RESUMO
Converging evidence indicates that extra-embryonic yolk sac is the source of both macrophages and endothelial cells in adult mouse tissues. Prevailing views are that these embryonically derived cells are maintained after birth by proliferative self-renewal in their differentiated states. Here we identify clonogenic endothelial-macrophage (EndoMac) progenitor cells in the adventitia of embryonic and postnatal mouse aorta, that are independent of Flt3-mediated bone marrow hematopoiesis and derive from an early embryonic CX3CR1+ and CSF1R+ source. These bipotent progenitors are proliferative and vasculogenic, contributing to adventitial neovascularization and formation of perfused blood vessels after transfer into ischemic tissue. We establish a regulatory role for angiotensin II, which enhances their clonogenic and differentiation properties and rapidly stimulates their proliferative expansion in vivo. Our findings demonstrate that embryonically derived EndoMac progenitors participate in local vasculogenic responses in the aortic wall by contributing to the expansion of endothelial cells and macrophages postnatally.
Assuntos
Aorta , Macrófagos , Animais , Macrófagos/citologia , Macrófagos/metabolismo , Aorta/citologia , Camundongos , Receptor 1 de Quimiocina CX3C/metabolismo , Receptor 1 de Quimiocina CX3C/genética , Células Endoteliais/citologia , Células Endoteliais/metabolismo , Diferenciação Celular , Receptores de Fator Estimulador das Colônias de Granulócitos e Macrófagos/metabolismo , Receptores de Fator Estimulador das Colônias de Granulócitos e Macrófagos/genética , Angiotensina II , Proliferação de Células , Células-Tronco/citologia , Células-Tronco/metabolismo , Camundongos Endogâmicos C57BL , Feminino , Neovascularização Fisiológica , Receptores de Quimiocinas/metabolismo , Receptores de Quimiocinas/genética , Masculino , Hematopoese/fisiologia , Tirosina Quinase 3 Semelhante a fmsRESUMO
Enoblituzumab, an immunotherapeutic agent targeting CD276, shows both safety and efficacy in activating T cells and oligodendrocyte-like cells against various cancers. Preclinical studies and mouse models suggest that therapies targeting CD276 may outperform PD1/PD-L1 blockade. However, data from mouse models indicate a significant non-responsive population to anti-CD276 treatment, with the mechanisms of resistance still unclear. In this study, we evaluate the activity of anti-CD276 antibodies in a chemically-induced murine model of head and neck squamous cell carcinoma. Using models of induced and orthotopic carcinogenesis, we identify ITGB6 as a key gene mediating differential responses to anti-CD276 treatment. Through single-cell RNA sequencing and gene-knockout mouse models, we find that ITGB6 regulates the expression of the tumor-associated chemokine CX3CL1, which recruits and activates PF4+ macrophages that express high levels of CX3CR1. Inhibition of the CX3CL1-CX3CR1 axis suppresses the infiltration and secretion of CXCL16 by PF4+ macrophages, thereby reinvigorating cytotoxic CXCR6+ CD8+ T cells and enhancing sensitivity to anti-CD276 treatment. Further investigations demonstrate that inhibiting ITGB6 restores sensitivity to PD1 antibodies in mice resistant to anti-PD1 treatment. In summary, our research reveals a resistance mechanism associated with immune checkpoint inhibitor therapy and identifies potential targets to overcome resistance in cancer treatment.
Assuntos
Antígenos B7 , Neoplasias de Cabeça e Pescoço , Camundongos Knockout , Animais , Camundongos , Antígenos B7/metabolismo , Antígenos B7/genética , Antígenos B7/antagonistas & inibidores , Humanos , Neoplasias de Cabeça e Pescoço/genética , Neoplasias de Cabeça e Pescoço/imunologia , Neoplasias de Cabeça e Pescoço/tratamento farmacológico , Neoplasias de Cabeça e Pescoço/patologia , Receptor 1 de Quimiocina CX3C/metabolismo , Receptor 1 de Quimiocina CX3C/genética , Resistencia a Medicamentos Antineoplásicos/genética , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Resistencia a Medicamentos Antineoplásicos/imunologia , Macrófagos/imunologia , Macrófagos/metabolismo , Linhagem Celular Tumoral , Camundongos Endogâmicos C57BL , Carcinoma de Células Escamosas de Cabeça e Pescoço/tratamento farmacológico , Carcinoma de Células Escamosas de Cabeça e Pescoço/imunologia , Carcinoma de Células Escamosas de Cabeça e Pescoço/genética , Carcinoma de Células Escamosas de Cabeça e Pescoço/patologia , Carcinoma de Células Escamosas de Cabeça e Pescoço/metabolismo , Modelos Animais de Doenças , Feminino , Antineoplásicos Imunológicos/farmacologia , Antineoplásicos Imunológicos/uso terapêutico , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacosRESUMO
Although oral tolerance is a critical system in regulating allergic disorders, the mechanisms by which dietary factors regulate the induction and maintenance of oral tolerance remain unclear. To address this, we explored the differentiation and function of various immune cells in the intestinal immune system under fasting and ad libitum-fed conditions before oral ovalbumin (OVA) administration. Fasting mitigated OVA-specific Treg expansion, which is essential for oral tolerance induction. This abnormality mainly resulted from functional defects in the CX3CR1+ cells responsible for the uptake of luminal OVA and reduction of tolerogenic CD103+ dendritic cells. Eventually, fasting impaired the preventive effect of oral OVA administration on asthma and allergic rhinitis development. Specific food ingredients, namely carbohydrates and arginine, were indispensable for oral tolerance induction by activating glycolysis and mTOR signaling. Overall, prior food intake and nutritional signals are critical for maintaining immune homeostasis by inducing tolerance to ingested food antigens.
Assuntos
Arginina , Células Dendríticas , Tolerância Imunológica , Ovalbumina , Linfócitos T Reguladores , Serina-Treonina Quinases TOR , Animais , Arginina/metabolismo , Linfócitos T Reguladores/imunologia , Ovalbumina/imunologia , Células Dendríticas/imunologia , Células Dendríticas/metabolismo , Camundongos , Serina-Treonina Quinases TOR/metabolismo , Camundongos Endogâmicos C57BL , Administração Oral , Receptor 1 de Quimiocina CX3C/metabolismo , Intestinos/imunologia , Antígenos CD/metabolismo , Cadeias alfa de Integrinas/metabolismo , Açúcares/metabolismo , Glicólise , Jejum , Transdução de Sinais , Mucosa Intestinal/imunologia , Mucosa Intestinal/metabolismo , FemininoRESUMO
Diabetes mellitus (DM) is the most common metabolic disease in humans, and its prevalence is increasing worldwide in parallel with the obesity pandemic. A lack of insulin or insulin resistance, and consequently hyperglycemia, leads to many systemic disorders, among which diabetic encephalopathy (DE) is a long-term complication of the central nervous system (CNS), characterized by cognitive impairment and motor dysfunctions. The role of oxidative stress and neuroinflammation in the pathomechanism of DE has been proven. Fractalkine (CX3CL1) has unique properties as an adhesion molecule and chemoattractant, and by acting on its only receptor, CX3CR1, it regulates the activity of microglia in physiological states and neuroinflammation. Depending on the clinical context, CX3CL1-CX3CR1 signaling may have neuroprotective effects by inhibiting the inflammatory process in microglia or, conversely, maintaining/intensifying inflammation and neurotoxicity. This review discusses the evidence supporting that the CX3CL1-CX3CR1 pair is neuroprotective and other evidence that it is neurotoxic. Therefore, interrupting the vicious cycle within neuron-microglia interactions by promoting neuroprotective effects or inhibiting the neurotoxic effects of the CX3CL1-CX3CR1 signaling axis may be a therapeutic goal in DE by limiting the inflammatory response. However, the optimal approach to prevent DE is simply tight glycemic control, because the elimination of dysglycemic states in the CNS abolishes the fundamental mechanisms that induce this vicious cycle.
Assuntos
Quimiocina CX3CL1 , Microglia , Transdução de Sinais , Humanos , Quimiocina CX3CL1/metabolismo , Animais , Microglia/metabolismo , Microglia/patologia , Receptor 1 de Quimiocina CX3C/metabolismoRESUMO
Oligodendrocyte death is common in aging and neurodegenerative disease. In these conditions, dying oligodendrocytes must be efficiently removed to allow remyelination and to prevent a feedforward degenerative cascade. Removal of this cellular debris is thought to primarily be carried out by resident microglia. To investigate the cellular dynamics underlying how microglia do this, we use a single-cell cortical demyelination model combined with longitudinal intravital imaging of dual-labeled transgenic mice. Following phagocytosis, single microglia clear the targeted oligodendrocyte and its myelin sheaths in one day via a precise, rapid, and stereotyped sequence. Deletion of the fractalkine receptor, CX3CR1, delays the microglial phagocytosis of the cell soma but has no effect on clearance of myelin sheaths. Unexpectedly, deletion of the phosphatidylserine receptor, MERTK, has no effect on oligodendrocyte or myelin sheath clearance. Thus, separate molecular signals are used to detect, engage, and clear distinct sub-compartments of dying oligodendrocytes to maintain tissue homeostasis.
Assuntos
Receptor 1 de Quimiocina CX3C , Microglia , Oligodendroglia , Fagocitose , c-Mer Tirosina Quinase , Animais , Oligodendroglia/metabolismo , Microglia/metabolismo , Receptor 1 de Quimiocina CX3C/metabolismo , Receptor 1 de Quimiocina CX3C/genética , c-Mer Tirosina Quinase/metabolismo , c-Mer Tirosina Quinase/genética , Camundongos , Bainha de Mielina/metabolismo , Camundongos Transgênicos , Camundongos Endogâmicos C57BL , Morte CelularRESUMO
Acute kidney injury (AKI) is a devastating consequence of sepsis, accompanied by high mortality rates. It was suggested that inflammatory pathways are closely linked to the pathogenesis of lipopolysaccharide (LPS)-induced AKI. Inflammatory signaling, including PCSK9, HMGB1/RAGE/TLR4/MYD88/NF-κB, NLRP3/caspase-1 and Fractalkine/CX3CR1 are considered major forerunners in this link. Alirocumab, PCSK9 inhibitor, with remarkable anti-inflammatory features. Accordingly, this study aimed to elucidate the antibacterial effect of alirocumab against E. coli in vitro. Additionally, evaluation of the potential nephroprotective effects of alirocumab against LPS-induced AKI in rats, highlighting the potential underlying mechanisms involved in these beneficial actions. Thirty-six adult male Wistar rats were assorted into three groups (n=12). Group I; was a normal control group, whereas sepsis-mediated AKI was induced in groups II and III through single-dose intraperitoneal injection of LPS on day 16. In group III, animals were given alirocumab. The results revealed that LPS-induced AKI was mitigated by alirocumab, evidenced by amelioration in renal function tests (creatinine, cystatin C, KIM-1, and NGAL); oxidative stress biomarkers (Nrf2, HO-1, TAC, and MDA); apoptotic markers and renal histopathological findings. Besides, alirocumab pronouncedly hindered LPS-mediated inflammatory response, confirmed by diminishing HMGB1, TNF-α, IL-1ß, and caspase-1 contents; the gene expression of PCSK9, RAGE, NF-á´B and Fractalkine/CX3CR1, along with mRNA expression of TLR4, MYD88, and NLRP3. Regarding the antibacterial actions, results showed that alirocumab displayed potential anti-bacterial activity against pathogenic gram-negative E. coli. In conclusion, alirocumab elicited nephroprotective activities against LPS-induced AKI via modulation of Nrf2/HO-1, PCSK9, HMGB1/RAGE/TLR4/MYD88/NF-á´B/NLRP3/Caspase-1, Fractalkine/CX3R1 and apoptotic axes.
Assuntos
Injúria Renal Aguda , Anticorpos Monoclonais Humanizados , Antioxidantes , Receptor 1 de Quimiocina CX3C , Quimiocina CX3CL1 , Proteína HMGB1 , Heme Oxigenase (Desciclizante) , Fator 2 Relacionado a NF-E2 , NF-kappa B , Proteína 3 que Contém Domínio de Pirina da Família NLR , Ratos Wistar , Sepse , Transdução de Sinais , Animais , Masculino , Sepse/complicações , Sepse/tratamento farmacológico , Sepse/metabolismo , Proteína HMGB1/metabolismo , Quimiocina CX3CL1/metabolismo , Injúria Renal Aguda/metabolismo , Injúria Renal Aguda/tratamento farmacológico , Injúria Renal Aguda/induzido quimicamente , Injúria Renal Aguda/patologia , Proteína 3 que Contém Domínio de Pirina da Família NLR/metabolismo , Ratos , Heme Oxigenase (Desciclizante)/metabolismo , Anticorpos Monoclonais Humanizados/farmacologia , NF-kappa B/metabolismo , Fator 2 Relacionado a NF-E2/metabolismo , Antioxidantes/farmacologia , Receptor 1 de Quimiocina CX3C/metabolismo , Transdução de Sinais/efeitos dos fármacos , Inflamação/tratamento farmacológico , Inflamação/metabolismo , Inflamação/patologia , Modelos Animais de Doenças , Lipopolissacarídeos , Inibidores de PCSK9 , Rim/efeitos dos fármacos , Rim/patologia , Rim/metabolismo , Pró-Proteína Convertase 9/metabolismo , Pró-Proteína Convertase 9/genética , Estresse Oxidativo/efeitos dos fármacos , Anti-Inflamatórios/farmacologiaRESUMO
Recent progress in single-cell profiling technologies has revealed significant phenotypic and transcriptional heterogeneity in tumor-infiltrating CD8+ T cells. However, the transition between the different states of intratumoral antigen-specific CD8+ T cells remains elusive. Here, we sought to examine the generation, transcriptomic states, and the clinical relevance of melanoma-infiltrating CD8+ T cells expressing a chemokine receptor and T-cell differentiation marker, CX3C chemokine receptor 1 (CX3CR1). Analysis of single-cell datasets revealed distinct human melanoma-infiltrating CD8+ T-cell clusters expressing genes associated with effector T-cell function but with distinguishing expression of CX3CR1 or PDCD1. No obvious impact of CX3CR1 expression in melanoma on the response to immune checkpoint inhibitor therapy was observed while increased pretreatment and on-treatment frequency of a CD8+ T-cell cluster expressing high levels of exhaustion markers was associated with poor response to the treatment. Adoptively transferred antigen-specific CX3CR1- CD8+ T cells differentiated into the CX3CR1+ subset in mice treated with FTY720, which inhibits lymphocyte egress from secondary lymphoid tissues, suggesting the intratumoral generation of CX3CR1+ CD8+ T cells rather than their trafficking from secondary lymphoid organs. Furthermore, analysis of adoptively transferred antigen-specific CD8+ T cells, in which the Cx3cr1 gene was replaced with a marker gene confirmed that CX3CR1+ CD8+ T cells could directly differentiate from the intratumoral CX3CR1- subset. These findings highlight that tumor antigen-specific CX3CR1- CD8+ T cells can fully differentiate outside the secondary lymphoid organs and generate CX3CR1+ CD8+ T cells in the tumor microenvironment, which are distinct from CD8+ T cells that express markers of exhaustion. SIGNIFICANCE: Intratumoral T cells are composed of heterogeneous subpopulations with various phenotypic and transcriptional states. This study illustrates the intratumoral generation of antigen-specific CX3CR1+ CD8+ T cells that exhibit distinct transcriptomic signatures and clinical relevance from CD8+ T cells expressing markers of exhaustion.
Assuntos
Linfócitos T CD8-Positivos , Receptor 1 de Quimiocina CX3C , Linfócitos do Interstício Tumoral , Melanoma , Transcriptoma , Receptor 1 de Quimiocina CX3C/genética , Receptor 1 de Quimiocina CX3C/metabolismo , Linfócitos T CD8-Positivos/imunologia , Linfócitos T CD8-Positivos/metabolismo , Melanoma/imunologia , Melanoma/genética , Melanoma/patologia , Animais , Humanos , Camundongos , Linfócitos do Interstício Tumoral/imunologia , Linfócitos do Interstício Tumoral/metabolismo , Inibidores de Checkpoint Imunológico/farmacologia , Inibidores de Checkpoint Imunológico/uso terapêutico , Cloridrato de Fingolimode/farmacologia , Cloridrato de Fingolimode/uso terapêutico , Camundongos Endogâmicos C57BL , Análise de Célula Única , Relevância ClínicaRESUMO
As a versatile element for maintaining homeostasis, the chemokine system has been reported to be implicated in the pathogenesis of immune thrombocytopenia (ITP). However, research pertaining to chemokine receptors and related ligands in adult ITP is still limited. The states of several typical chemokine receptors and cognate ligands in the circulation were comparatively assessed through various methodologies. Multiple variable analyses of correlation matrixes were conducted to characterize the correlation signatures of various chemokine receptors or candidate ligands with platelet counts. Our data illustrated a significant decrease in relative CXCR3 expression and elevated plasma levels of CXCL4, 9-11, 13, and CCL3 chemokines in ITP patients with varied platelet counts. Flow cytometry assays revealed eminently diminished CXCR3 levels on T and B lymphocytes and increased CXCR5 on cytotoxic T cell (Tc) subsets in ITP patients with certain platelet counts. Meanwhile, circulating CX3CR1 levels were markedly higher on T cells with a concomitant increase in plasma CX3CL1 level in ITP patients, highlighting the importance of aberrant alterations of the CX3CR1-CX3CL1 axis in ITP pathogenesis. Spearman's correlation analyses revealed a strong positive association of peripheral CXCL4 mRNA level, and negative correlations of plasma CXCL4 concentration and certain chemokine receptors with platelet counts, which might serve as a potential biomarker of platelet destruction in ITP development. Overall, these results indicate that the differential expression patterns and distinct activation states of peripheral chemokine network, and the subsequent expansion of circulating CXCR5+ Tc cells and CX3CR1+ T cells, may be a hallmark during ITP progression, which ultimately contributes to thrombocytopenia in ITP patients.
