4SS. Long-term colonisation of mice led to an impairment of cagPAI functionality. One of the bacterial clones re-isolated at four months post-infection revealed a mutation in the cagPAI gene cagW, resulting in a frame shift mutation, which prevented CagA translocation, possibly due to an impairment of T4SS function. Rescue of the mutant cagW re-established CagA translocation. Our data reveal intriguing insights into the adaptive abilities of PMSS1, suggesting functional modulation of the H. pylori cagPAI virulence attribute.
Assuntos
Proteínas de Bactérias/genética , Infecções por Helicobacter/microbiologia , Helicobacter pylori/genética , Helicobacter pylori/patogenicidade , Sistemas de Secreção Tipo IV/genética , Animais , Linhagem Celular Tumoral , Feminino , Humanos , Camundongos , Camundongos Endogâmicos C57BL , Família Multigênica , Virulência , Sequenciamento Completo do Genoma/métodosRESUMO
Mycobacterium tuberculosis (Mtb) primarily resides in the lung but can also persist in extrapulmonary sites. Macrophages are considered the prime cellular habitat in all tissues. Here we demonstrate that Mtb resides inside adipocytes of fat tissue where it expresses stress-related genes. Moreover, perigonadal fat of Mtb-infected mice disseminated the infection when transferred to uninfected animals. Adipose tissue harbors leukocytes in addition to adipocytes and other cell types and we observed that Mtb infection induces changes in adipose tissue biology depending on stage of infection. Mice infected via aerosol showed infiltration of inducible nitric oxide synthase (iNOS) or arginase 1 (Arg1)-negative F4/80+ cells, despite recruitment of CD3+, CD4+ and CD8+ T cells. Gene expression analysis of adipose tissue of aerosol Mtb-infected mice provided evidence for upregulated expression of genes associated with T cells and NK cells at 28 days post-infection. Strikingly, IFN-γ-producing NK cells and Mtb-specific CD8+ T cells were identified in perigonadal fat, specifically CD8+CD44-CD69+ and CD8+CD44-CD103+ subpopulations. Gene expression analysis of these cells revealed that they expressed IFN-γ and the lectin-like receptor Klrg1 and down-regulated CD27 and CD62L, consistent with an effector phenotype of Mtb-specific CD8+ T cells. Sorted NK cells expressed higher abundance of Klrg1 upon infection, as well. Our results reveal the ability of Mtb to persist in adipose tissue in a stressed state, and that NK cells and Mtb-specific CD8+ T cells infiltrate infected adipose tissue where they produce IFN-γ and assume an effector phenotype. We conclude that adipose tissue is a potential niche for Mtb and that due to infection CD8+ T cells and NK cells are attracted to this tissue.
Assuntos
Tecido Adiposo/imunologia , Tecido Adiposo/microbiologia , Tuberculose/imunologia , Tuberculose/microbiologia , Latência Viral/imunologia , Adipócitos/microbiologia , Animais , Linfócitos T CD8-Positivos/imunologia , Humanos , Células Matadoras Naturais/imunologia , Camundongos , Mycobacterium tuberculosis/imunologiaRESUMO
The bloodbrain barrier (BBB) and the environment of the central nervous system (CNS) guard the nervous tissue from peripheral immune cells. In the autoimmune disease multiple sclerosis, myelin-reactive T-cell blasts are thought to transgress the BBB and create a pro-inflammatory environment in the CNS, thereby making possible a second autoimmune attack that starts from the leptomeningeal vessels and progresses into the parenchyma. Using a Lewis rat model of experimental autoimmune encephalomyelitis, we show here that contrary to the expectations of this concept, T-cell blasts do not efficiently enter the CNS and are not required to prepare the BBB for immune-cell recruitment. Instead, intravenously transferred T-cell blasts gain the capacity to enter the CNS after residing transiently within the lung tissues. Inside the lung tissues, they move along and within the airways to bronchus-associated lymphoid tissues and lung-draining mediastinal lymph nodes before they enter the blood circulation from where they reach the CNS. Effector T cells transferred directly into the airways showed a similar migratory pattern and retained their full pathogenicity. On their way the T cells fundamentally reprogrammed their gene-expression profile, characterized by downregulation of their activation program and upregulation of cellular locomotion molecules together with chemokine and adhesion receptors. The adhesion receptors include ninjurin 1, which participates in T-cell intravascular crawling on cerebral blood vessels. We detected that the lung constitutes a niche not only for activated T cells but also for resting myelin-reactive memory T cells. After local stimulation in the lung, these cells strongly proliferate and, after assuming migratory properties, enter the CNS and induce paralytic disease. The lung could therefore contribute to the activation of potentially autoaggressive T cells and their transition to a migratory mode as a prerequisite to entering their target tissues and inducing autoimmune disease.
