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Regulatory T cells (Tregs) are key immune regulators that have shown promise in enhancing cardiac repair post-MI, although the mechanisms remain elusive. Here, we show that rapidly increasing Treg number in the circulation post-MI via systemic administration of exogenous Tregs improves cardiac function in male mice, by limiting cardiomyocyte death and reducing fibrosis. Mechanistically, exogenous Tregs quickly home to the infarcted heart and adopt an injury-specific transcriptome that mediates repair by modulating monocytes/macrophages. Specially, Tregs lead to a reduction in pro-inflammatory Ly6CHi CCR2+ monocytes/macrophages accompanied by a rapid shift of macrophages towards a pro-repair phenotype. Additionally, exogenous Treg-derived factors, including nidogen-1 and IL-10, along with a decrease in cardiac CD8+ T cell number, mediate the reduction of the pro-inflammatory monocyte/macrophage subset in the heart. Supporting the pivotal role of IL-10, exogenous Tregs knocked out for IL-10 lose their pro-repair capabilities. Together, this study highlights the beneficial use of a Treg-based therapeutic approach for cardiac repair with important mechanistic insights that could facilitate the development of novel immunotherapies for MI.
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Interleucina-10 , Macrófagos , Ratones Endogámicos C57BL , Infarto del Miocardio , Linfocitos T Reguladores , Animales , Infarto del Miocardio/inmunología , Infarto del Miocardio/genética , Infarto del Miocardio/patología , Linfocitos T Reguladores/inmunología , Macrófagos/inmunología , Macrófagos/metabolismo , Masculino , Ratones , Interleucina-10/metabolismo , Interleucina-10/genética , Fenotipo , Miocardio/patología , Miocardio/inmunología , Miocardio/metabolismo , Monocitos/inmunología , Monocitos/metabolismo , Miocitos Cardíacos/metabolismo , Miocitos Cardíacos/inmunología , Fibrosis , Linfocitos T CD8-positivos/inmunología , Modelos Animales de Enfermedad , Ratones NoqueadosRESUMEN
Although V(D)J recombination and immunoglobulin (Ig) production are traditionally recognised to occur only in B lymphocytes and plasma cells, the expression of Igs in non-lymphoid cells, which we call non B cell-derived Igs (non B Igs), has been documented by growing studies. It has been demonstrated that non B-Igs can be widely expressed in most cell types, including, but not limited to, epithelial cells, cardiomyocytes, hematopoietic stem/progenitor cells, myeloid cells, and cells from immune-privileged sites, such as neurons and spermatogenic cells. In particular, malignant tumour cells express high level of IgG. Moreover, different from B-Igs that mainly localised on the B cell membrane and in the serum and perform immune defence function mainly, non B-Igs have been found to distribute more widely and play critical roles in immune defence, maintaining cell proliferation and survival, and promoting progression. The findings of non B-Igs may provide a wealthier breakthrough point for more therapeutic strategies for a wide range of immune-related diseases.
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Inmunoglobulinas , Humanos , Animales , Inmunoglobulinas/genética , Inmunoglobulinas/metabolismo , Inmunoglobulinas/inmunología , Células Madre Hematopoyéticas/metabolismo , Células Madre Hematopoyéticas/inmunología , Células Madre Hematopoyéticas/citología , Linfocitos B/inmunología , Linfocitos B/metabolismo , Células Epiteliales/metabolismo , Células Epiteliales/inmunología , Miocitos Cardíacos/metabolismo , Miocitos Cardíacos/inmunología , Células Mieloides/inmunología , Células Mieloides/metabolismoRESUMEN
Introduction: Schistosomiasis (SM) is a parasitic disease caused by Schistosoma mansoni. SM causes chronic inflammation induced by parasitic eggs, with collagen/fibrosis deposition in the granuloma process in the liver, spleen, central nervous system, kidneys, and lungs. Pulmonary arterial hypertension (PAH) is a clinical manifestation characterized by high pressure in the pulmonary circulation and right ventricular overload. This study investigated the production of functional autoantibodies (fAABs) against the second loop of the G-protein-coupled receptor (GPCR) in the presence of hepatic and PAH forms of human SM. Methods: Uninfected and infected individuals presenting acute and chronic manifestations (e.g., hepatointestinal, hepato-splenic without PAH, and hepato-splenic with PAH) of SM were clinically evaluated and their blood was collected to identify fAABs/GPCRs capable of recognizing endothelin 1, angiotensin II, and a-1 adrenergic receptor. Human serum was analyzed in rat cardiomyocytes cultured in the presence of the receptor antagonists urapidil, losartan, and BQ123. Results: The fAABs/GPCRs from chronic hepatic and PAH SM individuals, but not from acute SM individuals, recognized the three receptors. In the presence of the antagonists, there was a reduction in beating rate changes in cultured cardiomyocytes. In addition, binding sites on the extracellular domain functionality of fAABs were identified, and IgG1 and/or IgG3 antibodies were found to be related to fAABs. Conclusion: Our data suggest that fAABs against GPCR play an essential role in vascular activity in chronic SM (hepatic and PAH) and might be involved in the development of hypertensive forms of SM.
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Autoanticuerpos , Receptores Acoplados a Proteínas G , Autoanticuerpos/inmunología , Autoanticuerpos/sangre , Humanos , Animales , Receptores Acoplados a Proteínas G/inmunología , Receptores Acoplados a Proteínas G/metabolismo , Ratas , Masculino , Femenino , Adulto , Hipertensión Pulmonar/inmunología , Hipertensión Pulmonar/etiología , Persona de Mediana Edad , Miocitos Cardíacos/inmunología , Miocitos Cardíacos/metabolismo , Miocitos Cardíacos/parasitología , Esquistosomiasis mansoni/inmunología , Schistosoma mansoni/inmunología , Esquistosomiasis/inmunologíaRESUMEN
Heart transplantation is associated with major hurdles, including the limited number of available organs for transplantation, the risk of rejection due to genetic discrepancies, and the burden of immunosuppression. In this study, we demonstrated the feasibility of permanent genetic engineering of the heart during ex vivo perfusion. Lentiviral vectors encoding for short hairpin RNAs targeting beta2-microglobulin (shß2m) and class II transactivator (shCIITA) were delivered to the graft during two hours of normothermic EVHP. Highly efficient genetic engineering was indicated by stable reporter gene expression in endothelial cells and cardiomyocytes. Remarkably, swine leucocyte antigen (SLA) class I and SLA class II expression levels were decreased by 66% and 76%, respectively, in the vascular endothelium. Evaluation of lactate, troponin T, and LDH levels in the perfusate and histological analysis showed no additional cell injury or tissue damage caused by lentiviral vectors. Moreover, cytokine secretion profiles (IL-6, IL-8, and TNF-α) of non-transduced and lentiviral vector-transduced hearts were comparable. This study demonstrated the ex vivo generation of genetically engineered hearts without compromising tissue integrity. Downregulation of SLA expression may contribute to reduce the immunogenicity of the heart and support graft survival after allogeneic or xenogeneic transplantation.
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Vectores Genéticos , Trasplante de Corazón , Antígenos de Histocompatibilidad Clase I , Lentivirus , Animales , Lentivirus/genética , Trasplante de Corazón/métodos , Vectores Genéticos/genética , Porcinos , Antígenos de Histocompatibilidad Clase I/genética , Antígenos de Histocompatibilidad Clase I/inmunología , Antígenos de Histocompatibilidad Clase I/metabolismo , Perfusión/métodos , Antígenos de Histocompatibilidad Clase II/genética , Antígenos de Histocompatibilidad Clase II/metabolismo , Antígenos de Histocompatibilidad Clase II/inmunología , Microglobulina beta-2/genética , Citocinas/metabolismo , Ingeniería Genética , Miocitos Cardíacos/metabolismo , Miocitos Cardíacos/inmunología , Humanos , ARN Interferente Pequeño/genética , Supervivencia de Injerto/inmunología , Supervivencia de Injerto/genética , Células Endoteliales/metabolismo , Células Endoteliales/inmunología , Proteínas Nucleares , TransactivadoresRESUMEN
Immunometabolism is an emerging field at the intersection of immunology and metabolism. Immune cell activation plays a critical role in the pathogenesis of cardiovascular diseases and is integral for regeneration during cardiac injury. We currently possess a limited understanding of the processes governing metabolic interactions between immune cells and cardiomyocytes. The impact of this intercellular crosstalk can manifest as alterations to the steady state flux of metabolites and impact cardiac contractile function. Although much of our knowledge is derived from acute inflammatory response, recent work emphasizes heterogeneity and flexibility in metabolism between cardiomyocytes and immune cells during pathological states, including ischemic, cardiometabolic, and cancer-associated disease. Metabolic adaptation is crucial because it influences immune cell activation, cytokine release, and potential therapeutic vulnerabilities. This review describes current concepts about immunometabolic regulation in the heart, focusing on intercellular crosstalk and intrinsic factors driving cellular regulation. We discuss experimental approaches to measure the cardio-immunologic crosstalk, which are necessary to uncover unknown mechanisms underlying the immune and cardiac interface. Deeper insight into these axes holds promise for therapeutic strategies that optimize cardioimmunology crosstalk for cardiac health.
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Miocitos Cardíacos , Humanos , Animales , Miocitos Cardíacos/metabolismo , Miocitos Cardíacos/inmunología , Metabolismo Energético , Cardiomiopatías/metabolismo , Cardiomiopatías/inmunología , Miocardio/metabolismo , Miocardio/inmunología , Miocardio/patologíaRESUMEN
Myocarditis is clinically characterized by chest pain, arrhythmias, and heart failure, and treatment is often supportive. Mutations in DSP, a gene encoding the desmosomal protein desmoplakin, have been increasingly implicated in myocarditis. To model DSP-associated myocarditis and assess the role of innate immunity, we generated engineered heart tissues (EHTs) using human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs) from patients with heterozygous DSP truncating variants (DSPtvs) and a gene-edited homozygous deletion cell line (DSP-/-). At baseline, DSP-/- EHTs displayed a transcriptomic signature of innate immune activation, which was mirrored by cytokine release. Importantly, DSP-/- EHTs were hypersensitive to Toll-like receptor (TLR) stimulation, demonstrating more contractile dysfunction compared with isogenic controls. Relative to DSP-/- EHTs, heterozygous DSPtv EHTs had less functional impairment. DSPtv EHTs displayed heightened sensitivity to TLR stimulation, and when subjected to strain, DSPtv EHTs developed functional deficits, indicating reduced contractile reserve compared with healthy controls. Colchicine or NF-κB inhibitors improved strain-induced force deficits in DSPtv EHTs. Genomic correction of DSP p.R1951X using adenine base editing reduced inflammatory biomarker release from EHTs. Thus, EHTs replicate electrical and contractile phenotypes seen in human myocarditis, implicating cytokine release as a key part of the myogenic susceptibility to inflammation. The heightened innate immune activation and sensitivity are targets for clinical intervention.
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Inmunidad Innata , Células Madre Pluripotentes Inducidas , Miocarditis , Miocitos Cardíacos , Humanos , Miocarditis/genética , Miocarditis/inmunología , Miocarditis/patología , Inmunidad Innata/genética , Células Madre Pluripotentes Inducidas/metabolismo , Células Madre Pluripotentes Inducidas/inmunología , Miocitos Cardíacos/metabolismo , Miocitos Cardíacos/inmunología , Miocitos Cardíacos/patología , Masculino , Predisposición Genética a la Enfermedad , FemeninoRESUMEN
Nuclear factor κ-B (NFκB) is activated in iPSC-cardiac myocytes from patients with arrhythmogenic cardiomyopathy (ACM) under basal conditions, and inhibition of NFκB signaling prevents disease in Dsg2mut/mut mice, a robust mouse model of ACM. Here, we used genetic approaches and single-cell RNA-Seq to define the contributions of immune signaling in cardiac myocytes and macrophages in the natural progression of ACM using Dsg2mut/mut mice. We found that NFκB signaling in cardiac myocytes drives myocardial injury, contractile dysfunction, and arrhythmias in Dsg2mut/mut mice. NFκB signaling in cardiac myocytes mobilizes macrophages expressing C-C motif chemokine receptor-2 (CCR2+ cells) to affected areas within the heart, where they mediate myocardial injury and arrhythmias. Contractile dysfunction in Dsg2mut/mut mice is caused both by loss of heart muscle and negative inotropic effects of inflammation in viable muscle. Single nucleus RNA-Seq and cellular indexing of transcriptomes and epitomes (CITE-Seq) studies revealed marked proinflammatory changes in gene expression and the cellular landscape in hearts of Dsg2mut/mut mice involving cardiac myocytes, fibroblasts, and CCR2+ macrophages. Changes in gene expression in cardiac myocytes and fibroblasts in Dsg2mut/mut mice were dependent on CCR2+ macrophage recruitment to the heart. These results highlight complex mechanisms of immune injury and regulatory crosstalk between cardiac myocytes, inflammatory cells, and fibroblasts in the pathogenesis of ACM.
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Desmogleína 2 , Modelos Animales de Enfermedad , Macrófagos , FN-kappa B , Receptores CCR2 , Transducción de Señal , Animales , Ratones , Macrófagos/metabolismo , Macrófagos/patología , Macrófagos/inmunología , Receptores CCR2/genética , Receptores CCR2/metabolismo , Desmogleína 2/genética , Desmogleína 2/metabolismo , FN-kappa B/metabolismo , FN-kappa B/genética , Miocitos Cardíacos/metabolismo , Miocitos Cardíacos/patología , Miocitos Cardíacos/inmunología , Humanos , Displasia Ventricular Derecha Arritmogénica/genética , Displasia Ventricular Derecha Arritmogénica/metabolismo , Displasia Ventricular Derecha Arritmogénica/patología , Miocardio/patología , Miocardio/metabolismo , Miocardio/inmunologíaRESUMEN
In contrast to adult mammals, adult zebrafish can fully regenerate injured cardiac tissue, and this regeneration process requires an adequate and tightly controlled immune response. However, which components of the immune response are required during regeneration is unclear. Here, we report positive roles for the antigen presentation-adaptive immunity axis during zebrafish cardiac regeneration. We find that following the initial innate immune response, activated endocardial cells (EdCs), as well as immune cells, start expressing antigen presentation genes. We also observe that T helper cells, a.k.a. Cd4+ T cells, lie in close physical proximity to these antigen-presenting EdCs. We targeted Major Histocompatibility Complex (MHC) class II antigen presentation by generating cd74a; cd74b mutants, which display a defective immune response. In these mutants, Cd4+ T cells and activated EdCs fail to efficiently populate the injured tissue and EdC proliferation is significantly decreased. cd74a; cd74b mutants exhibit additional defects in cardiac regeneration including reduced cardiomyocyte dedifferentiation and proliferation. Notably, Cd74 also becomes activated in neonatal mouse EdCs following cardiac injury. Altogether, these findings point to positive roles for antigen presentation during cardiac regeneration, potentially involving interactions between activated EdCs, classical antigen-presenting cells, and Cd4+ T cells.
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Presentación de Antígeno , Lesiones Cardíacas , Antígenos de Histocompatibilidad Clase II , Regeneración , Pez Cebra , Animales , Regeneración/inmunología , Presentación de Antígeno/inmunología , Lesiones Cardíacas/inmunología , Antígenos de Histocompatibilidad Clase II/metabolismo , Antígenos de Histocompatibilidad Clase II/inmunología , Antígenos de Histocompatibilidad Clase II/genética , Ratones , Linfocitos T CD4-Positivos/inmunología , Miocitos Cardíacos/inmunología , Miocitos Cardíacos/metabolismo , Proteínas de Pez Cebra/genética , Proteínas de Pez Cebra/metabolismo , Antígenos de Diferenciación de Linfocitos B/metabolismo , Antígenos de Diferenciación de Linfocitos B/genética , Proliferación Celular , Inmunidad Innata , Corazón/fisiopatología , Corazón/fisiología , Mutación , Inmunidad Adaptativa , Animales Modificados GenéticamenteRESUMEN
Sepsis often causes cell death via pyroptosis and hence results in septic cardiomyopathy. Triggering receptors expressed in myeloid cells-1 (TREM-1) may initiate cellular cascade pathways and, in turn, induce cell death and vital organ dysfunction in sepsis, but the evidence is limited. We set to investigate the role of TREM-1 on nucleotide-binding oligomerization domain-like receptors with pyrin domain-3 (NLRP3) inflammasome activation and cardiomyocyte pyroptosis in sepsis models using cardiac cell line (HL-1) and mice. In this study, TREM-1 was found to be significantly increased in HL-1 cells challenged with lipopolysaccharide (LPS). Pyroptosis was also significantly increased in the HL-1 cells challenged with lipopolysaccharide and an NLRP3 inflammasome activator, nigericin. The close interaction between TREM-1 and structural maintenance of chromosome 4 (SMC4) was also identified. Furthermore, inhibition of TREM-1 or SMC4 prevented the upregulation of NLRP3 and decreased Gasdermin-D, IL-1ß and caspase-1 cleavage. In mice subjected to caecal ligation and puncture, the TREM-1 inhibitor LR12 decreased the expression of NLRP3 and attenuated cardiomyocyte pyroptosis, leading to improved cardiac function and prolonged survival of septic mice. Our work demonstrates that, under septic conditions, TREM-1 plays a critical role in cardiomyocyte pyroptosis. Targeting TREM-1 and its associated molecules may therefore lead to novel therapeutic treatments for septic cardiomyopathy.
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Inflamasomas , Miocitos Cardíacos , Proteína con Dominio Pirina 3 de la Familia NLR , Piroptosis , Sepsis , Receptor Activador Expresado en Células Mieloides 1 , Animales , Humanos , Ratones , Adenosina Trifosfatasas/inmunología , Cardiomiopatías/etiología , Cardiomiopatías/genética , Cardiomiopatías/inmunología , Caspasa 1/genética , Proteínas Cromosómicas no Histona/genética , Proteínas Cromosómicas no Histona/inmunología , Cromosomas Humanos Par 4/inmunología , Inflamasomas/agonistas , Inflamasomas/genética , Inflamasomas/inmunología , Lipopolisacáridos/efectos adversos , Lipopolisacáridos/farmacología , Células Mieloides/inmunología , Miocitos Cardíacos/inmunología , Proteína con Dominio Pirina 3 de la Familia NLR/agonistas , Proteína con Dominio Pirina 3 de la Familia NLR/genética , Proteína con Dominio Pirina 3 de la Familia NLR/inmunología , Piroptosis/genética , Piroptosis/inmunología , Sepsis/complicaciones , Sepsis/genética , Sepsis/inmunología , Receptor Activador Expresado en Células Mieloides 1/antagonistas & inhibidores , Receptor Activador Expresado en Células Mieloides 1/genética , Receptor Activador Expresado en Células Mieloides 1/inmunologíaRESUMEN
BACKGROUND: CC chemokine receptor 5 (CCR5) has been demonstrated to be correlated to activation of pro-inflammatory immune cells and tissue injury. This study focused on the role of CCR5 in myocardial injury in rats with diabetic cardiomyopathy (DCM) and the mechanism of action. METHODS: A rat model of DCM was induced by streptozotocin (STZ). CCR5 was knocked down in rats to determine its role in myocardial injury and immune cell infiltration. The upstream regulators of CCR5 were bioinformatically predicted and the binding between nuclear receptor subfamily 4 group A member 2 (NR4A2) and CCR5 was validated. The portion of M1 and M2 macrophages in tissues was determined by flow cytometry or double-labeling immunofluorescence. Rat bone marrow mononuclear cells (BMMCs) were treated with granulocyte/macrophage colony stimulating factor (GM-CSF/M-CSF) and co-cultured with H9C2 cells for in vitro experiments. RESULTS: STZ-treated rats had impaired cardiac function and increased levels of creatine kinase-MB, cardiac troponin I and lactate dehydrogenase. CCR5 inhibition significantly alleviated myocardial injury in rats and reduced the portion of M1 macrophages in rat cardiac tissues. NR4A2, which could suppress CCR5 transcription, was poorly expressed in rats with DCM. NR4A2 overexpression played a similar myocardium-protective role in rats. In vitro, overexpression of NR4A2 induced M2 polarization of macrophages, which protected the co-cultured H9C2 cells from high glucose-induced damage, but the protective role was blocked after CCR5 overexpression. CONCLUSION: This study demonstrated that NR4A2 suppresses CCR5 expression and promotes M2 polarization of macrophages to alleviate cardiomyocyte loss and myocardial injury.
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Cardiomiopatías Diabéticas , Macrófagos , Miocitos Cardíacos , Miembro 2 del Grupo A de la Subfamilia 4 de Receptores Nucleares , Receptores CCR5 , Transcripción Genética , Animales , Masculino , Línea Celular , Técnicas de Cocultivo , Cardiomiopatías Diabéticas/genética , Cardiomiopatías Diabéticas/inmunología , Cardiomiopatías Diabéticas/metabolismo , Cardiomiopatías Diabéticas/patología , Modelos Animales de Enfermedad , Regulación hacia Abajo , Macrófagos/inmunología , Macrófagos/metabolismo , Macrófagos/patología , Miocitos Cardíacos/inmunología , Miocitos Cardíacos/metabolismo , Miocitos Cardíacos/patología , Miembro 2 del Grupo A de la Subfamilia 4 de Receptores Nucleares/genética , Miembro 2 del Grupo A de la Subfamilia 4 de Receptores Nucleares/metabolismo , Fenotipo , Ratas Sprague-Dawley , Receptores CCR5/genética , Receptores CCR5/metabolismo , Transducción de SeñalRESUMEN
COVID-19 affects multiple organs. Clinical data from the Mount Sinai Health System show that substantial numbers of COVID-19 patients without prior heart disease develop cardiac dysfunction. How COVID-19 patients develop cardiac disease is not known. We integrated cell biological and physiological analyses of human cardiomyocytes differentiated from human induced pluripotent stem cells (hiPSCs) infected with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) in the presence of interleukins (ILs) with clinical findings related to laboratory values in COVID-19 patients to identify plausible mechanisms of cardiac disease in COVID-19 patients. We infected hiPSC-derived cardiomyocytes from healthy human subjects with SARS-CoV-2 in the absence and presence of IL-6 and IL-1ß. Infection resulted in increased numbers of multinucleated cells. Interleukin treatment and infection resulted in disorganization of myofibrils, extracellular release of troponin I, and reduced and erratic beating. Infection resulted in decreased expression of mRNA encoding key proteins of the cardiomyocyte contractile apparatus. Although interleukins did not increase the extent of infection, they increased the contractile dysfunction associated with viral infection of cardiomyocytes, resulting in cessation of beating. Clinical data from hospitalized patients from the Mount Sinai Health System show that a significant portion of COVID-19 patients without history of heart disease have elevated troponin and interleukin levels. A substantial subset of these patients showed reduced left ventricular function by echocardiography. Our laboratory observations, combined with the clinical data, indicate that direct effects on cardiomyocytes by interleukins and SARS-CoV-2 infection might underlie heart disease in COVID-19 patients. IMPORTANCE SARS-CoV-2 infects multiple organs, including the heart. Analyses of hospitalized patients show that a substantial number without prior indication of heart disease or comorbidities show significant injury to heart tissue, assessed by increased levels of troponin in blood. We studied the cell biological and physiological effects of virus infection of healthy human iPSC-derived cardiomyocytes in culture. Virus infection with interleukins disorganizes myofibrils, increases cell size and the numbers of multinucleated cells, and suppresses the expression of proteins of the contractile apparatus. Viral infection of cardiomyocytes in culture triggers release of troponin similar to elevation in levels of COVID-19 patients with heart disease. Viral infection in the presence of interleukins slows down and desynchronizes the beating of cardiomyocytes in culture. The cell-level physiological changes are similar to decreases in left ventricular ejection seen in imaging of patients' hearts. These observations suggest that direct injury to heart tissue by virus can be one underlying cause of heart disease in COVID-19.
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COVID-19/inmunología , Células Madre Pluripotentes Inducidas , Interleucina-10/inmunología , Interleucina-1beta/inmunología , Interleucina-6/inmunología , Miocitos Cardíacos , Células Cultivadas , Humanos , Células Madre Pluripotentes Inducidas/inmunología , Células Madre Pluripotentes Inducidas/patología , Células Madre Pluripotentes Inducidas/virología , Miocitos Cardíacos/inmunología , Miocitos Cardíacos/patología , Miocitos Cardíacos/virologíaRESUMEN
AIMS: Myocardial infarction (MI) is the most common cause of heart failure (HF) worldwide. G protein-coupled receptor kinase 5 (GRK5) is upregulated in failing human myocardium and promotes maladaptive cardiac hypertrophy in animal models. However, the role of GRK5 in ischemic heart disease is still unknown. In this study, we evaluated whether myocardial GRK5 plays a critical role post-MI in mice and included the examination of specific cardiac immune and inflammatory responses. METHODS AND RESULTS: Cardiomyocyte-specific GRK5 overexpressing transgenic mice (TgGRK5) and non-transgenic littermate control (NLC) mice as well as cardiomyocyte-specific GRK5 knockout mice (GRK5cKO) and wild type (WT) were subjected to MI and, functional as well as structural changes together with outcomes were studied. TgGRK5 post-MI mice showed decreased cardiac function, augmented left ventricular dimension and decreased survival rate compared to NLC post-MI mice. Cardiac hypertrophy and fibrosis as well as fetal gene expression were increased post-MI in TgGRK5 compared to NLC mice. In TgGRK5 mice, GRK5 elevation produced immuno-regulators that contributed to the elevated and long-lasting leukocyte recruitment into the injured heart and ultimately to chronic cardiac inflammation. We found an increased presence of pro-inflammatory neutrophils and macrophages as well as neutrophils, macrophages and T-lymphocytes at 4-days and 8-weeks respectively post-MI in TgGRK5 hearts. Conversely, GRK5cKO mice were protected from ischemic injury and showed reduced early immune cell recruitment (predominantly monocytes) to the heart, improved contractility and reduced mortality compared to WT post-MI mice. Interestingly, cardiomyocyte-specific GRK2 transgenic mice did not share the same phenotype of TgGRK5 mice and did not have increased cardiac leukocyte migration and cytokine or chemokine production post-MI. CONCLUSIONS: Our study shows that myocyte GRK5 has a crucial and GRK-selective role on the regulation of leucocyte infiltration into the heart, cardiac function and survival in a murine model of post-ischemic HF, supporting GRK5 inhibition as a therapeutic target for HF.
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Quimiotaxis de Leucocito , Quinasa 5 del Receptor Acoplado a Proteína-G/metabolismo , Insuficiencia Cardíaca/enzimología , Leucocitos/metabolismo , Infarto del Miocardio/enzimología , Miocitos Cardíacos/enzimología , Función Ventricular Izquierda , Animales , Citocinas/genética , Citocinas/metabolismo , Modelos Animales de Enfermedad , Quinasa 5 del Receptor Acoplado a Proteína-G/genética , Insuficiencia Cardíaca/inmunología , Insuficiencia Cardíaca/patología , Insuficiencia Cardíaca/fisiopatología , Mediadores de Inflamación/metabolismo , Leucocitos/inmunología , Ratones Noqueados , Contracción Miocárdica , Infarto del Miocardio/inmunología , Infarto del Miocardio/patología , Infarto del Miocardio/fisiopatología , Miocitos Cardíacos/inmunología , Miocitos Cardíacos/patología , Transducción de Señal , Volumen Sistólico , Transcriptoma , Presión VentricularRESUMEN
AIMS: Angiotensin (Ang) II signalling has been suggested to promote cardiac fibrosis in inflammatory heart diseases; however, the underlying mechanisms remain obscure. Using Agtr1a-/- mice with genetic deletion of angiotensin receptor type 1 (ATR1) and the experimental autoimmune myocarditis (EAM) model, we aimed to elucidate the role of Ang II-ATR1 pathway in development of heart-specific autoimmunity and post-inflammatory fibrosis. METHODS AND RESULTS: EAM was induced in wild-type (WT) and Agtr1a-/- mice by subcutaneous injections with alpha myosin heavy chain peptide emulsified in complete Freund's adjuvant. Agtr1a-/- mice developed myocarditis to a similar extent as WT controls at day 21 but showed reduced fibrosis and better systolic function at day 40. Crisscross bone marrow chimaera experiments proved that ATR1 signalling in the bone marrow compartment was critical for cardiac fibrosis. Heart infiltrating, bone-marrow-derived cells produced Ang II, but lack of ATR1 in these cells reduced transforming growth factor beta (TGF-ß)-mediated fibrotic responses. At the molecular level, Agtr1a-/- heart-inflammatory cells showed impaired TGF-ß-mediated phosphorylation of Smad2 and TAK1. In WT cells, TGF-ß induced formation of RhoA-GTP and RhoA-A-kinase anchoring protein-Lbc (AKAP-Lbc) complex. In Agtr1a-/- cells, stabilization of RhoA-GTP and interaction of RhoA with AKAP-Lbc were largely impaired. Furthermore, in contrast to WT cells, Agtr1a-/- cells stimulated with TGF-ß failed to activate canonical Wnt pathway indicated by suppressed activity of glycogen synthase kinase-3 (GSK-3)ß and nuclear ß-catenin translocation and showed reduced expression of Wnts. In line with these in vitro findings, ß-catenin was detected in inflammatory regions of hearts of WT, but not Agtr1a-/- mice and expression of canonical Wnt1 and Wnt10b were lower in Agtr1a-/- hearts. CONCLUSION: Ang II-ATR1 signalling is critical for development of post-inflammatory fibrotic remodelling and dilated cardiomyopathy. Our data underpin the importance of Ang II-ATR1 in effective TGF-ß downstream signalling response including activation of profibrotic Wnt/ß-catenin pathway.
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Angiotensina II/metabolismo , Enfermedades Autoinmunes/metabolismo , Autoinmunidad , Linfocitos T CD4-Positivos/metabolismo , Miocarditis/metabolismo , Miocitos Cardíacos/metabolismo , Receptor de Angiotensina Tipo 1/metabolismo , Vía de Señalización Wnt , Animales , Enfermedades Autoinmunes/genética , Enfermedades Autoinmunes/inmunología , Enfermedades Autoinmunes/patología , Linfocitos T CD4-Positivos/inmunología , Proliferación Celular , Células Cultivadas , Modelos Animales de Enfermedad , Fibrosis , Mediadores de Inflamación/metabolismo , Activación de Linfocitos , Ratones Endogámicos BALB C , Ratones Noqueados , Miocarditis/genética , Miocarditis/inmunología , Miocarditis/patología , Miocitos Cardíacos/inmunología , Miocitos Cardíacos/patología , Receptor de Angiotensina Tipo 1/genética , Proteínas Wnt/genética , Proteínas Wnt/metabolismo , Proteína Wnt1/genética , Proteína Wnt1/metabolismo , beta Catenina/genética , beta Catenina/metabolismoRESUMEN
Benznidazole (Bzl), the drug of choice in many countries for the treatment of Chagas disease, leads to parasite clearance in the early stages of infection and contributes to immunomodulation. In addition to its parasiticidal effect, Bzl inhibits the NF-κB pathway. In this regard, we have previously described that this occurs through IL-10/STAT3/SOCS3 pathway. PI3K pathway is involved in the regulation of the immune system by inhibiting NF-κB pathway through STAT3. In this work, the participation of PI3K in the immunomodulatory effects of Bzl in cardiac and immune cells, the main targets of Chagas disease, was further studied. For that, we use a murine primary cardiomyocyte culture and a monocyte/macrophage cell line (RAW 264.7), stimulated with LPS in presence of LY294002, an inhibitor of PI3K. Under these conditions, Bzl could neither increase SOCS3 expression nor inhibit the NOS2 mRNA expression and the release of NOx, both in cardiomyocytes and macrophages. Macrophages are crucial in the development of Chronic Chagas Cardiomyopathy. Thus, to deepen our understanding of how Bzl acts, the expression profile of M1-M2 macrophage markers was evaluated. Bzl inhibited the release of NOx (M1 marker) and increased the expression of Arginase I (M2 marker) and a negative correlation was found between them. Besides, LPS increased the expression of pro-inflammatory cytokines. Bzl treatment not only inhibited this effect but also increased the expression of typical M2-macrophage markers like Mannose Receptor, TGF-ß, and VEGF-A. Moreover, Bzl increased the expression of PPAR-γ and PPAR-α, known as key regulators of macrophage polarization. PI3K directly regulates M1-to-M2 macrophage polarization. Since p110δ, catalytic subunit of PI3Kδ, is highly expressed in immune cells, experiments were carried out in presence of CAL-101, a specific inhibitor of this subunit. Under this condition, Bzl could neither increase SOCS3 expression nor inhibit NF-κB pathway. Moreover, Bzl not only failed to inhibit the expression of pro-inflammatory cytokines (M1 markers) but also could not increase M2 markers. Taken together these results demonstrate, for the first time, that the anti-inflammatory effect of Bzl depends on PI3K activity in a cell line of murine macrophages and in primary culture of neonatal cardiomyocytes. Furthermore, Bzl-mediated increase expression of M2-macrophage markers involves the participation of the p110δ catalytic subunit of PI3Kδ.
Asunto(s)
Antiinflamatorios/farmacología , Cardiomiopatía Chagásica/tratamiento farmacológico , Fosfatidilinositol 3-Quinasa Clase I/metabolismo , Nitroimidazoles/farmacología , Animales , Animales Recién Nacidos , Antiinflamatorios/uso terapéutico , Cardiomiopatía Chagásica/inmunología , Cromonas/farmacología , Fosfatidilinositol 3-Quinasa Clase I/antagonistas & inhibidores , Modelos Animales de Enfermedad , Femenino , Humanos , Lipopolisacáridos/inmunología , Macrófagos/efectos de los fármacos , Macrófagos/inmunología , Macrófagos/metabolismo , Masculino , Ratones , Morfolinas/farmacología , Miocitos Cardíacos/efectos de los fármacos , Miocitos Cardíacos/inmunología , Miocitos Cardíacos/metabolismo , Nitroimidazoles/uso terapéutico , Cultivo Primario de Células , Células RAW 264.7RESUMEN
Viral diseases are a major threat to modern society and the global health system. It is therefore of utter relevance to understand the way viruses affect the host as a basis to find new treatment solutions. The understanding of viral myocarditis (VMC) is incomplete and effective treatment options are lacking. This review will discuss the mechanism, effects, and treatment options of the most frequent myocarditis-causing viruses namely enteroviruses such as Coxsackievirus B3 (CVB3) and Parvovirus B19 (PVB19) on the human heart. Thereby, we focus on: 1. Viral entry: CVB3 use Coxsackievirus-Adenovirus-Receptor (CAR) and Decay Accelerating Factor (DAF) to enter cardiac myocytes while PVB19 use the receptor globoside (Gb4) to enter cardiac endothelial cells. 2. Immune system responses: The innate immune system mediated by activated cardiac toll-like receptors (TLRs) worsen inflammation in CVB3-infected mouse hearts. Different types of cells of the adaptive immune system are recruited to the site of inflammation that have either protective or adverse effects during VMC. 3. Autophagy: CVB3 evades autophagosomal degradation and misuses the autophasomal pathway for viral replication and release. 4. Viral replication sites: CVB3 promotes the formation of double membrane vesicles (DMVs), which it uses as replication sites. PVB19 uses the host cell nucleus as the replication site and uses the host cell DNA replication system. 5. Cell cycle manipulation: CVB3 attenuates the cell cycle at the G1/S phase, which promotes viral transcription and replication. PVB19 exerts cell cycle arrest in the S phase using its viral endonuclease activity. 6. Regulation of apoptosis: Enteroviruses prevent apoptosis during early stages of infection and promote cell death during later stages by using the viral proteases 2A and 3C, and viroporin 2B. PVB19 promotes apoptosis using the non-structural proteins NS1 and the 11 kDa protein. 7. Energy metabolism: Dysregulation of respiratory chain complex expression, activity and ROS production may be altered in CVB3- and PVB19-mediated myocarditis. 8. Ion channel modulation: CVB3-expression was indicated to alter calcium and potassium currents in Xenopus laevis oocytes and rodent cardiomyocytes. The phospholipase 2-like activity of PVB19 may alter several calcium, potassium and sodium channels. By understanding the general pathophysiological mechanisms of well-studied myocarditis-linked viruses, we might be provided with a guideline to handle other less-studied human viruses.
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Infecciones por Coxsackievirus/inmunología , Interacciones Huésped-Patógeno/inmunología , Miocarditis , Infecciones por Parvoviridae/inmunología , Parvovirus B19 Humano/fisiología , Replicación Viral , Infecciones por Coxsackievirus/patología , Humanos , Miocarditis/inmunología , Miocarditis/patología , Miocitos Cardíacos/inmunología , Miocitos Cardíacos/patología , Miocitos Cardíacos/virología , Infecciones por Parvoviridae/patología , Receptores Virales/inmunologíaRESUMEN
OBJECTIVES: This study is to investigate whether the cardiac microvascular endothelial cells (CMECs) can regulate the autophagy of cardiomyocytes (CMs) by secreting lncRNA-ANRIL/miR-181b exosomes, thus participating in the occurrence of uremic cardiovascular disease (CVD). METHODS: A 5/6 nephrectomy uremia model was established, with the mice injected with ANRIL-shRNA lentivirus vector, miR-181b agomir, and related control reagents, containing the serum creatinine and urea nitrogen measured. The renal tissue sections of mice were stained with Periodic Acid-Schiff (PAS), TUNEL, and Hematoxylin-Eosin (HE) performed on myocardial tissue sections of mice. ANRIL-shRNA, miR-181b mimics, and related control reagents were transfected into CMECs, in which the exosomes were extracted and co-cultured with CMs. The expressions of ANRIL, miR-181b and ATG5 were detected by qRT-PCR, and the expressions of autophagy related proteins by Western blot, as well as the binding of ANRIL and miR-181b by the double luciferase reporter gene experiment. RESULTS: ANRIL down-regulation or miR-181b up-regulation can increase the weight of mice with uremia, as well as the expressions of p62 and miR-181b, and reduce the content of serum creatinine and urea nitrogen, the damage of kidney and myocardial tissues, the number of apoptotic cells in myocardial tissues, as well as the expressions of ANRIL, ATG5, Beclin1, and LC3. CMs can absorb the exosomes of CMECs. Compared with IS+ CMEC-Exo group, the expressions of ANRIL and ATG5 in CMs of IS+ CMEC-Exo + sh lncRNA ANRIL and IS+CMEC-Exo+miR-181b mimics groups was down-regulated, as well as the expressions of ATG5, Beclin1, and LC3, while miR-181b expression was up-regulated as well as P62 expression. CONCLUSIONS: CMECs can regulate autophagy of CMs by releasing exosomes containing ANRIL and miR-181b.
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Proteína 5 Relacionada con la Autofagia/genética , Autofagia/genética , MicroARNs/metabolismo , ARN Largo no Codificante/metabolismo , Uremia/inmunología , Animales , Proteína 5 Relacionada con la Autofagia/metabolismo , Vasos Coronarios/citología , Vasos Coronarios/metabolismo , Modelos Animales de Enfermedad , Regulación hacia Abajo/inmunología , Células Endoteliales/citología , Células Endoteliales/metabolismo , Exosomas/metabolismo , Humanos , Masculino , Ratones , MicroARNs/genética , Microvasos/citología , Miocardio/citología , Miocardio/inmunología , Miocardio/patología , Miocitos Cardíacos/inmunología , Miocitos Cardíacos/patología , ARN Largo no Codificante/genética , Regulación hacia Arriba/inmunología , Uremia/genética , Uremia/patologíaRESUMEN
OBJECTIVE: Rheumatoid arthritis (RA) is a kind of chronic inflammatory disease characterized by the release of inflammatory cytokines and cardiomyocyte apoptosis, which lead to increased riskfor heart diseases. This study aims to explore the possible effect and mechanism of Celastrol on RA induced cardiac impairments in rats. METHODS: Collagen induced RA wistar rat models (CIA) were established for the measurement on secondary foot swelling degree, polyarthritis index score, spleen and thymus index. Pathological morphology was observed using H&E staining. Heart fibrosis was measured after Sirius red staining, while cell apoptosis was determined by TUNEL staining. For in vitro experiments, rat cardiomyocytes were isolated to determine the inflammatory cytokine secretion and cell apoptosis using ELISA and flow cytometry, respectively. Protein expressions of related index and autophagy were detected by Western blot and immunofluorescence. RESULTS: CIA rat model was successfully established and characterized by severe secondary foot swelling degree, and increased polyarthritis index score and spleen and thymus index. Synovial hyperplasia, disordered cardiomyocytes, cell infiltration and fibrosis were also observed in CIA rat model. Compared with CIA model, Celastrol treatment could suppress the release of inflammatory cytokines, including TNF-α, IL-6, IL-1ß, as well as inhibiting the expressions of Bax, cleaved caspase3, collagen I, collagen III and α-SMA. In addition to that, Celastrol treatment can attenuate cell apoptosis and fibrosis of cardiomyocytes and elevate Bcl-2 expression. RA induced cell autophagy can be suppressed by Celastrol through inhibiting the activation of TLR2/HMGB1 signal pathway. CONCLUSION: Celastrol can regulate TLR2/HMGB1 signal pathway to suppress autophagy and therefore exert cardioprotective effect in RA.
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Artritis Reumatoide/complicaciones , Autofagia/efectos de los fármacos , Cardiotónicos/farmacología , Proteína HMGB1/metabolismo , Cardiopatías/prevención & control , Triterpenos Pentacíclicos/farmacología , Receptor Toll-Like 2/metabolismo , Animales , Apoptosis/efectos de los fármacos , Artritis Reumatoide/inmunología , Artritis Reumatoide/metabolismo , Biomarcadores/metabolismo , Western Blotting , Cardiotónicos/uso terapéutico , Citocinas/metabolismo , Ensayo de Inmunoadsorción Enzimática , Técnica del Anticuerpo Fluorescente , Cardiopatías/etiología , Cardiopatías/metabolismo , Cardiopatías/patología , Etiquetado Corte-Fin in Situ , Ratones Endogámicos DBA , Miocitos Cardíacos/efectos de los fármacos , Miocitos Cardíacos/inmunología , Miocitos Cardíacos/metabolismo , Miocitos Cardíacos/patología , Triterpenos Pentacíclicos/uso terapéutico , Distribución Aleatoria , Ratas , Ratas Wistar , Transducción de Señal/efectos de los fármacosRESUMEN
OBJECTIVE: The NOD-like receptor protein 3 (NOD-like receptor protein 3, NLRP3) inflammasome is associated with many physiological processes related to aging. We investigated whether NLRP3 inflammasome activation contributes to the pathogenesis of cardiocytes aging dissected the underlying mechanism. METHODS: H9c2 cells were treated with different concentrations of D-galactose (D-gal, 0, 2, 10 and 50 g/L) for 24 hours. The cytochemical staining, flow cytometry and fluorescence microscope analysis were employed to detect the ß-galactosidase (ß-gal) activity. Western blot analysis was used to detect the age-associated proteins (P53, P21) and NLRP3 inflammasome proteins [NLRP3, apoptosis-associated speck-like protein (ASC)]. Confocal fluorescent images were applied to capture the colocalization of NLRP3 and caspase-1. Intracellular reactive oxygen species (ROS) was measured using 2'7'-dichlorodihydrofluorescein diacetate (DCFH-DA) by flow cytometry and visualized using a fluorescence microscope. The IL-1ß, IL-18 and lactate dehydrogenase (LDH) release were also detected. RESULTS: D-gal induced-H9c2 cells caused cardiocytes' aging changes (ß-gal staining, CellEvent™ Senescence Green staining, P53, P21) in a concentration-dependent manner. NLRP3 inflammasomes were activated, IL-1ß, IL-18 and LDH release and ROS generation were increased in the cardiocytes aging progress. When MCC950 inhibited NLRP3 inflammasomes, it attenuated the cardiocytes aging, yet the ROS generation was similar. Inhibition of ROS by NAC attenuated cardiocytes aging and inhibited the NLRP3 inflammasome activation at the same time. NLRP3 inflammasome activation by nigericin-induced cardiocytes cells aging progress. CONCLUSIONS: NLRP3 inflammasome activation contributes to the pathogenesis of cardiocytes aging, and ROS generation may serve as a potential mechanism by which NLRP3 inflammasome is activated.
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Envejecimiento/inmunología , Inflamasomas/inmunología , Miocitos Cardíacos/inmunología , Proteína con Dominio Pirina 3 de la Familia NLR/inmunología , Envejecimiento/genética , Animales , Caspasa 1/genética , Caspasa 1/inmunología , Senescencia Celular , Humanos , Inflamasomas/genética , Miocitos Cardíacos/citología , Proteína con Dominio Pirina 3 de la Familia NLR/genética , Ratas , Especies Reactivas de Oxígeno/inmunologíaRESUMEN
Purpose: Hydrogen (H2) is an antioxidant with anti-inflammatory and apoptosis functions.This study aimed to estimate the effects of H2 on acute myocardial infarction (AMI) in rats and its association with the inhibition of oxidative stress and cardiomyocyte pyroptosis. Methods: Sixty-four rats were randomly divided into three groups (Sham, AMI, and H2). The left anterior descending coronary artery (LAD) of rats in the AMI and H2 groups was ligated, while rats in the Sham group were threaded without ligation. In addition, 2% H2 was administered by inhalation for 24 h after ligation in the H2 group. Transthoracic echocardiography was performed after H2 inhalation, followed by collection of the serum and cardiac tissue of all rats. Results: H2 inhalation ameliorated the cardiac dysfunction, infarct size and inflammatory cell infiltration caused by AMI. Meanwhile, H2 inhalation reduced the concentration of serum Troponin I (TnI), brain natriuretic peptide (BNP), reactive oxygen species (ROS), cardiac malondialdehyde (MDA), and 8-OHdG. In addition, H2 inhalation inhibited cardiac inflammation and pyroptosis relative proteins expression. Conclusion: H2 effectively promoted heart functions in AMI rats by regulating oxidative stress and pyroptosis.
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Antioxidantes/administración & dosificación , Hidrógeno/administración & dosificación , Infarto del Miocardio/tratamiento farmacológico , