RESUMO
Sepsis is a life-threatening condition characterized by uncontrolled systemic inflammation and coagulation, leading to multiorgan failure. Therapeutic options to prevent sepsis-associated immunopathology remain scarce. Here, we established a mouse model of long-lasting disease tolerance during severe sepsis, manifested by diminished immunothrombosis and organ damage in spite of a high pathogen burden. We found that both neutrophils and B cells emerged as key regulators of tissue integrity. Enduring changes in the transcriptional profile of neutrophils include upregulated Cxcr4 expression in protected, tolerant hosts. Neutrophil Cxcr4 upregulation required the presence of B cells, suggesting that B cells promoted disease tolerance by improving tissue damage control via the suppression of neutrophils' tissue-damaging properties. Finally, therapeutic administration of a Cxcr4 agonist successfully promoted tissue damage control and prevented liver damage during sepsis. Our findings highlight the importance of a critical B-cell/neutrophil interaction during sepsis and establish neutrophil Cxcr4 activation as a potential means to promote disease tolerance during sepsis.
Assuntos
Infecções Bacterianas , Sepse , Animais , Infecções Bacterianas/metabolismo , Modelos Animais de Doenças , Camundongos , Insuficiência de Múltiplos Órgãos/metabolismo , Insuficiência de Múltiplos Órgãos/patologia , Neutrófilos/metabolismo , Sepse/metabolismoRESUMO
RATIONALE: Lung transplantation is the ultimate treatment option for patients with end-stage respiratory diseases but bears the highest mortality rate among all solid organ transplantations due to chronic lung allograft dysfunction (CLAD). The mechanisms leading to CLAD remain elusive due to an insufficient understanding of the complex post-transplant adaptation processes. OBJECTIVES: To better understand these lung adaptation processes after transplantation and to investigate their association with future changes in allograft function. METHODS: We performed an exploratory cohort study of bronchoalveolar lavage samples from 78 lung recipients and donors. We analysed the alveolar microbiome using 16S rRNA sequencing, the cellular composition using flow cytometry, as well as metabolome and lipidome profiling. MEASUREMENTS AND MAIN RESULTS: We established distinct temporal dynamics for each of the analysed data sets. Comparing matched donor and recipient samples, we revealed that recipient-specific as well as environmental factors, rather than the donor microbiome, shape the long-term lung microbiome. We further discovered that the abundance of certain bacterial strains correlated with underlying lung diseases even after transplantation. A decline in forced expiratory volume during the first second (FEV1) is a major characteristic of lung allograft dysfunction in transplant recipients. By using a machine learning approach, we could accurately predict future changes in FEV1 from our multi-omics data, whereby microbial profiles showed a particularly high predictive power. CONCLUSION: Bronchoalveolar microbiome, cellular composition, metabolome and lipidome show specific temporal dynamics after lung transplantation. The lung microbiome can predict future changes in lung function with high precision.
Assuntos
Transplante de Pulmão , Microbiota , Aloenxertos , Estudos de Coortes , Humanos , Pulmão , RNA Ribossômico 16S/genética , Estudos RetrospectivosRESUMO
Lipocalin 2 (LCN2) is a secreted glycoprotein with roles in multiple biological processes. It contributes to host defense by interference with bacterial iron uptake and exerts immunomodulatory functions in various diseases. Here, we aimed to characterize the function of LCN2 in lung macrophages and dendritic cells (DCs) using Lcn2-/- mice. Transcriptome analysis revealed strong LCN2-related effects in CD103+ DCs during homeostasis, with differential regulation of antigen processing and presentation and antiviral immunity pathways. We next validated the relevance of LCN2 in a mouse model of influenza infection, wherein LCN2 protected from excessive weight loss and improved survival. LCN2-deficiency was associated with enlarged mediastinal lymph nodes and increased lung T cell numbers, indicating a dysregulated immune response to influenza infection. Depletion of CD8+ T cells equalized weight loss between WT and Lcn2-/- mice, proving that LCN2 protects from excessive disease morbidity by dampening CD8+ T cell responses. In vivo T cell chimerism and in vitro T cell proliferation assays indicated that improved antigen processing by CD103+ DCs, rather than T cell intrinsic effects of LCN2, contribute to the exacerbated T cell response. Considering the antibacterial potential of LCN2 and that commensal microbes can modulate antiviral immune responses, we speculated that LCN2 might cause the observed influenza phenotype via the microbiome. Comparing the lung and gut microbiome of WT and Lcn2-/- mice by 16S rRNA gene sequencing, we observed profound effects of LCN2 on gut microbial composition. Interestingly, antibiotic treatment or co-housing of WT and Lcn2-/- mice prior to influenza infection equalized lung CD8+ T cell counts, suggesting that the LCN2-related effects are mediated by the microbiome. In summary, our results highlight a novel regulatory function of LCN2 in the modulation of antiviral immunity.
Assuntos
Influenza Humana/imunologia , Lipocalina-2/metabolismo , Microbiota/imunologia , Transcriptoma , Animais , Apresentação de Antígeno , Linfócitos T CD8-Positivos/imunologia , Células Dendríticas/imunologia , Células Dendríticas/virologia , Feminino , Microbioma Gastrointestinal , Homeostase , Humanos , Imunidade , Influenza Humana/virologia , Lipocalina-2/genética , Pulmão/imunologia , Pulmão/virologia , Ativação Linfocitária , Macrófagos/imunologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Organismos Livres de Patógenos EspecíficosRESUMO
STUDY QUESTION: Does signaling via the cannabinoid (CB1) receptor play a role in the pathogenesis of endometriosis in a mouse model? SUMMARY ANSWER: Mice treated with a CB1 agonist developed larger ectopic lesions, while less severe lesions developed in the absence of functional CB1 expression. WHAT IS KNOWN ALREADY: The expression of components of the endocannabinoid system has been demonstrated in both mouse and human uteri. CB1 receptors are expressed in human epithelial and stromal cell lines derived from eutopic endometrium and deep infiltrating endometriosis nodules. STUDY DESIGN, SIZE, DURATION: This was a randomized study in a mouse model of endometriosis. In a first set of experiments, mice with endometriosis were treated with the CB1 receptor agonist methanandamide (MET) (5 mg/kg, n = 20) on Days 1-5 and 8-12. In a second set of experiments, endometriosis development was evaluated in CB1-/- mice and in their wild-type (WT) littermates. PARTICIPANTS/MATERIALS, SETTING, METHODS: Endometriosis-like lesions were induced in Balb/c and C57/Bl6 mice. Two weeks after disease induction, the lesions were counted, measured and either included for immunohistochemistry analysis or frozen for gene expression profiling by semi-quantitative real-time PCR. To limit the role of chance, the experiments were conducted under standardized laboratory conditions with appropriate controls. MAIN RESULTS AND THE ROLE OF CHANCE: The lesion total volume was significantly higher in MET-treated compared with vehicle-treated mice (P < 0.05). Expression levels of mRNA for survivin, N-cadherin, integrin ß1 and interleukin-6 were increased in the ectopic endometrium of MET-treated versus vehicle-treated mice (P < 0.05). CB1-/- recipients that received endometrial tissue fragments from CB1-/- donors, WT recipients that received endometrial tissue fragments from CB1-/- donors and CB1-/- recipients that received endometrial tissue fragments from WT donors all showed a significant reduction in total lesion volume and lower expression of survivin and N-cadherin compared with WT recipients receiving uterine fragments from WT donors (P < 0.05). LARGE SCALE DATA: N/A. LIMITATIONS, REASONS FOR CAUTION: We provide evidence that endocannabinoid signaling via CB1 receptor plays a role in the development of endometriosis in a mouse model. However, the relative contribution of the CB1-mediated signaling pathways active in inflammatory, uterine and peritoneal cells remains to be ascertained. Since the study was performed in a mouse model, the significance of the findings in the human system warrants further investigation. WIDER IMPLICATIONS OF THE FINDINGS: Clarifying the function and regulation of CB1 and its molecular interactions with endogenous ligands, and how endocannabinoids levels are regulated in women with endometriosis, represent critical areas of research for the potential development of a novel medical treatment of the disease. STUDY FUNDING/COMPETING INTERESTS: A.M.S. was supported by a fellowship from Fondazione Giorgio Pardi. The authors have no conflicts of interest to declare.