Assuntos
Encéfalo/patologia , Movimento Celular , Encefalomielite Autoimune Experimental/imunologia , Encefalomielite Autoimune Experimental/patologia , Pulmão/patologia , Linfócitos T/patologia , Transferência Adotiva , Animais , Autoimunidade/imunologia , Barreira Hematoencefálica/imunologia , Encéfalo/citologia , Encéfalo/imunologia , Moléculas de Adesão Celular Neuronais/metabolismo , Circulação Cerebrovascular , Modelos Animais de Doenças , Perfilação da Expressão Gênica , Memória Imunológica , Pulmão/citologia , Pulmão/imunologia , Ativação Linfocitária , Bainha de Mielina/imunologia , Fatores de Crescimento Neural/metabolismo , Ratos , Ratos Endogâmicos Lew , Linfócitos T/citologia , Linfócitos T/imunologia , Linfócitos T/metabolismoRESUMO
We show that activation of Wnt/ß-catenin and attenuation of Bmp signals, by combined gain- and loss-of-function mutations of ß-catenin and Bmpr1a, respectively, results in rapidly growing, aggressive squamous cell carcinomas (SCC) in the salivary glands of mice. Tumours contain transplantable and hyperproliferative tumour propagating cells, which can be enriched by fluorescence activated cell sorting (FACS). Single mutations stimulate stem cells, but tumours are not formed. We show that ß-catenin, CBP and Mll promote self-renewal and H3K4 tri-methylation in tumour propagating cells. Blocking ß-catenin-CBP interaction with the small molecule ICG-001 and small-interfering RNAs against ß-catenin, CBP or Mll abrogate hyperproliferation and H3K4 tri-methylation, and induce differentiation of cultured tumour propagating cells into acini-like structures. ICG-001 decreases H3K4me3 at promoters of stem cell-associated genes in vitro and reduces tumour growth in vivo. Remarkably, high Wnt/ß-catenin and low Bmp signalling also characterize human salivary gland SCC and head and neck SCC in general. Our work defines mechanisms by which ß-catenin signals remodel chromatin and control induction and maintenance of tumour propagating cells. Further, it supports new strategies for the therapy of solid tumours.
Assuntos
Carcinoma de Células Escamosas/genética , Carcinoma de Células Escamosas/metabolismo , Histona-Lisina N-Metiltransferase/metabolismo , Proteína de Leucina Linfoide-Mieloide/metabolismo , Neoplasias das Glândulas Salivares/genética , Neoplasias das Glândulas Salivares/metabolismo , Via de Sinalização Wnt , beta Catenina/metabolismo , Animais , Proteínas Morfogenéticas Ósseas/metabolismo , Compostos Bicíclicos Heterocíclicos com Pontes/farmacologia , Proteína de Ligação a CREB/antagonistas & inibidores , Proteína de Ligação a CREB/metabolismo , Carcinoma de Células Escamosas/patologia , Proliferação de Células/efeitos dos fármacos , Epigênese Genética , Neoplasias de Cabeça e Pescoço/genética , Neoplasias de Cabeça e Pescoço/metabolismo , Neoplasias de Cabeça e Pescoço/patologia , Histona Metiltransferases , Humanos , Camundongos , Camundongos Endogâmicos NOD , Camundongos Mutantes , Camundongos SCID , Camundongos Transgênicos , Mutação , Células-Tronco Neoplásicas/metabolismo , Células-Tronco Neoplásicas/patologia , Pirimidinonas/farmacologia , Neoplasias das Glândulas Salivares/patologia , Transplante Heterólogo , Via de Sinalização Wnt/efeitos dos fármacos , beta Catenina/antagonistas & inibidoresRESUMO
For membrane-bound intracellular pathogens, the surrounding vacuole is the portal of communication with the host cell. The parasitophorous vacuole (PV) harboring intrahepatocytic Plasmodium parasites satisfies the parasites' needs of nutrition and protection from host defenses to allow the rapid parasite growth that occurs during the liver stage of infection. In this study, we visualized the PV membrane (PVM) and the associated tubovesicular network (TVN) through fluorescent tagging of two PVM-resident Plasmodium berghei proteins, UIS4 and IBIS1. This strategy revealed previously unrecognized dynamics with which these membranes extend throughout the host cell. We observed dynamic vesicles, elongated clusters of membranes and long tubules that rapidly extend and contract from the PVM in a microtubule-dependent manner. Live microscopy, correlative light-electron microscopy and fluorescent recovery after photobleaching enabled a detailed characterization of these membranous features, including velocities, the distribution of UIS4 and IBIS1, and the connectivity of PVM and TVN. Labeling of host cell compartments revealed association of late endosomes and lysosomes with the elongated membrane clusters. Moreover, the signature host autophagosome protein LC3 was recruited to the PVM and TVN and colocalized with UIS4. Together, our data demonstrate that the membranes surrounding intrahepatic Plasmodium are involved in active remodeling of host cells.
Assuntos
Fígado/parasitologia , Plasmodium/metabolismo , Animais , Membrana Celular/metabolismo , Interações Hospedeiro-Parasita , Plasmodium/patogenicidadeRESUMO
Multiple Sclerosis (MS) is a chronic autoimmune disorder affecting the central nervous system (CNS) through demyelination and neurodegeneration. Until recently, major therapeutic treatments have relied on agents requiring injection delivery. In September 2010, fingolimod/FTY720 (Gilenya, Novartis) was approved as the first oral treatment for relapsing forms of MS. Fingolimod causes down-modulation of S1P1 receptors on lymphocytes which prevents the invasion of autoaggressive T cells into the CNS. In astrocytes, down-modulation of S1P1 by the drug reduces astrogliosis, a hallmark of MS, thereby allowing restoration of productive astrocyte communication with other neural cells and the blood brain barrier. Animal data further suggest that the drug directly supports the recovery of nerve conduction and remyelination. In human MS, such mechanisms may explain the significant decrease in the number of inflammatory markers on brain magnetic resonance imaging in recent clinical trials, and the reduction of brain atrophy by the drug. Fingolimod binds to 4 of the 5 known S1P receptor subtypes, and significant efforts were made over the past 5 years to develop next generation S1P receptor modulators and determine the minimal receptor selectivity needed for maximal therapeutic efficacy in MS patients. Other approaches considered were competitive antagonists of the S1P1 receptor, inhibitors of the S1P lyase to prevent S1P degradation, and anti-S1P antibodies. Below we discuss the current status of the field, and the functional properties of the most advanced compounds. This article is part of a Special Issue entitled New Frontiers in Sphingolipid Biology.
Assuntos
Medicina Clínica , Imunossupressores/farmacologia , Lisofosfolipídeos/metabolismo , Propilenoglicóis/farmacologia , Projetos de Pesquisa , Transdução de Sinais/efeitos dos fármacos , Esfingosina/análogos & derivados , Pesquisa Biomédica , Cloridrato de Fingolimode , Humanos , Receptores de Lisoesfingolipídeo/metabolismo , Esfingosina/metabolismo , Esfingosina/farmacologiaRESUMO
Recently, we showed that endothelial heparan sulfate facilitates entry of a bacterial pathogen into the central nervous system. Here, we show that normal bactericidal activity of neutrophils is influenced by the sulfation pattern of heparan sulfate. Inactivation of heparan sulfate uronyl 2-O-sulfotransferase (Hs2st) in neutrophils substantially reduced their bactericidal activity, and Hs2st deficiency rendered mice more susceptible to systemic infection with the pathogenic bacterium group B Streptococcus. Specifically, altered sulfation of heparan sulfate in mutant neutrophils affected formation of neutrophil extracellular traps while not influencing phagocytosis, production of reactive oxygen species, or secretion of granular proteases. Heparan sulfate proteoglycan(s) is present in neutrophil extracellular traps, modulates histone affinity, and modulates their microbial activity. Hs2st-deficient brain endothelial cells show enhanced binding to group B Streptococcus and are more susceptible to apoptosis, likely contributing to the observed increase in dissemination of group B Streptococcus into the brain of Hs2st-deficient mice following intravenous challenge. Taken together, our data provide strong evidence that heparan sulfate from both neutrophils and the endothelium plays important roles in modulating innate immunity.
Assuntos
Células Endoteliais/imunologia , Proteoglicanas de Heparan Sulfato/imunologia , Imunidade Inata/imunologia , Neutrófilos/imunologia , Animais , Western Blotting , Células Cultivadas , Modelos Animais de Doenças , Armadilhas Extracelulares/imunologia , Proteoglicanas de Heparan Sulfato/metabolismo , Camundongos , Microscopia Eletrônica de Varredura , Infecções Estreptocócicas/imunologia , Streptococcus agalactiae/imunologia , Sulfotransferases/metabolismoRESUMO
Chlamydia trachomatis (Ctr), an obligate intracellular bacterium, survives and replicates within a membrane-bound vacuole, termed the inclusion, which intercepts host exocytic pathways to acquire nutrients. Ctr subverts cellular trafficking pathways from the Golgi by targeting small GTPases, including Rab proteins, to sustain intracellular bacterial replication; however, the precise mechanisms involved remain incompletely understood. Here, we show that Chlamydia infection in human epithelial cells induces microtubule remodeling, in particular the formation of detyrosinated stable MTs, to recruit Golgi ministacks, but not recycling endosomes, to the inclusion. These stable microtubules show increased resistance to chemically induced depolymerization, and their selective depletion results in reduced bacterial infectivity. Rab6 knockdown reversibly prevented not only Golgi ministack formation but also detyrosinated microtubule association with the inclusion. Our data demonstrate that Chlamydia co-opts the function of stable microtubules to support its development.
Assuntos
Infecções por Chlamydia/patologia , Chlamydia trachomatis/crescimento & desenvolvimento , Complexo de Golgi/metabolismo , Microtúbulos/metabolismo , Infecções por Chlamydia/metabolismo , Células HeLa , Células Hep G2 , Humanos , Microtúbulos/patologia , Nocodazol/farmacologia , Moduladores de Tubulina/farmacologia , Proteínas rab de Ligação ao GTP/genética , Proteínas rab de Ligação ao GTP/metabolismoRESUMO
General interest in the biological functions of IFN type I in Mycobacterium tuberculosis (Mtb) infection increased after the recent identification of a distinct IFN gene expression signature in tuberculosis (TB) patients. Here, we demonstrate that TB-susceptible mice lacking the receptor for IFN I (IFNAR1) were protected from death upon aerogenic infection with Mtb. Using this experimental model to mimic primary progressive pulmonary TB, we dissected the immune processes affected by IFN I. IFNAR1 signaling did not affect T-cell responses, but markedly altered migration of inflammatory monocytes and neutrophils to the lung. This process was orchestrated by IFNAR1 expressed on both immune and tissue-resident radioresistant cells. IFNAR1-driven TB susceptibility was initiated by augmented Mtb replication and in situ death events, along with CXCL5/CXCL1-driven accumulation of neutrophils in alveoli, followed by the discrete compartmentalization of Mtb in lung phagocytes. Early depletion of neutrophils rescued TB-susceptible mice to levels observed in mice lacking IFNAR1. We conclude that IFN I alters early innate events at the site of Mtb invasion leading to fatal immunopathology. These data furnish a mechanistic explanation for the detrimental role of IFN I in pulmonary TB and form a basis for understanding the complex roles of IFN I in chronic inflammation.
Assuntos
Interferon Tipo I/imunologia , Pulmão/imunologia , Fagócitos/imunologia , Transdução de Sinais/imunologia , Tuberculose Pulmonar/imunologia , Animais , Células Cultivadas , Quimiocina CXCL1/imunologia , Quimiocina CXCL5/imunologia , Inflamação/imunologia , Camundongos , Camundongos Endogâmicos C57BL , Monócitos/imunologia , Mycobacterium tuberculosis/imunologia , Neutrófilos/imunologia , Alvéolos Pulmonares/imunologia , Receptor de Interferon alfa e beta/imunologia , Linfócitos T/imunologiaRESUMO
The obligate intracellular bacterium Chlamydia trachomatis survives and replicates within a membrane-bound vacuole, termed the inclusion, which intercepts host exocytic pathways to obtain nutrients. Like many other intracellular pathogens, C. trachomatis has a marked requirement for host cell lipids, such as sphingolipids and cholesterol, produced in the endoplasmic reticulum and the Golgi apparatus. However, the mechanisms by which intracellular pathogens acquire host cell lipids are not well understood. In particular, no host cell protein responsible for transporting Golgi-derived lipids to the chlamydial inclusions has yet been identified. Here we show that Chlamydia infection in human epithelial cells induces Golgi fragmentation to generate Golgi ministacks surrounding the bacterial inclusion. Ministack formation is triggered by the proteolytic cleavage of the Golgi matrix protein golgin-84. Inhibition of golgin-84 truncation prevents Golgi fragmentation, causing a block in lipid acquisition and maturation of C. trachomatis. Golgi fragmentation by means of RNA-interference-mediated knockdown of distinct Golgi matrix proteins before infection enhances bacterial maturation. Our data functionally connect bacteria-induced golgin-84 cleavage, Golgi ministack formation, lipid acquisition and intracellular pathogen growth. We show that C. trachomatis subverts the structure and function of an entire host cell organelle for its own advantage.
