RESUMO
Combining data from experiments on multispecies studies provides invaluable contributions to the understanding of basic disease mechanisms and pathophysiology of pathogens crossing species boundaries. The task of multispecies gene expression analysis, however, is often challenging given annotation inconsistencies and in cases of small sample sizes due to bias caused by batch effects. In this work we aim to demonstrate that an alternative approach to standard differential expression analysis in single cell RNA-sequencing (scRNA-seq) based on effect size profiles is suitable for the fusion of data from small samples and multiple organisms. The analysis pipeline is based on effect size metric profiles of samples in specific cell clusters. The effect size substitutes standard differentiation analyses based on p-values and profiles identified based on these effect size metrics serve as a tool to link cell type clusters between the studied organisms. The algorithms were tested on published scRNA-seq data sets derived from several species and subsequently validated on own data from human and bovine peripheral blood mononuclear cells stimulated with Mycobacterium tuberculosis. Correlation of the effect size profiles between clusters allowed for the linkage of human and bovine cell types. Moreover, effect size ratios were used to identify differentially regulated genes in control and stimulated samples. The genes identified through effect size profiling were confirmed experimentally using qPCR. We demonstrate that in situations where batch effects dominate cell type variation in single cell small sample size multispecies studies, effect size profiling is a valid alternative to traditional statistical inference techniques.
Assuntos
Mycobacterium tuberculosis , Análise de Célula Única , Análise de Célula Única/métodos , Animais , Humanos , Bovinos , Mycobacterium tuberculosis/genética , Perfilação da Expressão Gênica/métodos , Algoritmos , Leucócitos Mononucleares/metabolismo , Análise de Sequência de RNA/métodosRESUMO
The gastroesophageal squamocolumnar junction (GE-SCJ) is a critical tissue interface between the esophagus and stomach, with significant relevance in the pathophysiology of gastrointestinal diseases. Despite this, the molecular mechanisms underlying GE-SCJ development remain unclear. Using single-cell transcriptomics, organoids, and spatial analysis, we examine the cellular heterogeneity and spatiotemporal dynamics of GE-SCJ development from embryonic to adult mice. We identify distinct transcriptional states and signaling pathways in the epithelial and mesenchymal compartments of the esophagus and stomach during development. Fibroblast-epithelial interactions are mediated by various signaling pathways, including WNT, BMP, TGF-ß, FGF, EGF, and PDGF. Our results suggest that fibroblasts predominantly send FGF and TGF-ß signals to the epithelia, while epithelial cells mainly send PDGF and EGF signals to fibroblasts. We observe differences in the ligands and receptors involved in cell-cell communication between the esophagus and stomach. Our findings provide insights into the molecular mechanisms underlying GE-SCJ development and fibroblast-epithelial crosstalk involved, paving the way to elucidate mechanisms during adaptive metaplasia development and carcinogenesis.
Assuntos
Fator de Crescimento Epidérmico , Junção Esofagogástrica , Animais , Camundongos , Fator de Crescimento Epidérmico/metabolismo , Junção Esofagogástrica/metabolismo , Fator de Crescimento Transformador beta/metabolismo , Fibroblastos/metabolismo , Análise de Célula ÚnicaRESUMO
Colibactin, a bacterial genotoxin produced by E. coli strains harboring the pks genomic island, induces cytopathic effects, such as DNA breaks, cell cycle arrest, and apoptosis. Patients with inflammatory bowel diseases, such as ulcerative colitis, display changes in their microbiota with the expansion of E. coli. Whether and how colibactin affects the integrity of the colonic mucosa and whether pks+ E. coli contributes to the pathogenesis of colitis is not clear. Using a gnotobiotic mouse model, we show that under homeostatic conditions, pks+ E. coli do not directly interact with the epithelium or affect colonic integrity. However, upon short-term chemical disruption of mucosal integrity, pks+ E. coli gain direct access to the epithelium, causing epithelial injury and chronic colitis, while mice colonized with an isogenic ΔclbR mutant incapable of producing colibactin show a rapid recovery. pks+ E. coli colonized mice are unable to reestablish a functional barrier. In turn, pks+ E. coli remains in direct contact with the epithelium, perpetuating the process and triggering chronic mucosal inflammation that morphologically and transcriptionally resembles human ulcerative colitis. This state is characterized by impaired epithelial differentiation and high proliferative activity, which is associated with high levels of stromal R-spondin 3. Genetic overexpression of R-spondin 3 in colon myofibroblasts is sufficient to mimic barrier disruption and expansion of E. coli. Together, our data reveal that pks+ E. coli are pathobionts that promote severe injury and initiate a proinflammatory trajectory upon contact with the colonic epithelium, resulting in a chronic impairment of tissue integrity.