Assuntos
Ácidos Araquidônicos/farmacologia , Agonistas de Receptores de Canabinoides/farmacologia , Endometriose/patologia , Doenças Peritoneais/patologia , Receptor CB1 de Canabinoide/metabolismo , Animais , Caderinas/metabolismo , Proliferação de Células/efeitos dos fármacos , Modelos Animais de Doenças , Endometriose/metabolismo , Endométrio/efeitos dos fármacos , Endométrio/metabolismo , Endométrio/patologia , Feminino , Proteínas Inibidoras de Apoptose/metabolismo , Integrina beta1/metabolismo , Interleucina-6/metabolismo , Camundongos , Doenças Peritoneais/metabolismo , Peritônio/efeitos dos fármacos , Peritônio/metabolismo , Peritônio/patologia , Proteínas Repressoras/metabolismo , Transdução de Sinais/efeitos dos fármacos , Transdução de Sinais/fisiologia , SurvivinaRESUMO
Hemolysis drives susceptibility to bacterial infections and predicts poor outcome from sepsis. These detrimental effects are commonly considered to be a consequence of heme-iron serving as a nutrient for bacteria. We employed a Gram-negative sepsis model and found that elevated heme levels impaired the control of bacterial proliferation independently of heme-iron acquisition by pathogens. Heme strongly inhibited phagocytosis and the migration of human and mouse phagocytes by disrupting actin cytoskeletal dynamics via activation of the GTP-binding Rho family protein Cdc42 by the guanine nucleotide exchange factor DOCK8. A chemical screening approach revealed that quinine effectively prevented heme effects on the cytoskeleton, restored phagocytosis and improved survival in sepsis. These mechanistic insights provide potential therapeutic targets for patients with sepsis or hemolytic disorders.
Assuntos
Infecções por Bactérias Gram-Negativas/imunologia , Fatores de Troca do Nucleotídeo Guanina/metabolismo , Heme/metabolismo , Hemólise/imunologia , Macrófagos/imunologia , Fagocitose , Sepse/imunologia , Animais , Antibacterianos/uso terapêutico , Citoesqueleto/metabolismo , Feminino , Infecções por Bactérias Gram-Negativas/tratamento farmacológico , Fatores de Troca do Nucleotídeo Guanina/genética , Heme Oxigenase-1/genética , Hemólise/efeitos dos fármacos , Humanos , Evasão da Resposta Imune , Macrófagos/efeitos dos fármacos , Macrófagos/microbiologia , Proteínas de Membrana/genética , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Fagocitose/efeitos dos fármacos , Quinina/uso terapêutico , Células RAW 264.7 , Sepse/tratamento farmacológico , Proteína cdc42 de Ligação ao GTP/metabolismoRESUMO
Extravillous trophoblasts (EVTs) play a central role in educating maternal leukocytes, endometrial stromal and endothelial cells to generate a receptive decidual microenvironment tailored to accept the semi-allogeneic fetus. HLA-G, a non-classical HLA class I molecule endowed with immune-regulatory functions, is primarily expressed on EVTs lining the placenta and on the naturally occurring tolerogenic dendritic cells, named DC-10, which are enriched in the human first trimester decidua. Decidual DC-10 are involved in HLA-G-mediated tolerance at the maternal-fetal interface. EVTs not only establish a tolerogenic microenvironment through the interaction with maternal innate and adaptive cells but also orchestrate placenta vascular and tissue remodeling, leading to a successful pregnancy. Here, we discuss the potential implications of the HLA-G-mediated cross-talk among the cells present at the maternal-fetal interface, and its role in maintaining a positive relationship between the mother and the fetus.