Assuntos
Chlamydia trachomatis/crescimento & desenvolvimento , Chlamydia trachomatis/patogenicidade , Complexo de Golgi/microbiologia , Complexo de Golgi/patologia , Chlamydia muridarum/crescimento & desenvolvimento , Células Epiteliais/microbiologia , Células Epiteliais/patologia , Técnicas de Silenciamento de Genes , Complexo de Golgi/metabolismo , Proteínas da Matriz do Complexo de Golgi , Células HeLa , Humanos , Metabolismo dos Lipídeos , Proteínas de Membrana/metabolismo , Processamento de Proteína Pós-Traducional , Interferência de RNA , Proteínas de Transporte VesicularRESUMO
RATIONALE: Myeloid cells encompass distinct populations with unique functions during homeostasis and disease. Recently, a novel subset of innate cells, myeloid-derived suppressor cells (MDSCs), has been described in cancer, which suppresses T-cell responses and fosters disease progression. The role of MDSCs in infection is insufficiently addressed. OBJECTIVES: To examine the presence and function of MDSCs during experimental pulmonary tuberculosis (TB) and further understand the immunologic consequences of direct interactions between MDSCs and lung bacterial pathogens. METHODS: Using cell-based approaches and experimental mouse models for pulmonary TB we characterized MDSCs as novel myeloid populations directly interacting with Mycobacterium tuberculosis (Mtb). MEASUREMENTS AND MAIN RESULTS: MDSCs readily phagocytosed Mtb, and released proinflammatory (IL-6, IL-1α) and immunomodulatory (IL-10) cytokines while retaining their suppressive capacity. MDSCs were identified at the site of infection in the lung in disease-resistant and -susceptible mice during pulmonary TB. Excessive MDSC accumulation in lungs correlated with elevated surface expression of IL-4Rα and heightened TB lethality, whereas targeted depletion of MDSCs ameliorated disease. CONCLUSIONS: Our data reveal that MDSCs provide a niche for pathogen survival and tailor immunity in TB. These findings suggest MDSCs as amenable targets for host-directed therapies and emphasize them as cellular-immune regulators during chronic inflammatory conditions, including chronic infections and microbial complications of neoplastic disorders.
Assuntos
Células Mieloides/fisiologia , Tuberculose Pulmonar/imunologia , Animais , Camundongos , Camundongos Endogâmicos C57BLRESUMO
BACKGROUND: The aim of this prospective randomized study was to analyze migration and strain transmission of the Metha™ and Nanos™ femoral prostheses. MATERIALS AND METHODS: Between 1 January 2011 and 2 April 2013, 50 patients were randomized to receive short-stemmed femoral prostheses. Metha™ stems were implanted in 24 patients (12 female, 12 male; mean age 58.7 years; mean body mass index [BMI] 27.4) and Nanos™ stems in 26 patients (10 female, 16 male; mean age 59.7 years; mean BMI 27.1). Longitudinal stem migration, varus-valgus alignment, changes of center of rotation (COR), femoral offset and caput-collum-diaphyseal angle, leg length discrepancy, periprosthetic radiolucent lines incidence, and dual-energy X-ray absorptiometry (DEXA) scans were analysed after an average of 98 and 381 days. RESULTS: There was no significant change of varus-valgus alignment or clinically relevant migration of the Metha™ or Nanos™ prostheses during postoperative follow-up. After 12.3 months, the DEXA scans showed small but significant differences of bone mineral density in Gruen zones 1 (minus ~8 %) and 6 (plus ~9 %) for the Metha™ and in Gruen zone 1 (minus ~14 %) for the Nanos™ (paired t test). Visual analog scale (VAS) and Harris Hip Score (HHS) improved significantly for both implants (Nanos™/Metha™ 12.3 months postoperatively HSS 96.5/96.2; VAS 0.7/0.8, respectively). COR or offset did not change significantly after surgery. CONCLUSIONS: Neither implant showed signs of impaired osseointegration. DEXA demonstrated proximally located load transfer with only moderate proximal stress shielding. LEVEL OF EVIDENCE: II.