Assuntos
Colite Ulcerativa , Microbioma Gastrointestinal , Policetídeos , Humanos , Camundongos , Animais , Escherichia coli/genética , Escherichia coli/metabolismo , Colite Ulcerativa/patologia , Policetídeos/metabolismo , Mucosa Intestinal/metabolismoRESUMO
The stomach corpus epithelium is organized into anatomical units that consist of glands and pits. Mechanisms that control the cellular organization of corpus glands and enable their recovery upon injury are not well understood. R-spondin 3 (RSPO3) is a WNT-signaling enhancer that regulates stem cell behavior in different organs. Here, we investigated the function of RSPO3 in the corpus during homeostasis, upon chief and/or parietal cell loss, and during chronic Helicobacter pylori infection. Using organoid culture and conditional mouse models, we demonstrate that RSPO3 is a critical driver of secretory cell differentiation in the corpus gland toward parietal and chief cells, while its absence promoted pit cell differentiation. Acute loss of chief and parietal cells induced by high dose tamoxifen - or merely the depletion of LGR5+ chief cells - caused an upregulation of RSPO3 expression, which was required for the initiation of a coordinated regenerative response via the activation of yes-associated protein (YAP) signaling. This response enabled a rapid recovery of the injured secretory gland cells. However, in the context of chronic H. pylori infection, the R-spondin-driven regeneration was maintained long term, promoting severe glandular hyperproliferation and the development of premalignant metaplasia.
Assuntos
Infecções por Helicobacter , Helicobacter pylori , Neoplasias Gástricas , Camundongos , Animais , Helicobacter pylori/metabolismo , Infecções por Helicobacter/metabolismo , Receptores Acoplados a Proteínas G/genética , Receptores Acoplados a Proteínas G/metabolismo , Mucosa Gástrica/metabolismo , Metaplasia/metabolismo , Metaplasia/patologia , Estômago/patologia , Regeneração , Neoplasias Gástricas/metabolismoRESUMO
BACKGROUND: The intestinal microbiota fundamentally guides the development of a normal intestinal physiology, the education, and functioning of the mucosal immune system. The Citrobacter rodentium-carrier model in germ-free (GF) mice is suitable to study the influence of selected microbes on an otherwise blunted immune response in the absence of intestinal commensals. RESULTS: Here, we describe that colonization of adult carrier mice with 14 selected commensal microbes (OMM12 + MC2) was sufficient to reestablish the host immune response to enteric pathogens; this conversion was facilitated by maturation and activation of the intestinal blood vessel system and the step- and timewise stimulation of innate and adaptive immunity. While the immature colon of C. rodentium-infected GF mice did not allow sufficient extravasation of neutrophils into the gut lumen, colonization with OMM12 + MC2 commensals initiated the expansion and activation of the visceral vascular system enabling granulocyte transmigration into the gut lumen for effective pathogen elimination. CONCLUSIONS: Consortium modeling revealed that the addition of two facultative anaerobes to the OMM12 community was essential to further progress the intestinal development. Moreover, this study demonstrates the therapeutic value of a defined consortium to promote intestinal maturation and immunity even in adult organisms. Video Abstract.
Assuntos
Citrobacter rodentium , Mucosa Intestinal , Animais , Citrobacter rodentium/fisiologia , Sistema Imunitário , Imunocompetência , Intestinos , CamundongosRESUMO
Helicobacter pylori is a pathogen that colonizes the stomach and causes chronic gastritis. Helicobacter pylori can colonize deep inside gastric glands, triggering increased R-spondin 3 (Rspo3) signaling. This causes an expansion of the "gland base module," which consists of self-renewing stem cells and antimicrobial secretory cells and results in gland hyperplasia. The contribution of Rspo3 receptors Lgr4 and Lgr5 is not well explored. Here, we identified that Lgr4 regulates Lgr5 expression and is required for H. pylori-induced hyperplasia and inflammation, while Lgr5 alone is not. Using conditional knockout mice, we reveal that R-spondin signaling via Lgr4 drives proliferation of stem cells and also induces NF-κB activity in the proliferative stem cells. Upon exposure to H. pylori, the Lgr4-driven NF-κB activation is responsible for the expansion of the gland base module and simultaneously enables chemokine expression in stem cells, resulting in gland hyperplasia and neutrophil recruitment. This demonstrates a connection between R-spondin-Lgr and NF-κB signaling that links epithelial stem cell behavior and inflammatory responses to gland-invading H. pylori.
Assuntos
Helicobacter pylori , Animais , Hiperplasia/metabolismo , Hiperplasia/patologia , Inflamação/patologia , Camundongos , NF-kappa B/metabolismo , Receptores Acoplados a Proteínas G/genética , Receptores Acoplados a Proteínas G/metabolismo , Células-Tronco/metabolismo , EstômagoRESUMO
Helicobacter pylori causes gastric inflammation, gland hyperplasia and is linked to gastric cancer. Here, we studied the interplay between gastric epithelial stem cells and their stromal niche under homeostasis and upon H. pylori infection. We find that gastric epithelial stem cell differentiation is orchestrated by subsets of stromal cells that either produce BMP inhibitors in the gland base, or BMP ligands at the surface. Exposure to BMP ligands promotes a feed-forward loop by inducing Bmp2 expression in the epithelial cells themselves, enforcing rapid lineage commitment to terminally differentiated mucous pit cells. H. pylori leads to a loss of stromal and epithelial Bmp2 expression and increases expression of BMP inhibitors, promoting self-renewal of stem cells and accumulation of gland base cells, which we mechanistically link to IFN-γ signaling. Mice that lack IFN-γ signaling show no alterations of BMP gradient upon infection, while exposure to IFN-γ resembles H. pylori-driven mucosal responses.
Assuntos
Infecções por Helicobacter , Helicobacter pylori , Animais , Células Epiteliais/metabolismo , Mucosa Gástrica/metabolismo , Infecções por Helicobacter/metabolismo , Inflamação/metabolismo , Ligantes , CamundongosRESUMO
Coinfections with pathogenic microbes continually confront cervical mucosa, yet their implications in pathogenesis remain unclear. Lack of in-vitro models recapitulating cervical epithelium has been a bottleneck to study coinfections. Using patient-derived ectocervical organoids, we systematically modeled individual and coinfection dynamics of Human papillomavirus (HPV)16 E6E7 and Chlamydia, associated with carcinogenesis. The ectocervical stem cells were genetically manipulated to introduce E6E7 oncogenes to mimic HPV16 integration. Organoids from these stem cells develop the characteristics of precancerous lesions while retaining the self-renewal capacity and organize into mature stratified epithelium similar to healthy organoids. HPV16 E6E7 interferes with Chlamydia development and induces persistence. Unique transcriptional and post-translational responses induced by Chlamydia and HPV lead to distinct reprogramming of host cell processes. Strikingly, Chlamydia impedes HPV-induced mechanisms that maintain cellular and genome integrity, including mismatch repair in the stem cells. Together, our study employing organoids demonstrates the hazard of multiple infections and the unique cellular microenvironment they create, potentially contributing to neoplastic progression.
Assuntos
Chlamydia , Coinfecção , Infecções por Papillomavirus , Neoplasias do Colo do Útero , Reprogramação Celular/genética , Feminino , Papillomavirus Humano 16/genética , Humanos , Organoides , Microambiente Tumoral , Neoplasias do Colo do Útero/genéticaRESUMO
Rationale: Suboptimal vaccine immunogenicity and antigenic mismatch, compounded by poor uptake, means that influenza remains a major global disease. T cells recognizing peptides derived from conserved viral proteins could enhance vaccine-induced cross-strain protection. Objectives: To investigate the kinetics, phenotypes, and function of influenza virus-specific CD8+ resident memory T (Trm) cells in the lower airway and infer the molecular pathways associated with their response to infection in vivo. Methods: Healthy volunteers, aged 18-55, were inoculated intranasally with influenza A/California/4/09(H1N1). Blood, upper airway, and (in a subgroup) lower airway samples were obtained throughout infection. Symptoms were assessed by using self-reported diaries, and the nasal viral load was assessed by using quantitative PCR. T-cell responses were analyzed by using a three-color FluoroSpot assay, flow cytometry with MHC I-peptide tetramers, and RNA sequencing, with candidate markers being confirmed by using the immunohistochemistry results for endobronchial biopsy specimens. Measurements and Main Results: After challenge, 57% of participants became infected. Preexisting influenza-specific CD8+ T cells in blood correlated strongly with a reduced viral load, which peaked at Day 3. Influenza-specific CD8+ T cells in BAL fluid were highly enriched and predominantly expressed the Trm markers CD69 and CD103. Comparison between preinfection CD8+ T cells in BAL fluid and blood by using RNA sequencing revealed 3,928 differentially expressed genes, including all major Trm-cell markers. However, gene set enrichment analysis of BAL-fluid CD8+ T cells showed primarily innate cell-related pathways and, during infection, included upregulation of innate chemokines (Cxcl1, Cxcl10, and Cxcl16) that were also expressed by CD8+ cells in bronchial tissues. Conclusions: CD8+ Trm cells in the human lung display innate-like gene and protein expression that demonstrates blurred divisions between innate and adaptive immunity. Clinical study registered with www.clinicaltrials.gov (NCT02755948).
Assuntos
Linfócitos T CD8-Positivos/imunologia , Imunidade Inata/genética , Vírus da Influenza A Subtipo H1N1/imunologia , Influenza Humana/imunologia , Imunidade Adaptativa/genética , Adolescente , Adulto , Antígenos CD/genética , Antígenos CD/metabolismo , Antígenos de Diferenciação de Linfócitos T/genética , Antígenos de Diferenciação de Linfócitos T/metabolismo , Biomarcadores/metabolismo , Líquido da Lavagem Broncoalveolar/imunologia , Líquido da Lavagem Broncoalveolar/virologia , Linfócitos T CD8-Positivos/metabolismo , Quimiocinas/metabolismo , Feminino , Expressão Gênica , Perfilação da Expressão Gênica , Voluntários Saudáveis , Humanos , Influenza Humana/genética , Influenza Humana/virologia , Cadeias alfa de Integrinas/genética , Cadeias alfa de Integrinas/metabolismo , Cinética , Lectinas Tipo C/genética , Lectinas Tipo C/metabolismo , Masculino , Pessoa de Meia-Idade , Fenótipo , Sistema Respiratório/imunologia , Sistema Respiratório/virologia , Carga Viral , Adulto JovemRESUMO
Cellular stress has been associated with inflammation, yet precise underlying mechanisms remain elusive. In this study, various unrelated stress inducers were employed to screen for sensors linking altered cellular homeostasis and inflammation. We identified the intracellular pattern recognition receptors NOD1/2, which sense bacterial peptidoglycans, as general stress sensors detecting perturbations of cellular homeostasis. NOD1/2 activation upon such perturbations required generation of the endogenous metabolite sphingosine-1-phosphate (S1P). Unlike peptidoglycan sensing via the leucine-rich repeats domain, cytosolic S1P directly bound to the nucleotide binding domains of NOD1/2, triggering NF-κB activation and inflammatory responses. In sum, we unveiled a hitherto unknown role of NOD1/2 in surveillance of cellular homeostasis through sensing of the cytosolic metabolite S1P. We propose S1P, an endogenous metabolite, as a novel NOD1/2 activator and NOD1/2 as molecular hubs integrating bacterial and metabolic cues.
Assuntos
Inflamação/metabolismo , Lisofosfolipídeos/metabolismo , Proteína Adaptadora de Sinalização NOD1/metabolismo , Proteína Adaptadora de Sinalização NOD2/metabolismo , Esfingosina/análogos & derivados , Animais , Linhagem Celular , Linhagem Celular Tumoral , Feminino , Células HEK293 , Células HeLa , Humanos , Camundongos , NF-kappa B/metabolismo , Peptidoglicano/metabolismo , Transdução de Sinais/fisiologia , Esfingosina/metabolismo , Células THP-1RESUMO
Colonization of the mosquito host by Plasmodium parasites is achieved by sexually differentiated gametocytes. Gametocytogenesis, gamete formation and fertilization are tightly regulated processes, and translational repression is a major regulatory mechanism for stage conversion. Here, we present a characterization of a Plasmodium berghei RNA binding protein, UIS12, that contains two conserved eukaryotic RNA recognition motifs (RRM). Targeted gene deletion resulted in viable parasites that replicate normally during blood infection, but form fewer gametocytes. Upon transmission to Anopheles stephensi mosquitoes, both numbers and size of midgut-associated oocysts were reduced and their development stopped at an early time point. As a consequence, no salivary gland sporozoites were formed indicative of a complete life cycle arrest in the mosquito vector. Comparative transcript profiling in mutant and wild-type infected red blood cells revealed a decrease in transcript abundance of mRNAs coding for signature gamete-, ookinete-, and oocyst-specific proteins in uis12(-) parasites. Together, our findings indicate multiple roles for UIS12 in regulation of gene expression after blood infection in good agreement with the pleiotropic defects that terminate successful sporogony and onward transmission to a new vertebrate host.
Assuntos
Anopheles , Plasmodium berghei , Animais , Oocistos , Proteínas de Ligação a RNA , EsporozoítosRESUMO
Apart from the constitutive proteasome, the immunoproteasome that comprises the three proteolytic subunits LMP2, MECL-1, and LMP7 is expressed in most immune cells. In this study, we describe opposing roles for immunoproteasomes in regulating the tumor microenvironment (TME). During chronic inflammation, immunoproteasomes modulated the expression of protumorigenic cytokines and chemokines and enhanced infiltration of innate immune cells, thus triggering the onset of colitis-associated carcinogenesis (CAC) in wild-type mice. Consequently, immunoproteasome-deficient animals (LMP2/MECL-1/LMP7-null mice) were almost completely resistant to CAC development. In patients with ulcerative colitis with high risk for CAC, immunoproteasome-induced protumorigenic mediators were upregulated. In melanoma tumors, the role of immunoproteasomes is relatively unknown. We found that high expression of immunoproteasomes in human melanoma was associated with better prognosis. Similarly, our data revealed that the immunoproteasome has antitumorigenic activity in a mouse model of melanoma. The antitumor immunity against melanoma was compromised in immunoproteasome-deficient mice because of the impaired activity of CD8+ CTLs, CD4+ Th1 cells, and antigen-presenting cells. These findings show that immunoproteasomes may exert opposing roles with either pro- or antitumoral properties in a context-dependent manner.
Assuntos
Cisteína Endopeptidases/metabolismo , Melanoma Experimental/imunologia , Complexo de Endopeptidases do Proteassoma/metabolismo , Linfócitos T Citotóxicos/imunologia , Microambiente Tumoral/imunologia , Animais , Antineoplásicos/uso terapêutico , Linhagem Celular Tumoral , Colite/patologia , Cisteína Endopeptidases/deficiência , Cisteína Endopeptidases/genética , Citocinas/metabolismo , Feminino , Antígenos de Histocompatibilidade Classe I/metabolismo , Humanos , Melanoma Experimental/tratamento farmacológico , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Complexo de Endopeptidases do Proteassoma/genética , Linfócitos T Citotóxicos/metabolismoRESUMO
BACKGROUND AND AIMS: A histopathological hallmark of chronic hepatitis B virus (HBV) infection is the presence of ground glass hepatocytes (GGHs). GGHs are liver cells that exhibit eosinophilic, granular, glassy cytoplasm in light microscopy and are characterized by accumulation of HBV surface (HBs) proteins in the endoplasmic reticulum (ER). More important, GGHs have been accepted as a precursor of HCC and may represent preneoplastic lesions of the liver. METHODS: Here we show that the reason for ground glass phenotype of hepatocytes in patients with chronic hepatitis B (CHB) and in HBs transgenic mice is a complex formation between HBs proteins and lipid droplets (LDs) within the ER. RESULTS: As fat is a main component of LDs their presence reduces the protein density of HBs aggregates. Therefore, they adsorb less amount of eosin during hematoxylin-eosin staining and appear dull in light microscopy. However, after induction of interferon response in the liver LDs were not only co-localized with HBs but also distributed throughout the cytoplasm of hepatocytes. The uniform distribution of LDs weakens the contrast between HBs aggregates and the rest of the cytoplasm and complicates the identification of GGHs. Suppression of interferon response restored the ground glass phenotype of hepatocytes. CONCLUSIONS: Complex formation between HBs and LDs represents a very important feature of CHB that could affect LDs functions in hepatocytes. The strain specific activation of the interferon response in the liver of HBs/c mice prevented the development of GGHs. Thus, manipulation of LDs could provide a new treatment strategy in the prevention of liver cancer.
Assuntos
Hepatócitos/metabolismo , Interferons/metabolismo , Metabolismo dos Lipídeos , Animais , Retículo Endoplasmático/metabolismo , Humanos , Fígado/metabolismo , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Modelos Biológicos , FenótipoRESUMO
SARS-CoV-2, the agent that causes COVID-19, invades epithelial cells, including those of the respiratory and gastrointestinal mucosa, using angiotensin-converting enzyme-2 (ACE2) as a receptor. Subsequent inflammation can promote rapid virus clearance, but severe cases of COVID-19 are characterized by an inefficient immune response that fails to clear the infection. Using primary epithelial organoids from human colon, we explored how the central antiviral mediator IFN-γ, which is elevated in COVID-19, affects epithelial cell differentiation, ACE2 expression, and susceptibility to infection with SARS-CoV-2. In mouse and human colon, ACE2 is mainly expressed by surface enterocytes. Inducing enterocyte differentiation in organoid culture resulted in increased ACE2 production. IFN-γ treatment promoted differentiation into mature KRT20+ enterocytes expressing high levels of ACE2, increased susceptibility to SARS-CoV-2 infection, and resulted in enhanced virus production in infected cells. Similarly, infection-induced epithelial interferon signaling promoted enterocyte maturation and enhanced ACE2 expression. We here reveal a mechanism by which IFN-γ-driven inflammatory responses induce a vulnerable epithelial state with robust replication of SARS-CoV-2, which may have an impact on disease outcome and virus transmission.
Assuntos
COVID-19/etiologia , Interferon gama/imunologia , Modelos Imunológicos , SARS-CoV-2 , Enzima de Conversão de Angiotensina 2/genética , Enzima de Conversão de Angiotensina 2/metabolismo , Animais , COVID-19/imunologia , COVID-19/patologia , Diferenciação Celular/imunologia , Colo/imunologia , Colo/patologia , Colo/virologia , Suscetibilidade a Doenças , Enterócitos/metabolismo , Enterócitos/patologia , Enterócitos/virologia , Expressão Gênica , Interações entre Hospedeiro e Microrganismos/imunologia , Humanos , Interferon gama/administração & dosagem , Mucosa Intestinal/imunologia , Mucosa Intestinal/patologia , Mucosa Intestinal/virologia , Camundongos , Organoides/imunologia , Organoides/patologia , Organoides/virologia , SARS-CoV-2/genética , SARS-CoV-2/imunologia , SARS-CoV-2/patogenicidade , Replicação Viral/imunologiaRESUMO
Emerging mosquito-borne RNA viruses cause massive health complications worldwide. The Zika virus (ZIKV), in particular, has spread dramatically since 2007 and has provoked epidemics in the Americas and the South Pacific. The lack of antiviral therapy and vaccination has focused research on the investigation of ZIKV-host interactions, in order to understand underlying molecular infection mechanisms. We have established an approach for the analysis of ZIKV host dependency factors in a human trophoblast cell line and applied genome-wide CRISPR/Cas9 knockout mutagenesis. The presented method is especially of value for the identification of factors that are essential for placental infection with the potential to serve as targets for antiviral treatment.
Assuntos
Sistemas CRISPR-Cas , Infecção por Zika virus , Zika virus , Animais , Feminino , Humanos , Placenta/virologia , Gravidez , Trofoblastos , Replicação Viral , Zika virus/genética , Infecção por Zika virus/diagnósticoRESUMO
Successful asexual reproduction of intracellular pathogens depends on their potential to exploit host resources and subvert antimicrobial defense. In this work, we deployed two prevalent apicomplexan parasites of mammalian cells, namely Toxoplasma gondii and Eimeria falciformis, to identify potential host determinants of infection. Expression analyses of the young adult mouse colonic (YAMC) epithelial cells upon infection by either parasite showed regulation of several distinct transcripts, indicating that these two pathogens program their intracellular niches in a tailored manner. Conversely, parasitized mouse embryonic fibroblasts (MEFs) displayed a divergent transcriptome compared to corresponding YAMC epithelial cells, suggesting that individual host cells mount a fairly discrete response when encountering a particular pathogen. Among several host transcripts similarly altered by T. gondii and E. falciformis, we identified cFos, a master transcription factor, that was consistently induced throughout the infection. Indeed, asexual growth of both parasites was strongly impaired in MEF host cells lacking cFos expression. Last but not the least, our differential transcriptomics of the infected MEFs (parental and cFos-/- mutant) and YAMC epithelial cells disclosed a cFos-centered network, underlying signal cascades, as well as a repertoire of nucleotides- and ion-binding proteins, which presumably act in consort to acclimatize the mammalian cell and thereby facilitate the parasite development.
RESUMO
The transition zones of the squamous and columnar epithelia constitute hotspots for the emergence of cancer, often preceded by metaplasia, in which one epithelial type is replaced by another. It remains unclear how the epithelial spatial organization is maintained and how the transition zone niche is remodelled during metaplasia. Here we used single-cell RNA sequencing to characterize epithelial subpopulations and the underlying stromal compartment of endo- and ectocervix, encompassing the transition zone. Mouse lineage tracing, organoid culture and single-molecule RNA in situ hybridizations revealed that the two epithelia derive from separate cervix-resident lineage-specific stem cell populations regulated by opposing Wnt signals from the stroma. Using a mouse model of cervical metaplasia, we further show that the endocervical stroma undergoes remodelling and increases expression of the Wnt inhibitor Dickkopf-2 (DKK2), promoting the outgrowth of ectocervical stem cells. Our data indicate that homeostasis at the transition zone results from divergent stromal signals, driving the differential proliferation of resident epithelial lineages.
Assuntos
Colo do Útero/patologia , Epitélio/patologia , Homeostase , Via de Sinalização Wnt , Adenocarcinoma/genética , Adenocarcinoma/patologia , Animais , Biomarcadores Tumorais/metabolismo , Carcinoma de Células Escamosas/genética , Carcinoma de Células Escamosas/patologia , Diferenciação Celular , Linhagem da Célula , Microambiente Celular , Receptores ErbB/metabolismo , Feminino , Regulação Neoplásica da Expressão Gênica , Humanos , Queratinas/metabolismo , Metaplasia , Camundongos Endogâmicos C57BL , Organoides/patologia , Receptores Notch/metabolismo , Células-Tronco/patologia , Células Estromais/patologia , Transcrição Gênica , Neoplasias do Colo do Útero/genética , Neoplasias do Colo do Útero/patologiaRESUMO
The Th2 cytokine IL-13 is involved in biliary epithelial injury and liver fibrosis in patients as well as in animal models. The aim of this study was to investigate IL-13 as a therapeutic target during short term and chronic intrahepatic cholestasis in an Abcb4-knockout mouse model (Abcb4-/-). Lack of IL-13 protected Abcb4-/- mice transiently from cholestasis. This decrease in serum bile acids was accompanied by an enhanced excretion of bile acids and a normalization of fecal bile acid composition. In Abcb4-/-/IL-13-/- double knockout mice, bacterial translocation to the liver was significantly reduced and the intestinal microbiome resembled the commensal composition in wild type animals. In addition, 52-week-old Abcb4-/-IL-13-/- mice showed significantly reduced hepatic fibrosis. Abcb4-/- mice devoid of IL-13 transiently improved cholestasis and converted the composition of the gut microbiota towards healthy conditions. This highlights IL-13 as a potential therapeutic target in biliary diseases.
Assuntos
Colestase Intra-Hepática/terapia , Disbiose/terapia , Interleucina-13/metabolismo , Animais , Modelos Animais de Doenças , Humanos , Camundongos , Camundongos KnockoutRESUMO
An amendment to this paper has been published and can be accessed via a link at the top of the paper.
RESUMO
Rationale: Platelets are generated in the capillaries of the lung, control hemostasis, and display immunological functions. Tuberculosis primarily affects the lung, and patients show platelet changes and hemoptysis. A role of platelets in immunopathology of pulmonary tuberculosis requires careful assessment.Objectives: To identify the dynamics and interaction partners of platelets in the respiratory tissue and establish their impact on the outcome of pulmonary tuberculosis.Methods: Investigations were primarily performed in murine models of primary progressive pulmonary tuberculosis, by analysis of mouse strains with variable susceptibility to Mycobacterium tuberculosis infection using platelet depletion and delivery of antiplatelet drugs.Measurements and Main Results: Platelets were present at the site of infection and formed aggregates with different myeloid subsets during experimental tuberculosis. Such aggregates were also detected in patients with tuberculosis. Platelets were detrimental during the early phase of infection, and this effect was uncoupled from their canonical activation. Platelets left lung cell dynamics and patterns of antimycobacterial T-cell responses unchanged but hampered antimicrobial defense by restricting production of reactive oxygen species in lung-residing myeloid cells.Conclusions: Platelets are detrimental in primary progressive pulmonary tuberculosis, orchestrate lung immunity by modulating innate immune responsiveness, and may be amenable to new interventions for this deadly disease.