RESUMO
Background: Chemotactic cytokines play a crucial role in the development of acute myeloid leukemia (AML). Thus, investigating the mechanisms of chemotactic cytokine-related genes (CCRGs) in AML is of paramount importance. Methods: Using the TCGA-AML, GSE114868, and GSE12417 datasets, differential expression analysis identified differentially expressed CCRGs (DE-CCRGs). These genes were screened by overlapping differentially expressed genes (DEGs) between AML and control groups with CCRGs. Subsequently, functional enrichment analysis and the construction of a protein-protein interaction (PPI) network were conducted to explore the functions of the DE-CCRGs. Univariate Cox regression, least absolute shrinkage and selection operator (LASSO), and multivariate Cox regression analyses identified relevant prognostic genes and developed a prognostic model. Survival analysis of the prognostic gene was performed, followed by functional similarity analysis, immune analysis, enrichment analysis, and drug prediction analysis. Results: Differential expression analysis revealed 6,743 DEGs, of which 29 DE-CCRGs were selected for this study. Functional enrichment analysis indicated that DE-CCRGs were primarily involved in chemotactic cytokine-related functions and pathways. Six prognostic genes (CXCR3, CXCR2, CXCR6, CCL20, CCL4, and CCR2) were identified and incorporated into the risk model. The model's performance was validated using the GSE12417 dataset. Survival analysis showed significant differences in AML overall survival (OS) between prognostic gene high and low expression groups, indicating that prognostic gene might be significantly associated with patient survival. Additionally, nine different immune cells were identified between the two risk groups. Correlation analysis revealed that CCR2 had the most significant positive correlation with monocytes and the most significant negative correlation with resting mast cells. The tumor immune dysfunction and exclusion score was lower in the high-risk group. Conclusion: CXCR3, CXCR2, CXCR6, CCL20, CCL4, and CCR2 were identified as prognostic genes correlated to AML and the tumor immune microenvironment. These findings offerred novel insights into the prevention and treatment of AML.
Assuntos
Leucemia Mieloide Aguda , Mapas de Interação de Proteínas , Receptores CCR2 , Receptores de Interleucina-8B , Humanos , Leucemia Mieloide Aguda/genética , Leucemia Mieloide Aguda/mortalidade , Prognóstico , Receptores de Interleucina-8B/genética , Receptores CCR2/genética , Mapas de Interação de Proteínas/genética , Quimiocina CCL4/genética , Quimiocina CCL20/genética , Quimiocina CCL20/metabolismo , Feminino , Masculino , Quimiocinas/genética , Perfilação da Expressão Gênica , Pessoa de Meia-Idade , Biomarcadores Tumorais/genética , Receptores CXCR3RESUMO
BACKGROUND: Chronic obstructive pulmonary disease (COPD) remains a leading cause of morbidity and mortality worldwide, characterized by persistent respiratory symptoms and airflow limitation. The involvement of C-C motif chemokine ligand 2 (CCL2) in COPD pathogenesis, particularly in macrophage regulation and activation, is poorly understood despite its recognized role in chronic inflammation. Our study aims to elucidate the regulatory role and molecular mechanisms of CCL2 in the pathogenesis of COPD, providing new insights for therapeutic strategies. METHODS: This study focused on the CCL2-CCR2 signaling pathway, exploring its role in COPD pathogenesis using both Ccl2 knockout (KO) mice and pharmacological inhibitors. To dissect the underlying mechanisms, we employed various in vitro and in vivo methods to analyze the secretion patterns and pathogenic effects of CCL2 and its downstream molecular signaling through the CCL2-CCR2 axis. RESULTS: Elevated Ccl2 expression was confirmed in the lungs of COPD mice and was associated with enhanced recruitment and activation of macrophages. Deletion of Ccl2 in knockout mice, as well as treatment with a Ccr2 inhibitor, resulted in protection against CS- and LPS-induced alveolar injury and airway remodeling. Mechanistically, CCL2 was predominantly secreted by bronchial epithelial cells in a process dependent on STAT1 phosphorylation and acted through the CCR2 receptor on macrophages. This interaction activated the PI3K-AKT signaling pathway, which was pivotal for macrophage activation and the secretion of inflammatory cytokines, further influencing the progression of COPD. CONCLUSIONS: The study highlighted the crucial role of CCL2 in mediating inflammatory responses and remodeling in COPD. It enhanced our understanding of COPD's molecular mechanisms, particularly how CCL2's interaction with the CCR2 activates critical signaling pathways. Targeting the CCL2-CCR2 axis emerged as a promising strategy to alleviate COPD pathology.
Assuntos
Quimiocina CCL2 , Macrófagos , Camundongos Knockout , Fosfatidilinositol 3-Quinases , Proteínas Proto-Oncogênicas c-akt , Doença Pulmonar Obstrutiva Crônica , Receptores CCR2 , Transdução de Sinais , Doença Pulmonar Obstrutiva Crônica/metabolismo , Doença Pulmonar Obstrutiva Crônica/patologia , Animais , Quimiocina CCL2/metabolismo , Quimiocina CCL2/genética , Receptores CCR2/metabolismo , Receptores CCR2/genética , Proteínas Proto-Oncogênicas c-akt/metabolismo , Transdução de Sinais/efeitos dos fármacos , Fosfatidilinositol 3-Quinases/metabolismo , Macrófagos/metabolismo , Macrófagos/efeitos dos fármacos , Camundongos , Humanos , Camundongos Endogâmicos C57BL , MasculinoAssuntos
Modelos Animais de Doenças , Macrófagos , NF-kappa B , Receptores CCR2 , Transdução de Sinais , Animais , Camundongos , Macrófagos/metabolismo , Macrófagos/patologia , Receptores CCR2/metabolismo , Receptores CCR2/genética , NF-kappa B/metabolismo , NF-kappa B/genética , Miocárdio/metabolismo , Miocárdio/patologia , Humanos , Displasia Arritmogênica Ventricular Direita/genética , Displasia Arritmogênica Ventricular Direita/metabolismo , Displasia Arritmogênica Ventricular Direita/patologia , Camundongos KnockoutRESUMO
The ligands of chemokine receptors 2 and 5 (CCR2 and CCR5, respectively) are associated with the pathomechanism of neuropathic pain development, but their role in painful diabetic neuropathy remains unclear. Therefore, the aim of our study was to examine the function of these factors in the hypersensitivity accompanying diabetes. Additionally, we analyzed the analgesic effect of cenicriviroc (CVC), a dual CCR2/CCR5 antagonist, and its influence on the effectiveness of morphine. An increasing number of experimental studies have shown that targeting more than one molecular target is advantageous compared with the coadministration of individual pharmacophores in terms of their analgesic effect. The advantage of using bifunctional compounds is that they gain simultaneous access to two receptors at the same dose, positively affecting their pharmacokinetics and pharmacodynamics and consequently leading to improved analgesia. Experiments were performed on male and female Swiss albino mice with a streptozotocin (STZ, 200 mg/kg, i.p.) model of diabetic neuropathy. We found that the blood glucose level increased, and the mechanical and thermal hypersensitivity developed on the 7th day after STZ administration. In male mice, we observed increased mRNA levels of Ccl2, Ccl5, and Ccl7, while in female mice, we observed additional increases in Ccl8 and Ccl12 levels. We have demonstrated for the first time that a single administration of cenicriviroc relieves pain to a similar extent in male and female mice. Moreover, repeated coadministration of cenicriviroc with morphine delays the development of opioid tolerance, while the best and longest-lasting analgesic effect is achieved by repeated administration of cenicriviroc alone, which reduces pain hypersensitivity in STZ-exposed mice, and unlike morphine, no tolerance to the analgesic effects of CVC is observed until Day 15 of treatment. Based on these results, we suggest that targeting CCR2 and CCR5 with CVC is a potent therapeutic option for novel pain treatments in diabetic neuropathy patients.
Assuntos
Antagonistas dos Receptores CCR5 , Neuropatias Diabéticas , Modelos Animais de Doenças , Receptores CCR2 , Receptores CCR5 , Animais , Camundongos , Neuropatias Diabéticas/tratamento farmacológico , Masculino , Receptores CCR2/antagonistas & inibidores , Receptores CCR2/metabolismo , Feminino , Receptores CCR5/metabolismo , Receptores CCR5/genética , Antagonistas dos Receptores CCR5/farmacologia , Antagonistas dos Receptores CCR5/uso terapêutico , Morfina/farmacologia , Morfina/uso terapêutico , Diabetes Mellitus Experimental/tratamento farmacológico , Diabetes Mellitus Experimental/complicações , Analgésicos/farmacologia , Analgésicos/uso terapêutico , Hiperalgesia/tratamento farmacológico , Imidazóis , SulfóxidosRESUMO
BACKGROUND: Macrophages are key players in obesity-associated cardiovascular diseases, which are marked by inflammatory and immune alterations. However, the pathophysiological mechanisms underlying macrophage's role in obesity-induced cardiac inflammation are incompletely understood. Our study aimed to identify the key macrophage population involved in obesity-induced cardiac dysfunction and investigate the molecular mechanism that contributes to the inflammatory response. METHODS: In this study, we used single-cell RNA-sequencing analysis of Cd45+CD11b+F4/80+ cardiac macrophages to explore the heterogeneity of cardiac macrophages. The CCR2+ (C-C chemokine receptor 2) macrophages were specifically removed by a dual recombinase approach, and the macrophage CCR2 was deleted to investigate their functions. We also performed cleavage under target and tagmentation analysis, chromatin immunoprecipitation-polymerase chain reaction, luciferase assay, and macrophage-specific lentivirus transfection to define the impact of lysozyme C in macrophages on obesity-induced inflammation. RESULTS: We find that the Ccr2 cluster undergoes a functional transition from homeostatic maintenance to proinflammation. Our data highlight specific changes in macrophage behavior during cardiac dysfunction under metabolic challenge. Consistently, inducible ablation of CCR2+CX3CR1+ macrophages or selective deletion of macrophage CCR2 prevents obesity-induced cardiac dysfunction. At the mechanistic level, we demonstrate that the obesity-induced functional shift of CCR2-expressing macrophages is mediated by the CCR2/activating transcription factor 3/lysozyme 1/NF-κB (nuclear factor kappa B) signaling. Finally, we uncover a noncanonical role for lysozyme 1 as a transcription activator, binding to the RelA promoter, driving NF-κB signaling, and strongly promoting inflammation and cardiac dysfunction in obesity. CONCLUSIONS: Our findings suggest that lysozyme 1 may represent a potential target for the diagnosis of obesity-induced inflammation and the treatment of obesity-induced heart disease.
Assuntos
Macrófagos , Muramidase , Obesidade , Receptores CCR2 , Animais , Obesidade/complicações , Obesidade/metabolismo , Macrófagos/metabolismo , Receptores CCR2/metabolismo , Receptores CCR2/genética , Camundongos , Muramidase/metabolismo , Muramidase/genética , Camundongos Endogâmicos C57BL , Masculino , Camundongos Knockout , Transdução de Sinais , Inflamação/metabolismo , Inflamação/genética , Cardiopatias/etiologia , Cardiopatias/metabolismo , Cardiopatias/genéticaRESUMO
Osteoarthritis (OA) is characterized by a complex interplay of molecular signals orchestrated by the CCL2/CCR2 axis. The pathogenesis of OA has been revealed to be influenced by a multifaceted effect of CCL2/CCR2 signaling on inflammation, cartilage degradation, and joint homeostasis. The CCL2/CCR2 axis promotes immune cell recruitment and tips the balance toward degeneration by influencing chondrocyte behavior. Insights into these intricate pathways will offer novel therapeutic approaches, paving the way for targeted interventions that may redefine OA management in the future. This review article explores the molecular symphony through the lens of the CCL2/CCR2 axis, providing a harmonious blend of current knowledge and future directions on OA treatment. Furthermore, in this study, through a meticulous review of recent research, the key players and molecular mechanisms that amplify the catabolic cascade within the joint microenvironment are identified, and therapeutic approaches to targeting the CCL2/CCR axis are discussed.
Assuntos
Quimiocina CCL2 , Osteoartrite , Receptores CCR2 , Transdução de Sinais , Humanos , Quimiocina CCL2/metabolismo , Receptores CCR2/metabolismo , Osteoartrite/metabolismo , Osteoartrite/imunologia , Osteoartrite/terapia , Animais , Condrócitos/metabolismo , Condrócitos/imunologia , Cartilagem Articular/metabolismo , Cartilagem Articular/patologia , Cartilagem Articular/imunologia , Terapia de Alvo MolecularRESUMO
Benign Prostatic Hyperplasia (BPH) is a complex condition leading to Lower Urinary Tract Symptoms in aging men, characterized by cellular proliferation, smooth muscle dysfunction, inflammation, and fibrosis. While BPH is known to involve heightened macrophage infiltration, the specific contribution of infiltrating monocytes/macrophages to the disease mechanism remains uncertain. This research explores the impact of reducing circulating monocytes and subsequently limiting their tissue infiltration by using Ccr2 knockout (Ccr2-KO) mice. Ccr2-KO and wild type mice were implanted with testosterone and estradiol (T + E2, 25 mg + 2.5 mg) pellets. Urinary function was assessed via weekly void spot assays over 12 weeks, and prostatic macrophage levels were visualized and quantified in tissue sections using an F4/80 antibody. Additionally, Ki-67 staining was used to evaluate cell proliferation, and picrosirius red staining to assess collagen accumulation. Increased voiding frequency which developed in T + E2 mice, was significantly ameliorated in Ccr2-KO mice, however, both Ccr2-KO and wild type (WT) mice showed increased bladder weights after three month, representing a hypertrophic response to bladder outlet obstruction. T + E2 substantially increased the density of macrophages in WT but not Ccr2-KO mouse prostate. Proliferation rate, as indicated by Ki-67 positivity, was elevated in the vental and anterior prostate lobes but was only marginally reduced in Ccr2-KO mice. Most importantly, a significant prostatic collagen accumulation was observed in WT mice that was markedly reduced by Ccr2 deficiency post T + E2 treatment. The absence of Ccr2 mitigates urinary dysfunction and alters prostatic macrophage levels and collagen accumulation in steroid hormone imbalance. These findings suggest a crucial role for monocyte infiltration, giving rise to macrophages or other cell derivatives, to drive fibrosis.
Assuntos
Estradiol , Fibrose , Macrófagos , Camundongos Knockout , Monócitos , Próstata , Receptores CCR2 , Testosterona , Animais , Masculino , Receptores CCR2/metabolismo , Macrófagos/metabolismo , Camundongos , Monócitos/metabolismo , Próstata/metabolismo , Próstata/patologia , Testosterona/metabolismo , Estradiol/metabolismo , Estradiol/farmacologia , Hiperplasia Prostática/metabolismo , Hiperplasia Prostática/patologia , Proliferação de Células , Camundongos Endogâmicos C57BLRESUMO
Targeting the CCL2/CCR2 chemokine axis has been shown to be effective at relieving pain in rodent models of inflammatory and neuropathic pain, therefore representing a promising avenue for the development of non-opioid analgesics. However, clinical trials targeting this receptor for inflammatory conditions and painful neuropathies have failed to meet expectations and have all been discontinued due to lack of efficacy. To overcome the poor selectivity of CCR2 chemokine receptor antagonists, we generated and characterized the function of intracellular cell-penetrating allosteric modulators targeting CCR2, namely pepducins. In vivo, chronic intrathecal administration of the CCR2-selective pepducin PP101 was effective in alleviating neuropathic and bone cancer pain. In the setting of bone metastases, we found that T cells infiltrate dorsal root ganglia (DRG) and induce long-lasting pain hypersensitivity. By acting on CCR2-expressing DRG neurons, PP101 attenuated the altered phenotype of sensory neurons as well as the neuroinflammatory milieu of DRGs, and reduced bone cancer pain by blocking CD4+ and CD8+ T cell infiltration. Notably, PP101 demonstrated its efficacy in targeting the neuropathic component of bone cancer pain, as evidenced by its anti-nociceptive effects in a model of chronic constriction injury of the sciatic nerve. Importantly, PP101-induced reduction of CCR2 signaling in DRGs did not result in deleterious tumor progression or adverse behavioral effects. Thus, targeting neuroimmune crosstalk through allosteric inhibition of CCR2 could represent an effective and safe avenue for the management of chronic pain.
Assuntos
Dor Crônica , Gânglios Espinais , Neuralgia , Receptores CCR2 , Animais , Receptores CCR2/antagonistas & inibidores , Receptores CCR2/metabolismo , Dor Crônica/tratamento farmacológico , Gânglios Espinais/efeitos dos fármacos , Gânglios Espinais/metabolismo , Neuralgia/tratamento farmacológico , Neuralgia/metabolismo , Humanos , Dor do Câncer/tratamento farmacológico , Neoplasias Ósseas/tratamento farmacológico , Neoplasias Ósseas/secundário , Analgésicos/farmacologia , Analgésicos/uso terapêutico , Masculino , Camundongos , Feminino , Camundongos Endogâmicos C57BLRESUMO
The interactions between chemokines and their receptors, particularly in the context of inflammation, are complex, with individual receptors binding multiple ligands and individual ligands interacting with multiple receptors. In addition, there are numerous reports of simultaneous coexpression of multiple inflammatory chemokine receptors on individual inflammatory leukocyte subtypes. Overall, this has previously been interpreted as redundancy and proposed as a protective mechanism to ensure that the inflammatory response is robust. By contrast, we have hypothesized that the system is not redundant but exquisitely subtle. Our interests relate to the receptors CCR1, CCR2, CCR3, and CCR5, which, together, regulate nonneutrophilic myeloid cell recruitment to inflammatory sites. In this study, we demonstrate that although most murine monocytes exclusively express CCR2, there is a small subpopulation that is expanded during inflammation and coexpresses CCR1 and CCR2. Combinations of transcript and functional analysis demonstrate that this is not redundant expression and that coexpression of CCR1 and CCR2 marks a phenotypically distinct population of monocytes characterized by expression of genes otherwise typically associated with neutrophils. Single-cell RNA sequencing confirms this as a monodisperse population of atypical monocytes. This monocytic population has previously been described as having immunosuppressive activity. Overall, our data confirm combinatorial chemokine receptor expression by a subpopulation of monocytes but demonstrate that this is not redundant expression and marks a discrete monocytic population.
Assuntos
Monócitos , Receptores CCR1 , Receptores CCR2 , Receptores CCR1/genética , Receptores CCR1/metabolismo , Receptores CCR2/genética , Receptores CCR2/metabolismo , Monócitos/imunologia , Monócitos/metabolismo , Animais , Camundongos , Camundongos Endogâmicos C57BL , Inflamação/imunologiaRESUMO
The inhibition of hepatic macrophage and Kupfer cell recruitment and activation is a potential strategy for treating insulin resistance and nonalcoholic steatohepatitis (NASH). Cenicriviroc (CVC), a dual C-C chemokine receptor 2 (CCR2) and CCR5 antagonist, has shown antifibrotic activity in murine models of NASH and has been evaluated in clinical trials on patients with NASH. This study investigated the effects of CVC on macrophage infiltration and polarization in a lipotoxic model of NASH. C57BL/6 mice were fed a high-cholesterol, high-fat (CL) diet or a CL diet containing 0.015% CVC (CL + CVC) for 12 weeks. Macrophage recruitment and activation were assayed by immunohistochemistry and flow cytometry. CVC supplementation attenuated excessive hepatic lipid accumulation and peroxidation and alleviated glucose intolerance and hyperinsulinemia in the mice that were fed the CL diet. Flow cytometry analysis revealed that compared with the CL group, mice fed the CL + CVC diet had fewer M1-like macrophages, more M2-like macrophages, and fewer T cell counts, indicating that CVC caused an M2-dominant shift of macrophages in the liver. Similarly, CVC decreased lipopolysaccharide-stimulated M1-like macrophage activation, whereas it increased interleukin-4-induced M2-type macrophage polarization in vitro. In addition, CVC attenuated hepatic fibrosis by repressing hepatic stellate cell activation. Lastly, CVC reversed insulin resistance as well as steatosis, inflammation, and fibrosis of the liver in mice with pre-existing NASH. In conclusion, CVC prevented and reversed hepatic steatosis, insulin resistance, inflammation, and fibrogenesis in the liver of NASH mice via M2 macrophage polarization.
Assuntos
Fígado , Macrófagos , Camundongos Endogâmicos C57BL , Hepatopatia Gordurosa não Alcoólica , Animais , Macrófagos/efeitos dos fármacos , Macrófagos/metabolismo , Camundongos , Hepatopatia Gordurosa não Alcoólica/tratamento farmacológico , Hepatopatia Gordurosa não Alcoólica/patologia , Masculino , Fígado/efeitos dos fármacos , Fígado/patologia , Fígado/metabolismo , Dieta Hiperlipídica/efeitos adversos , Receptores CCR2/metabolismo , Sulfóxidos/farmacologia , Ativação de Macrófagos/efeitos dos fármacos , Antagonistas dos Receptores CCR5/farmacologia , Antagonistas dos Receptores CCR5/uso terapêutico , Resistência à Insulina , ImidazóisRESUMO
Peripheral nerves exhibit long-term residual motor dysfunction following injury. The length of the denervation period before nerve and muscle reconnection is an important factor in motor function recovery. We aimed to investigate whether repeated nerve crush injuries to the same site every 7 days would preserve the conditioning lesion (CL) response and to determine the number of nerve crush injuries required to create an experimental animal model that would prolong the denervation period while maintaining peripheral nerve continuity. Rats were grouped according to the number of sciatic nerve crushes. A significant decrease in the soleus muscle fiber cross-sectional area was observed with increased crushes. After a single crush, macrophage accumulation and macrophage chemotaxis factor CCL2 expression in dorsal root ganglia were markedly increased, which aligned with the gene expression of Ccl2 and its receptor Ccr2. Macrophage numbers, histological CCL2 expression, and Ccl2 and Ccr2 gene expression levels decreased, depending on the number of repeated crushes. Histological analysis and gene expression analysis in the group with four repeated crushes did not differ significantly when compared with uninjured animals. Our findings indicated that repeated nerve crushes at the same site every 7 days sustained innervation loss and caused a loss of the CL response. The experimental model did not require nerve stump suturing and is useful for exploring factors causing prolonged denervation-induced motor dysfunction. SIGNIFICANCE STATEMENT: This study elucidates the effects of repeated nerve crush injury to the same site on innervation and conditioning lesion responses and demonstrates the utility of an experimental animal model that recapitulates the persistent residual motor deficits owing to prolonged denervation without requiring nerve transection and transection suturing.
Assuntos
Quimiocina CCL2 , Modelos Animais de Doenças , Compressão Nervosa , Nervo Isquiático , Animais , Nervo Isquiático/lesões , Masculino , Compressão Nervosa/métodos , Quimiocina CCL2/metabolismo , Quimiocina CCL2/genética , Músculo Esquelético/inervação , Músculo Esquelético/metabolismo , Gânglios Espinais/metabolismo , Ratos , Receptores CCR2/metabolismo , Receptores CCR2/genética , Macrófagos/metabolismo , Traumatismos dos Nervos Periféricos/metabolismo , Traumatismos dos Nervos Periféricos/fisiopatologia , Ratos Sprague-Dawley , Denervação/métodos , Regeneração Nervosa/fisiologia , Neuropatia Ciática/patologia , Neuropatia Ciática/fisiopatologiaRESUMO
Low back pain due to epidural fibrosis is a major complication after spine surgery. Macrophages infiltrate the wound area post laminectomy, but the role of macrophages in epidural fibrosis remains largely elusive. In a mouse model of laminectomy, macrophage depletion decreased epidural fibrosis. CD146, an adhesion molecule involved in cell migration, is expressed by macrophages. CD146-defective macrophages exhibited impaired migration, which was mediated by reduced expression of CCR2 and suppression of the MAPK/ERK signaling pathway. CD146-defective macrophages suppress the MAPK/ERK signaling pathway by increasing Erdr1. In vivo, CD146 deficiency decreased macrophage infiltration and reduced extracellular matrix deposition in wound tissues. Moreover, the anti-CD146 antibody AA98 suppressed macrophage infiltration and epidural fibrosis. Taken together, these findings demonstrated that CD146 deficiency alleviates epidural fibrosis by decreasing the migration of macrophages via the Erdr1/ERK/CCR2 pathway. Blocking CD146 and macrophage infiltration may help alleviate epidural fibrosis.
Assuntos
Antígeno CD146 , Fibrose , Macrófagos , Camundongos Endogâmicos C57BL , Receptores CCR2 , Animais , Receptores CCR2/metabolismo , Receptores CCR2/genética , Macrófagos/imunologia , Macrófagos/metabolismo , Camundongos , Antígeno CD146/metabolismo , Antígeno CD146/genética , Movimento Celular , Camundongos Knockout , Espaço Epidural/patologia , Masculino , Sistema de Sinalização das MAP Quinases/imunologia , Laminectomia , Modelos Animais de Doenças , Transdução de Sinais , HumanosRESUMO
Abdominal adhesion, a serious complication of abdominal surgery, often resists mitigation by current drug administration and physical barriers. To address this issue, we developed an injectable, antifouling hydrogel through the free-radical polymerization of methacrylate chondroitin sulfate (CS-GMA) and 2-methacryloyloxyethyl phosphorylcholine (MPC) monomers, dubbed the CGM hydrogel. We systematically analyzed its physicochemical properties, including rheological strength, biocompatibility, and antifouling capabilities. A rat abdominal cecum adhesion model was constructed to assess the effectiveness of CGM hydrogel in preventing postoperative adhesion and recurrent adhesion. In addition, multi-omics analyses identified the relationship between adhesion development and CCL2/CCR2 interaction. Notably, CGM hydrogel can thwart the recruitment and aggregation of fibroblasts and macrophages by inhibiting the CCL2/CCR2 interaction. Moreover, CGM hydrogel significantly dampens the activity of fibrosis-linked cytokines (TGF-ßR1) and recalibrates extracellular matrix deposition-related cytokines (t-PA and PAI-1, Col â and MMP-9). Cumulatively, the dual action of CGM hydrogel-as a physical barrier and cytokine regulator-highlights its promising potential in clinical application for abdominal adhesion prevention.
Assuntos
Quimiocina CCL2 , Hidrogéis , Ratos Sprague-Dawley , Receptores CCR2 , Animais , Aderências Teciduais/prevenção & controle , Aderências Teciduais/metabolismo , Hidrogéis/química , Hidrogéis/farmacologia , Quimiocina CCL2/metabolismo , Ratos , Receptores CCR2/metabolismo , Fosforilcolina/análogos & derivados , Fosforilcolina/química , Fosforilcolina/farmacologia , Metacrilatos/química , Metacrilatos/farmacologia , Incrustação Biológica/prevenção & controle , Sulfatos de Condroitina/química , Sulfatos de Condroitina/farmacologia , Camundongos , Abdome/cirurgia , Injeções , Masculino , Macrófagos/metabolismo , Macrófagos/efeitos dos fármacosRESUMO
Cardiovascular diseases are an array of age-related disorders, and accumulating evidence suggests a link between cardiac resident macrophages (CRMs) and the age-related disorders. However, how does CRMs alter with aging remains elusive. In the present study, aged mice (20 months old) have been employed to check for their cardiac structural and functional alterations, and the changes in the proportion of CRM subsets as well, followed by sorting of CRMs, including C-C Motif Chemokine Receptor 2 (CCR2)+ and CCR2- CRMs, which were subjected to Smart-Seq. Integrated analysis of the Smart-Seq data with three publicly available single-cell RNA-seq datasets revealed that inflammatory genes were drastic upregulated for both CCR2+ and CCR2- CRMs with aging, but genes germane to wound healing were downregulated for CCR2- CRMs, suggesting the differential functions of these two subsets. More importantly, inflammatory genes involved in damage sensing, complement cascades, and phagocytosis were largely upregulated in CCR2- CRMs, implying the imbalance of inflammatory response upon aging. Our work provides a comprehensive framework and transcriptional resource for assessing the impact of aging on CRMs with a potential for further understanding cardiac aging.
Assuntos
Envelhecimento , Perfilação da Expressão Gênica , Macrófagos , Camundongos Endogâmicos C57BL , Receptores CCR2 , Animais , Macrófagos/metabolismo , Envelhecimento/genética , Envelhecimento/metabolismo , Camundongos , Receptores CCR2/metabolismo , Receptores CCR2/genética , Transcriptoma , Miocárdio/metabolismo , Masculino , Análise de Célula Única , Inflamação/genética , Inflamação/metabolismo , Inflamação/patologia , Transdução de Sinais , FagocitoseRESUMO
BACKGROUND: Since the 1990s, evidence has accumulated that macrophages promote peripheral nerve regeneration and are required for enhancing regeneration in the conditioning lesion (CL) response. After a sciatic nerve injury, macrophages accumulate in the injury site, the nerve distal to that site, and the axotomized dorsal root ganglia (DRGs). In the peripheral nervous system, as in other tissues, the macrophage response is derived from both resident macrophages and recruited monocyte-derived macrophages (MDMs). Unresolved questions are: at which sites do macrophages enhance nerve regeneration, and is a particular population needed. METHODS: Ccr2 knock-out (KO) and Ccr2gfp/gfp knock-in/KO mice were used to prevent MDM recruitment. Using these strains in a sciatic CL paradigm, we examined the necessity of MDMs and residents for CL-enhanced regeneration in vivo and characterized injury-induced nerve inflammation. CL paradigm variants, including the addition of pharmacological macrophage depletion methods, tested the role of various macrophage populations in initiating or sustaining the CL response. In vivo regeneration, measured from bilateral proximal test lesions (TLs) after 2 d, and macrophages were quantified by immunofluorescent staining. RESULTS: Peripheral CL-enhanced regeneration was equivalent between crush and transection CLs and was sustained for 28 days in both Ccr2 KO and WT mice despite MDM depletion. Similarly, the central CL response measured in dorsal roots was unchanged in Ccr2 KO mice. Macrophages at both the TL and CL, but not between them, stained for the pro-regenerative marker, arginase 1. TL macrophages were primarily CCR2-dependent MDMs and nearly absent in Ccr2 KO and Ccr2gfp/gfp KO mice. However, there were only slightly fewer Arg1+ macrophages in CCR2 null CLs than controls due to resident macrophage compensation. Zymosan injection into an intact WT sciatic nerve recruited Arg1+ macrophages but did not enhance regeneration. Finally, clodronate injection into Ccr2gfp KO CLs dramatically reduced CL macrophages. Combined with the Ccr2gfp KO background, depleting MDMs and TL macrophages, and a transection CL, physically removing the distal nerve environment, nearly all macrophages in the nerve were removed, yet CL-enhanced regeneration was not impaired. CONCLUSIONS: Macrophages in the sciatic nerve are neither necessary nor sufficient to produce a CL response.
Assuntos
Macrófagos , Regeneração Nervosa , Traumatismos dos Nervos Periféricos , Receptores CCR2 , Degeneração Walleriana , Animais , Macrófagos/metabolismo , Macrófagos/patologia , Camundongos , Regeneração Nervosa/fisiologia , Degeneração Walleriana/patologia , Receptores CCR2/metabolismo , Receptores CCR2/genética , Receptores CCR2/deficiência , Traumatismos dos Nervos Periféricos/patologia , Traumatismos dos Nervos Periféricos/metabolismo , Camundongos Endogâmicos C57BL , Camundongos Knockout , Neuropatia Ciática/patologia , Axônios/patologia , Camundongos Transgênicos , Modelos Animais de Doenças , Nervo Isquiático/lesões , Nervo Isquiático/patologia , Gânglios Espinais/metabolismo , Gânglios Espinais/patologia , Proteínas de Fluorescência Verde/genética , Proteínas de Fluorescência Verde/metabolismoRESUMO
Immune checkpoint inhibitors remained the standard-of-care treatment for advanced non-small cell lung cancer (NSCLC) for the past decade. In unselected patients, anti-PD-(L)1 monotherapy achieved an overall response rate of about 20%. In this analysis, we developed a pharmacokinetic and pharmacodynamic module for our previously calibrated quantitative systems pharmacology model (QSP) to simulate the effectiveness of macrophage-targeted therapies in combination with PD-L1 inhibition in advanced NSCLC. By conducting in silico clinical trials, the model confirmed that anti-CD47 treatment is not an optimal option of second- and later-line treatment for advanced NSCLC resistant to PD-(L)1 blockade. Furthermore, the model predicted that inhibition of macrophage recruitment, such as using CCR2 inhibitors, can potentially improve tumor size reduction when combined with anti-PD-(L)1 therapy, especially in patients who are likely to respond to anti-PD-(L)1 monotherapy and those with a high level of tumor-associated macrophages. Here, we demonstrate the application of the QSP platform on predicting the effectiveness of novel drug combinations involving immune checkpoint inhibitors based on preclinical or early-stage clinical trial data.
Assuntos
Antígeno B7-H1 , Carcinoma Pulmonar de Células não Pequenas , Inibidores de Checkpoint Imunológico , Neoplasias Pulmonares , Humanos , Carcinoma Pulmonar de Células não Pequenas/tratamento farmacológico , Carcinoma Pulmonar de Células não Pequenas/patologia , Carcinoma Pulmonar de Células não Pequenas/imunologia , Neoplasias Pulmonares/tratamento farmacológico , Neoplasias Pulmonares/patologia , Neoplasias Pulmonares/imunologia , Inibidores de Checkpoint Imunológico/farmacologia , Inibidores de Checkpoint Imunológico/uso terapêutico , Inibidores de Checkpoint Imunológico/administração & dosagem , Inibidores de Checkpoint Imunológico/farmacocinética , Antígeno B7-H1/antagonistas & inibidores , Antígeno B7-H1/metabolismo , Protocolos de Quimioterapia Combinada Antineoplásica/farmacologia , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapêutico , Antígeno CD47/antagonistas & inibidores , Antígeno CD47/metabolismo , Macrófagos/metabolismo , Macrófagos/efeitos dos fármacos , Macrófagos/imunologia , Receptores CCR2/antagonistas & inibidores , Receptores CCR2/metabolismo , Farmacologia em Rede/métodos , Simulação por Computador , Modelos Biológicos , Macrófagos Associados a Tumor/efeitos dos fármacos , Macrófagos Associados a Tumor/imunologia , Macrófagos Associados a Tumor/metabolismoRESUMO
Although the pathogenesis of choroidal neovascularization (CNV) is largely unknown in age-related macular degeneration (AMD), inflammasomes may contribute to CNV development and progression. To understand the role NLRP3 inflammasomes in CNV, we used Ccr2RFPCx3cr1GFP dual-reporter mice and immunostaining techniques to confirm localization of NLRP3 inflammasomes in the laser-induced CNV (LCNV) lesions. Confocal microscopy was used to image and quantify LCNV volumes. MCC950 was used as NLRP3 inhibitor. ELISA and quantitative RT-PCR were used to confirm the activation of NLRP3 by monitoring the expression of IL-1ß protein and mRNA in choroidal tissues from LCNV mice. In addition, NLRP3 (-/-) LCNV mice were used to investigate whether NLRP3 inflammasomes contribute to the development of LCNV lesions. We observed that red fluorescent protein (RFP)-positive monocyte-derived macrophages and GFP-positive microglia-derived macrophages, in addition to other cell types, were localized in LCNV lesions at day 7 post-laser injury. In addition, NLRP3 inflammasomes are associated with LCNV lesions. Inhibition of NLRP3 inflammasomes, using MCC950, caused an increased Ccr2RFP-positive macrophages, Cx3cr1GFP-positive microglia, and other cells, resulting in an increase in total lesion size. NLRP3 (-/-) LCNV mice showed significantly increased lesion size compared with age-matched controls. Inhibition of NLRP3 resulted in decreased IL-1ß mRNA and protein expression in the choroidal tissues, suggesting that increased lesion size may not be directly related to IL-1ß.
Assuntos
Neovascularização de Coroide , Indenos , Inflamassomos , Interleucina-1beta , Microglia , Monócitos , Proteína 3 que Contém Domínio de Pirina da Família NLR , Animais , Proteína 3 que Contém Domínio de Pirina da Família NLR/metabolismo , Proteína 3 que Contém Domínio de Pirina da Família NLR/genética , Neovascularização de Coroide/metabolismo , Neovascularização de Coroide/patologia , Camundongos , Inflamassomos/metabolismo , Interleucina-1beta/metabolismo , Microglia/metabolismo , Monócitos/metabolismo , Camundongos Knockout , Sulfonas/farmacologia , Camundongos Endogâmicos C57BL , Furanos/farmacologia , Receptores CCR2/metabolismo , Receptores CCR2/genética , Macrófagos/metabolismo , Macrófagos/imunologia , Sulfonamidas/farmacologia , Compostos Heterocíclicos de 4 ou mais Anéis/farmacologia , Proteínas de Transporte/metabolismo , Proteínas de Transporte/genética , Corioide/metabolismo , Corioide/patologia , Modelos Animais de Doenças , Lasers/efeitos adversos , Degeneração Macular/patologia , Degeneração Macular/metabolismo , Degeneração Macular/genéticaRESUMO
Introduction: Myocardial infarction (MI) is a significant contributor to morbidity and mortality worldwide. Many individuals who survive the acute event continue to experience heart failure (HF), with inflammatory and healing processes post-MI playing a pivotal role. Polymorphonuclear neutrophils (PMN) and monocytes infiltrate the infarcted area, where PMN release high amounts of the heme enzyme myeloperoxidase (MPO). MPO has numerous inflammatory properties and MPO plasma levels are correlated with prognosis and severity of MI. While studies have focused on MPO inhibition and controlling PMN infiltration into the infarcted tissue, less is known on MPO's role in monocyte function. Methods and results: Here, we combined human data with mouse and cell studies to examine the role of MPO on monocyte activation and migration. We revealed a correlation between plasma MPO levels and monocyte activation in a patient study. Using a mouse model of MI, we demonstrated that MPO deficiency led to an increase in splenic monocytes and a decrease in cardiac monocytes compared to wildtype mice (WT). In vitro studies further showed that MPO induces monocyte migration, with upregulation of the chemokine receptor CCR2 and upregulation of inflammatory pathways identified as underlying mechanisms. Conclusion: Taken together, we identify MPO as a pro-inflammatory mediator of splenic monocyte recruitment and activation post-MI and provide mechanistic insight for novel therapeutic strategies after ischemic injury.
Assuntos
Monócitos , Infarto do Miocárdio , Peroxidase , Animais , Infarto do Miocárdio/imunologia , Infarto do Miocárdio/patologia , Infarto do Miocárdio/metabolismo , Peroxidase/metabolismo , Monócitos/imunologia , Monócitos/metabolismo , Humanos , Camundongos , Masculino , Movimento Celular , Modelos Animais de Doenças , Camundongos Endogâmicos C57BL , Feminino , Neutrófilos/imunologia , Neutrófilos/metabolismo , Camundongos Knockout , Receptores CCR2/metabolismo , Pessoa de Meia-IdadeRESUMO
Background: Schistosomiasis is a common cause of pulmonary hypertension (PH) worldwide. Type 2 inflammation contributes to the development of Schistosoma-induced PH. Specifically, interstitial macrophages (IMs) derived from monocytes play a pivotal role by producing thrombospondin-1 (TSP-1), which in turn activates TGF-ß, thereby driving the pathology of PH. Resident and recruited IM subpopulations have recently been identified. We hypothesized that in Schistosoma-PH, one IM subpopulation expresses monocyte recruitment factors, whereas recruited monocytes become a separate IM subpopulation that expresses TSP-1. Methods: Mice were intraperitoneally sensitized and then intravenously challenged with S. mansoni eggs. Flow cytometry on lungs and blood was performed on wildtype and reporter mice to identify IM subpopulations and protein expression. Single-cell RNA sequencing (scRNAseq) was performed on flow-sorted IMs from unexposed and at day 1, 3 and 7 following Schistosoma exposure to complement flow cytometry based IM characterization and identify gene expression. Results: Flow cytometry and scRNAseq both identified 3 IM subpopulations, characterized by CCR2, MHCII, and FOLR2 expression. Following Schistosoma exposure, the CCR2+ IM subpopulation expanded, suggestive of circulating monocyte recruitment. Schistosoma exposure caused increased monocyte-recruitment ligand CCL2 expression in the resident FOLR2+ IM subpopulation. In contrast, the vascular pathology-driving protein TSP-1 was greatest in the CCR2+ IM subpopulation. Conclusion: Schistosoma-induced PH involves crosstalk between IM subpopulations, with increased expression of monocyte recruitment ligands by resident FOLR2+ IMs, and the recruitment of CCR2+ IMs which express TSP-1 that activates TGF-ß and causes PH.
Assuntos
Hipertensão Pulmonar , Macrófagos , Animais , Hipertensão Pulmonar/etiologia , Hipertensão Pulmonar/parasitologia , Hipertensão Pulmonar/imunologia , Hipertensão Pulmonar/patologia , Camundongos , Macrófagos/imunologia , Macrófagos/parasitologia , Fenótipo , Schistosoma mansoni/imunologia , Camundongos Endogâmicos C57BL , Esquistossomose/imunologia , Esquistossomose/complicações , Esquistossomose/parasitologia , Modelos Animais de Doenças , Esquistossomose mansoni/imunologia , Esquistossomose mansoni/parasitologia , Esquistossomose mansoni/complicações , Esquistossomose mansoni/patologia , Trombospondina 1/genética , Trombospondina 1/metabolismo , Monócitos/imunologia , Receptores CCR2/genética , Receptores CCR2/metabolismo , Feminino , Schistosoma/imunologia , Schistosoma/fisiologia , Pulmão/imunologia , Pulmão/parasitologia , Pulmão/patologiaRESUMO
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.