RESUMO
Endometriosis is caused by the displacement of endometrium outside the uterus contributing heavily to infertility and debilitating pelvic pain. Ectopic adhesion and growth are believed to occur under the influence of a favorable hormonal environment and immunological factors. The objective of this study is to analyze the effect of a targeted therapy with an antibody-based pharmacodelivery of interleukin 4 (F8-IL4) in a mouse model of experimentally induced endometriosis. Endometriosis-like lesions were induced in Balb/c mice. The animals were treated intravenously with F8-IL4 or with untargeted IL4 (KSF-IL4). Twelve days after disease induction, the lesions were isolated. A significant reduction in the number of total lesions/mouse and in the total volume of lesions/mouse was observed in mice treated with F8-IL4 compared to controls (P = .029 and P = .006, respectively), while no difference was found between KSF-IL4-treated mice and their controls. Gene expression was evaluated by quantitative real-time polymerase chain reaction. Expression of genes involved in cell adhesion, extracellular matrix invasion, and neovascularization was significantly downregulated in F8-IL4-treated mice compared to their controls (integrin ß1: P = .02; metalloproteinase [MMP] 3: P = .02; MMP9: P = .04; vascular endothelial growth factor: P = .04). Gene expression of inflammatory cytokines (tumor necrosis factor α, IL1ß, IL1α, and IL6) did not vary in the ectopic lesions isolated from F8-IL4-treated mice compared to their controls. Immunohistochemistry demonstrated a significantly reduced expression of E-cadherin and ß-catenin in the lesions of mice treated with F8-IL4. Our results show that the antibody-mediated targeted delivery of IL4 inhibits the development of endometriosis in a syngeneic mouse model by likely impairing adhesion, invasion, and vascularization of the ectopic endometrium.
Assuntos
Anticorpos Monoclonais/farmacologia , Sistemas de Liberação de Medicamentos , Endometriose/prevenção & controle , Endométrio/efeitos dos fármacos , Imunoconjugados/farmacologia , Região Variável de Imunoglobulina/farmacologia , Interleucina-4/farmacologia , Animais , Anticorpos Monoclonais/administração & dosagem , Anticorpos Monoclonais/metabolismo , Anticorpos Monoclonais Humanizados , Caderinas/genética , Caderinas/metabolismo , Adesão Celular/efeitos dos fármacos , Movimento Celular/efeitos dos fármacos , Química Farmacêutica , Citocinas/genética , Citocinas/metabolismo , Citoproteção , Modelos Animais de Doenças , Endometriose/genética , Endometriose/metabolismo , Endometriose/patologia , Endométrio/irrigação sanguínea , Endométrio/metabolismo , Endométrio/patologia , Feminino , Regulação da Expressão Gênica , Imunoconjugados/administração & dosagem , Imunoconjugados/metabolismo , Região Variável de Imunoglobulina/administração & dosagem , Região Variável de Imunoglobulina/metabolismo , Mediadores da Inflamação/metabolismo , Injeções Intravenosas , Interleucina-4/administração & dosagem , Interleucina-4/metabolismo , Camundongos Endogâmicos BALB C , Neovascularização Patológica , Proteínas Recombinantes de Fusão/farmacologia , Fatores de Tempo , beta Catenina/genética , beta Catenina/metabolismoRESUMO
OBJECTIVE: To analyze the WNT/ß-catenin signaling pathway in luteinized granulosa cells from women with and without endometriosis in relation to cellular apoptosis. DESIGN: Basic. SETTING: University hospital. PATIENT(S): Patients with a laparoscopic diagnosis of endometriosis (n = 30) and women undergoing intracytoplasmic sperm injection for male infertility (control group n = 39). INTERVENTION(S): Isolation of luteinized granulosa cells. MAIN OUTCOME MEASURE(S): Gene expression analysis of components of the WNT/ß-catenin pathway, protein expression levels of ß-catenin, and cell cycle studies in luteinized granulosa cells. RESULT(S): Compared with luteinized granulosa cells from control women, cells derived from endometriosis patients had significantly higher transcript levels of the ß-catenin-independent molecules WNT4 and WNT5a and lower levels of the ß-catenin-dependent molecule WNT1. A decrease of total ß-catenin as well as of its dephosphorylated active form, together with an aberrant gene expression of the downstream targets survivin and BMP4, was detected in cells from affected women. Flow cytometry analysis confirmed an enhanced apoptosis of luteinized granulosa cells from patients with endometriosis. CONCLUSION(S): The concomitant dysregulation of specific members of the WNT pathway and of its pivot molecule ß-catenin in granulosa cells characterized by an increased apoptosis suggests that the WNT/ß-catenin signaling pathway might be involved in leading to granulosa cell atresia.