Assuntos
Receptor 1 de Quimiocina CX3C , Púrpura Trombocitopênica Idiopática , Receptores CXCR3 , Receptores CXCR5 , Humanos , Receptores CXCR3/metabolismo , Púrpura Trombocitopênica Idiopática/sangue , Púrpura Trombocitopênica Idiopática/imunologia , Receptor 1 de Quimiocina CX3C/metabolismo , Masculino , Receptores CXCR5/metabolismo , Feminino , Adulto , Pessoa de Meia-Idade , Contagem de Plaquetas , Fator Plaquetário 4/sangue , Fator Plaquetário 4/metabolismo , Idoso , Linfócitos B/imunologia , Linfócitos B/metabolismoRESUMO
Neutralizing antibodies are considered a correlate of protection against severe human respiratory syncytial virus (HRSV) disease. Currently, HRSV neutralization assays are performed on immortalized cell lines like Vero or A549 cells. It is known that assays on these cell lines exclusively detect neutralizing antibodies (nAbs) directed to the fusion (F) protein. For the detection of nAbs directed to the glycoprotein (G), ciliated epithelial cells expressing the cellular receptor CX3CR1 are required, but generation of primary cell cultures is expensive and labor-intensive. Here, we developed a high-throughput neutralization assay based on the interaction between clinically relevant HRSV grown on primary cells with ciliated epithelial cells, and validated this assay using a panel of infant sera. To develop the high-throughput neutralization assay, we established a culture of differentiated apical-out airway organoids (Ap-O AO). CX3CR1 expression was confirmed, and both F- and G-specific monoclonal antibodies neutralized HRSV in the Ap-O AO. In a side-by-side neutralization assay on Vero cells and Ap-O AO, neutralizing antibody levels in sera from 125 infants correlated well, although titers on Ap-O AO were consistently lower. We speculate that these lower titers might be an actual reflection of the neutralizing antibody capacity in vivo. The organoid-based neutralization assay described here holds promise for further characterization of correlates of protection against HRSV disease.
Assuntos
Anticorpos Neutralizantes , Receptor 1 de Quimiocina CX3C , Testes de Neutralização , Organoides , Infecções por Vírus Respiratório Sincicial , Vírus Sincicial Respiratório Humano , Humanos , Vírus Sincicial Respiratório Humano/imunologia , Anticorpos Neutralizantes/imunologia , Organoides/metabolismo , Organoides/imunologia , Organoides/virologia , Organoides/citologia , Animais , Testes de Neutralização/métodos , Chlorocebus aethiops , Células Vero , Infecções por Vírus Respiratório Sincicial/imunologia , Infecções por Vírus Respiratório Sincicial/virologia , Receptor 1 de Quimiocina CX3C/metabolismo , Receptor 1 de Quimiocina CX3C/imunologia , Anticorpos Antivirais/imunologia , Proteínas Virais de Fusão/imunologia , Proteínas Virais de Fusão/metabolismo , Lactente , Células Epiteliais/metabolismo , Células Epiteliais/imunologia , Células Epiteliais/virologia , Anticorpos Monoclonais/imunologiaRESUMO
Inflammatory bowel disease (IBD) is a chronic inflammatory intestinal disease, characterized by dysregulated immune response. HDAC3 is reported to be an epigenetic brake in inflammation, playing critical roles in macrophages. However, its role in IBD is unclear. In our study, we found HDAC3 was upregulated in CX3CR1-positive cells in the mucosa from IBD mice. Conditional knockout (cKO) of Hdac3 in CX3CR1 positive cells attenuated the disease severity of Dextran Sulfate Sodium (DSS)-induced colitis. In addition, inhibition of HDAC3 with RGFP966 could also alleviate the DSS-induced tissue injury and inflammation in IBD. The RNA sequencing results revealed that Hdac3 cKO restrained DSS-induced upregulation of genes in the pathways of cytokine-cytokine receptor interaction, complement and coagulation cascades, chemokine signaling, and extracellular matrix receptor interaction. We also identified that Guanylate-Binding Protein 5 (GBP5) was transcriptionally regulated by HDAC3 in monocytes by RNA sequencing. Inhibition of HDAC3 resulted in decreased transcriptional activity of interferon-gamma-induced expression of GBP5 in CX3CR1-positive cells, such as macrophages and microglia. Overexpression of HDAC3 upregulated the transcriptional activity of GBP5 reporter. Lastly, conditional knockout of Hdac3 in macrophages (Hdac3 mKO) attenuated the disease severity of DSS-induced colitis. In conclusion, inhibition of HDAC3 in macrophages could ameliorate the disease severity and inflammatory response in colitis by regulating GBP5-NLRP3 axis, identifying a new therapeutic avenue for the treatment of colitis.
Assuntos
Colite , Sulfato de Dextrana , Histona Desacetilases , Macrófagos , Camundongos Knockout , Proteína 3 que Contém Domínio de Pirina da Família NLR , Transdução de Sinais , Animais , Sulfato de Dextrana/toxicidade , Sulfato de Dextrana/efeitos adversos , Histona Desacetilases/metabolismo , Histona Desacetilases/genética , Camundongos , Macrófagos/metabolismo , Macrófagos/imunologia , Proteína 3 que Contém Domínio de Pirina da Família NLR/metabolismo , Proteína 3 que Contém Domínio de Pirina da Família NLR/genética , Colite/induzido quimicamente , Colite/genética , Colite/patologia , Colite/metabolismo , Humanos , Transdução de Sinais/efeitos dos fármacos , Doenças Inflamatórias Intestinais/induzido quimicamente , Doenças Inflamatórias Intestinais/genética , Doenças Inflamatórias Intestinais/patologia , Doenças Inflamatórias Intestinais/metabolismo , Doenças Inflamatórias Intestinais/tratamento farmacológico , Proteínas de Ligação ao GTP/genética , Proteínas de Ligação ao GTP/metabolismo , Proteínas de Ligação ao GTP/antagonistas & inibidores , Modelos Animais de Doenças , Receptor 1 de Quimiocina CX3C/metabolismo , Receptor 1 de Quimiocina CX3C/genética , Camundongos Endogâmicos C57BL , Inibidores de Histona Desacetilases/farmacologia , Inibidores de Histona Desacetilases/uso terapêutico , Mucosa Intestinal/patologia , Mucosa Intestinal/metabolismo , Mucosa Intestinal/efeitos dos fármacos , Acrilamidas , FenilenodiaminasRESUMO
The cellular distribution and changes in CX3CL1/fractalkine and its receptor CX3CR1 protein levels in the trigeminal subnucleus caudalis (TSC) of rats with unilateral infraorbital nerve ligation (IONL) were investigated on postoperation days 1, 3, 7, and 14 (POD1, POD3, POD7, and POD14, respectively) and compared with those of sham-operated and naïve controls. Behavioral tests revealed a significant increase in tactile hypersensitivity bilaterally in the vibrissal pads of both sham- and IONL-operated animals from POD1 to POD7, with a trend towards normalization in sham controls at POD14. Image analysis revealed increased CX3CL1 immunofluorescence (IF) intensities bilaterally in the TSC neurons of both sham- and IONL-operated rats at all survival periods. Reactive astrocytes in the ipsilateral TSC also displayed CX3CL1-IF from POD3 to POD14. At POD1 and POD3, microglial cells showed high levels of CX3CR1-IF, which decreased by POD7 and POD14. Conversely, CX3CR1 was increased in TSC neurons and reactive astrocytes at POD7 and POD14, which coincided with high levels of CX3CL1-IF and ADAM17-IF. This indicates that CX3CL1/CX3CR1 may be involved in reciprocal signaling between TSC neurons and reactive astrocytes. The level of CatS-IF in microglial cells suggests that soluble CX3CL1 may be involved in neuron-microglial cell signaling at POD3 and POD7, while ADAM17 allows this release at all studied time points. These results indicate an extended CX3CL1/CX3CR1 signaling axis and its role in the crosstalk between TSC neurons and glial cells during the development of trigeminal neuropathic pain.
Assuntos
Receptor 1 de Quimiocina CX3C , Quimiocina CX3CL1 , Transdução de Sinais , Animais , Quimiocina CX3CL1/metabolismo , Ratos , Receptor 1 de Quimiocina CX3C/metabolismo , Receptor 1 de Quimiocina CX3C/genética , Masculino , Microglia/metabolismo , Neuralgia do Trigêmeo/metabolismo , Neuralgia do Trigêmeo/patologia , Neurônios/metabolismo , Astrócitos/metabolismo , Neuralgia/metabolismo , Neuralgia/patologia , Ratos Sprague-DawleyRESUMO
Acute brain slices represent a workhorse model for studying the central nervous system (CNS) from nanoscale events to complex circuits. While slice preparation inherently involves tissue damage, it is unclear how microglia, the main immune cells and damage sensors of the CNS react to this injury and shape neuronal activity ex vivo. To this end, we investigated microglial phenotypes and contribution to network organization and functioning in acute brain slices. We reveal time-dependent microglial phenotype changes influenced by complex extracellular ATP dynamics through P2Y12R and CX3CR1 signalling, which is sustained for hours in ex vivo mouse brain slices. Downregulation of P2Y12R and changes of microglia-neuron interactions occur in line with alterations in the number of excitatory and inhibitory synapses over time. Importantly, functional microglia modulate synapse sprouting, while microglial dysfunction results in markedly impaired ripple activity both ex vivo and in vivo. Collectively, our data suggest that microglia are modulators of complex neuronal networks with important roles to maintain neuronal network integrity and activity. We suggest that slice preparation can be used to model time-dependent changes of microglia-neuron interactions to reveal how microglia shape neuronal circuits in physiological and pathological conditions.
Assuntos
Trifosfato de Adenosina , Encéfalo , Receptor 1 de Quimiocina CX3C , Microglia , Neurônios , Receptores Purinérgicos P2Y12 , Sinapses , Animais , Microglia/metabolismo , Trifosfato de Adenosina/metabolismo , Camundongos , Neurônios/metabolismo , Receptor 1 de Quimiocina CX3C/metabolismo , Receptor 1 de Quimiocina CX3C/genética , Receptores Purinérgicos P2Y12/metabolismo , Receptores Purinérgicos P2Y12/genética , Encéfalo/metabolismo , Sinapses/metabolismo , Camundongos Endogâmicos C57BL , Fenótipo , Masculino , Transdução de SinaisRESUMO
Multiple sclerosis (MS) is a complex autoimmune disease of central nervous system (CNS) characterized by the myelin sheath destruction and compromised nerve signal transmission. Understanding molecular mechanisms driving MS development is critical due to its early onset, chronic course, and therapeutic approaches based only on symptomatic treatment. Cytokines are known to play a pivotal role in the MS pathogenesis with interleukin-6 (IL-6) being one of the key mediators. This study investigates contribution of IL-6 produced by microglia and dendritic cells to the development of experimental autoimmune encephalomyelitis (EAE), a widely used mouse model of MS. Mice with conditional inactivation of IL-6 in the CX3CR1+ cells, including microglia, or CD11c+ dendritic cells, displayed less severe symptoms as compared to their wild-type counterparts. Mice with microglial IL-6 deletion exhibited an elevated proportion of regulatory T cells and reduced percentage of pathogenic IFNγ-producing CD4+ T cells, accompanied by the decrease in pro-inflammatory monocytes in the CNS at the peak of EAE. At the same time, deletion of IL-6 from microglia resulted in the increase of CCR6+ T cells and GM-CSF-producing T cells. Conversely, mice with IL-6 deficiency in the dendritic cells showed not only the previously described increase in the proportion of regulatory T cells and decrease in the proportion of TH17 cells, but also reduction in the production of GM-CSF and IFNγ in the secondary lymphoid organs. In summary, IL-6 functions during EAE depend on both the source and localization of immune response: the microglial IL-6 exerts both pathogenic and protective functions specifically in the CNS, whereas the dendritic cell-derived IL-6, in addition to being critically involved in the balance of regulatory T cells and TH17 cells, may stimulate production of cytokines associated with pathogenic functions of T cells.
Assuntos
Células Dendríticas , Modelos Animais de Doenças , Encefalomielite Autoimune Experimental , Interleucina-6 , Microglia , Esclerose Múltipla , Animais , Células Dendríticas/metabolismo , Células Dendríticas/imunologia , Camundongos , Interleucina-6/metabolismo , Esclerose Múltipla/imunologia , Esclerose Múltipla/metabolismo , Esclerose Múltipla/patologia , Microglia/metabolismo , Encefalomielite Autoimune Experimental/metabolismo , Encefalomielite Autoimune Experimental/imunologia , Encefalomielite Autoimune Experimental/patologia , Camundongos Endogâmicos C57BL , Receptor 1 de Quimiocina CX3C/metabolismo , Receptor 1 de Quimiocina CX3C/genética , Linfócitos T Reguladores/imunologia , Linfócitos T Reguladores/metabolismo , Receptores CCR6/metabolismo , Receptores CCR6/genética , FemininoRESUMO
OBJECTIVE: We aimed to describe the association between CX3CR1, CX3CL1, and ITGAV immunoexpression with PNI and adverse oncologic outcomes in patients with OSCC. STUDY DESIGN: Expression CX3CR1, CX3CL1, and ITGAV was assessed by immunohistochemistry in a cohort of 50 paraffin-embedded resections of OSCC. Survival analysis, Cox, and binary logistic regressions were undertaken to determine the impact on patient survival and predictive value for PNI. RESULTS: CX3CL1 positive nerves exhibited a significant association with tumor budding (TB) (P = .043), whereas nerves positive for ITGAV were associated with PNI (P = .021), T3-T4 tumor size (P = .029), and III-IV stage (P = .044). Cases with ITGAV-positive nerves exhibited an odds ratio of 9.603 (P = .008) for PNI, whereas cases with CX3CL1-positive nerves exhibited and odds ratio of 4.682 (P = .033) for TB. A trend toward decreased 5-year overall survival (P = .078) and 5-year disease-specific survival (P = .09) was observed in relation to ITGAV-positive nerves. However, no independent predictors for poor survival were identified. CONCLUSIONS: The expression of ITGAV was associated with PNI and advanced disease, whereas the expression of CX3CL1 was related to TB, suggesting that ITGAV and CX3CL1 are involved in their respective developments. Therefore, further investigations are encouraged to assess the potential utility of targeted therapies against CX3CL1 receptors in OSCC.
Assuntos
Biomarcadores Tumorais , Receptor 1 de Quimiocina CX3C , Carcinoma de Células Escamosas , Quimiocina CX3CL1 , Imuno-Histoquímica , Neoplasias Bucais , Adulto , Idoso , Idoso de 80 Anos ou mais , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Biomarcadores Tumorais/metabolismo , Carcinoma de Células Escamosas/metabolismo , Carcinoma de Células Escamosas/patologia , Quimiocina CX3CL1/metabolismo , Receptor 1 de Quimiocina CX3C/metabolismo , Neoplasias Bucais/patologia , Neoplasias Bucais/metabolismo , Invasividade Neoplásica , Estadiamento de Neoplasias , Valor Preditivo dos Testes , Prognóstico , Análise de SobrevidaRESUMO
Respiratory syncytial virus (RSV) is the most prevalent cause of acute lower respiratory infection in young children. Currently, the first RSV vaccines are approved by the FDA. Recently, N6-methyladenosine (m6A) RNA methylation has been implicated in the regulation of the viral life cycle and replication of many viruses, including RSV. m6A methylation of RSV RNA has been demonstrated to promote replication and prevent anti-viral immune responses by the host. Whether m6A is also involved in viral entry and whether m6A can also affect RSV infection via different mechanisms than methylation of viral RNA is poorly understood. Here, we identify m6A reader YTH domain-containing protein 1 (YTHDC1) as a novel negative regulator of RSV infection. We demonstrate that YTHDC1 abrogates RSV infection by reducing the expression of RSV entry receptor CX3C motif chemokine receptor 1 (CX3CR1) on the cell surface of lung epithelial cells. Altogether, these data reveal a novel role for m6A methylation and YTHDC1 in the viral entry of RSV. These findings may contribute to the development of novel treatment options to control RSV infection.
Assuntos
Receptor 1 de Quimiocina CX3C , Regulação para Baixo , Fatores de Processamento de RNA , Infecções por Vírus Respiratório Sincicial , Humanos , Células A549 , Adenosina/análogos & derivados , Adenosina/metabolismo , Linhagem Celular , Receptor 1 de Quimiocina CX3C/metabolismo , Receptor 1 de Quimiocina CX3C/genética , Células Epiteliais/virologia , Células Epiteliais/metabolismo , Interações Hospedeiro-Patógeno , Metilação , Proteínas do Tecido Nervoso , Infecções por Vírus Respiratório Sincicial/virologia , Infecções por Vírus Respiratório Sincicial/metabolismo , Vírus Sincicial Respiratório Humano/fisiologia , Fatores de Processamento de RNA/metabolismo , Fatores de Processamento de RNA/genética , RNA Viral/genética , RNA Viral/metabolismo , Internalização do Vírus , Replicação ViralRESUMO
Monocytes, as well as downstream macrophages and dendritic cells, are essential players in the immune system, fulfilling key roles in homeostasis as well as in inflammatory conditions. Conventionally, driven by studies on reporter models, mouse monocytes are categorized into a classical and a non-classical subset based on their inversely correlated surface expression of Ly6C/CCR2 and CX3CR1. Here, we aimed to challenge this concept by antibody staining and reporter mouse models. Therefore, we took advantage of Cx3cr1GFP and Ccr2RFP reporter mice, in which the respective gene was replaced by a fluorescent reporter protein gene. We analyzed the expression of CX3CR1 and CCR2 by flow cytometry using several validated fluorochrome-coupled antibodies and compared them with the reporter gene signal in these reporter mouse strains. Although we were able to validate the specificity of the fluorochrome-coupled flow cytometry antibodies, mouse Ly6Chigh classical and Ly6Clow non-classical monocytes showed no differences in CX3CR1 expression levels in the peripheral blood and spleen when stained with these antibodies. On the contrary, in Cx3cr1GFP reporter mice, we were able to reproduce the inverse correlation of the CX3CR1 reporter gene signal and Ly6C surface expression. Furthermore, differential CCR2 surface expression correlating with the expression of Ly6C was observed by antibody staining, but not in Ccr2RFP reporter mice. In conclusion, our data suggest that phenotyping strategies for mouse monocyte subsets should be carefully selected. In accordance with the literature, the suitability of CX3CR1 antibody staining is limited, whereas for CCR2, caution should be applied when using reporter mice.
Assuntos
Receptor 1 de Quimiocina CX3C , Citometria de Fluxo , Monócitos , Receptores CCR2 , Animais , Receptores CCR2/metabolismo , Receptores CCR2/genética , Monócitos/metabolismo , Receptor 1 de Quimiocina CX3C/metabolismo , Receptor 1 de Quimiocina CX3C/genética , Camundongos , Anticorpos/imunologia , Genes Reporter , Fenótipo , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Proteínas de Fluorescência Verde/metabolismo , Antígenos Ly/metabolismo , Antígenos Ly/genéticaRESUMO
Background: The crucial role of inflammation in aortic aneurysm (AA) is gaining prominence, while there is still a lack of key cytokines or targets for effective clinical translation. Methods: Mendelian randomization (MR) analysis was performed to identify the causal relationship between 91 circulating inflammatory proteins and AA and between 731 immune traits and AA. Bulk RNA sequencing data was utilized to demonstrate the expression profile of the paired ligand-receptor. Gene enrichment analysis, Immune infiltration, and correlation analysis were employed to deduce the potential role of CX3CR1. We used single-cell RNA sequencing data to pinpoint the localization of CX3CL1 and CX3CR1, which was further validated by multiplex immunofluorescence staining. Cellchat analysis was utilized to infer the CX3C signaling pathway. Trajectory analysis and the Cytosig database were exploited to determine the downstream effect of CX3CL1-CX3CR1. Results: We identified 4 candidates (FGF5, CX3CL1, IL20RA, and SCF) in multiple two-sample MR analyses. Subsequent analysis of the expression profile of the paired receptor revealed the significant upregulation of CX3CR1 in AA and its positive correlation with pro-inflammatory macrophages. Two sample MR between immune cell traits and AA demonstrated the potential causality between intermediate monocytes and AA. We finally deciphered in single-cell sequencing data that CX3CL1 sent by endothelial cells (ECs) acted on CX3CR1 of intermediated monocytes, leading to its recruitment and pro-inflammatory responses. Conclusion: Our study presented a genetic insight into the pathogenetic role of CX3CL1-CX3CR1 in AA, and further deciphered the CX3C signaling pathway between ECs and intermediate monocytes.
Assuntos
Aneurisma Aórtico , Receptor 1 de Quimiocina CX3C , Quimiocina CX3CL1 , Análise da Randomização Mendeliana , Receptor 1 de Quimiocina CX3C/genética , Receptor 1 de Quimiocina CX3C/metabolismo , Humanos , Quimiocina CX3CL1/genética , Quimiocina CX3CL1/metabolismo , Aneurisma Aórtico/genética , Aneurisma Aórtico/metabolismo , Perfilação da Expressão Gênica , Transcriptoma , Transdução de Sinais , Predisposição Genética para DoençaRESUMO
Identifying tumor-relevant T cell subsets in the peripheral blood (PB) has become a potential strategy for cancer treatment. However, the subset of PB that could be used to treat cancer remains poorly defined. Here, we found that the CX3CR1+ T cell subset in the blood of patients with lung cancer exhibited effector properties and had a higher TCR matching ratio with tumor-infiltrating lymphocytes (TILs) compared to CX3CR1- T cells, as determined by paired single-cell RNA and TCR sequencing. Meanwhile, the anti-tumor activities, effector cytokine production, and mitochondrial function were enhanced in CX3CR1+ T cells both in vitro and in vivo. However, in the co-culture system of H322 cells with T cells, the percentages of apoptotic cells and Fas were substantially higher in CX3CR1+ T cells than those in CX3CR1- T cells. Fas-mediated apoptosis was rescued by treatment with an anti-PD-1 antibody. Accordingly, the combination of adoptive transfer of CX3CR1+ T cells and anti-PD-1 treatment considerably decreased Fas expression and improved the survival of lung xenograft mice. Moreover, an increased frequency of CX3CR1+ T cells in the PB correlated with a better response and prolonged survival of patients with lung cancer who received anti-PD-1 therapy. These findings indicate the promising potential of adoptive transfer of peripheral CX3CR1+ T cells as an individual cancer immunotherapy.
Assuntos
Receptor 1 de Quimiocina CX3C , Inibidores de Checkpoint Imunológico , Neoplasias Pulmonares , Linfócitos do Interstício Tumoral , Receptor de Morte Celular Programada 1 , Animais , Neoplasias Pulmonares/tratamento farmacológico , Neoplasias Pulmonares/imunologia , Neoplasias Pulmonares/patologia , Receptor 1 de Quimiocina CX3C/metabolismo , Humanos , Camundongos , Inibidores de Checkpoint Imunológico/farmacologia , Inibidores de Checkpoint Imunológico/uso terapêutico , Receptor de Morte Celular Programada 1/antagonistas & inibidores , Receptor de Morte Celular Programada 1/metabolismo , Linfócitos do Interstício Tumoral/imunologia , Linfócitos do Interstício Tumoral/efeitos dos fármacos , Linfócitos do Interstício Tumoral/metabolismo , Ensaios Antitumorais Modelo de Xenoenxerto , Linhagem Celular Tumoral , Feminino , Apoptose/efeitos dos fármacos , Masculino , Subpopulações de Linfócitos T/imunologia , Subpopulações de Linfócitos T/efeitos dos fármacos , Subpopulações de Linfócitos T/metabolismo , Linfócitos T/imunologia , Linfócitos T/efeitos dos fármacos , Linfócitos T/metabolismoRESUMO
The chemotactic cytokine fractalkine (FKN, chemokine CX3CL1) has unique properties resulting from the combination of chemoattractants and adhesion molecules. The soluble form (sFKN) has chemotactic properties and strongly attracts T cells and monocytes. The membrane-bound form (mFKN) facilitates diapedesis and is responsible for cell-to-cell adhesion, especially by promoting the strong adhesion of leukocytes (monocytes) to activated endothelial cells with the subsequent formation of an extracellular matrix and angiogenesis. FKN signaling occurs via CX3CR1, which is the only known member of the CX3C chemokine receptor subfamily. Signaling within the FKN-CX3CR1 axis plays an important role in many processes related to inflammation and the immune response, which often occur simultaneously and overlap. FKN is strongly upregulated by hypoxia and/or inflammation-induced inflammatory cytokine release, and it may act locally as a key angiogenic factor in the highly hypoxic tumor microenvironment. The importance of the FKN/CX3CR1 signaling pathway in tumorigenesis and cancer metastasis results from its influence on cell adhesion, apoptosis, and cell migration. This review presents the role of the FKN signaling pathway in the context of angiogenesis in inflammation and cancer. The mechanisms determining the pro- or anti-tumor effects are presented, which are the cause of the seemingly contradictory results that create confusion regarding the therapeutic goals.
Assuntos
Receptor 1 de Quimiocina CX3C , Carcinogênese , Quimiocina CX3CL1 , Inflamação , Neovascularização Patológica , Transdução de Sinais , Humanos , Quimiocina CX3CL1/metabolismo , Neovascularização Patológica/metabolismo , Inflamação/metabolismo , Inflamação/patologia , Receptor 1 de Quimiocina CX3C/metabolismo , Receptor 1 de Quimiocina CX3C/genética , Animais , Carcinogênese/metabolismo , Carcinogênese/patologia , Neoplasias/metabolismo , Neoplasias/patologia , Neoplasias/etiologia , Microambiente Tumoral , AngiogêneseRESUMO
BACKGROUND: Cardiac damage induced by ischemic stroke, such as arrhythmia, cardiac dysfunction, and even cardiac arrest, is referred to as cerebral-cardiac syndrome (CCS). Cardiac macrophages are reported to be closely associated with stroke-induced cardiac damage. However, the role of macrophage subsets in CCS is still unclear due to their heterogeneity. Sympathetic nerves play a significant role in regulating macrophages in cardiovascular disease. However, the role of macrophage subsets and sympathetic nerves in CCS is still unclear. METHODS AND RESULTS: In this study, a middle cerebral artery occlusion mouse model was used to simulate ischemic stroke. ECG and echocardiography were used to assess cardiac function. We used Cx3cr1GFPCcr2RFP mice and NLRP3-deficient mice in combination with Smart-seq2 RNA sequencing to confirm the role of macrophage subsets in CCS. We demonstrated that ischemic stroke-induced cardiac damage is characterized by severe cardiac dysfunction and robust infiltration of monocyte-derived macrophages into the heart. Subsequently, we identified that cardiac monocyte-derived macrophages displayed a proinflammatory profile. We also observed that cardiac dysfunction was rescued in ischemic stroke mice by blocking macrophage infiltration using a CCR2 antagonist and NLRP3-deficient mice. In addition, a cardiac sympathetic nerve retrograde tracer and a sympathectomy method were used to explore the relationship between sympathetic nerves and cardiac macrophages. We found that cardiac sympathetic nerves are significantly activated after ischemic stroke, which contributes to the infiltration of monocyte-derived macrophages and subsequent cardiac dysfunction. CONCLUSIONS: Our findings suggest a potential pathogenesis of CCS involving the cardiac sympathetic nerve-monocyte-derived macrophage axis.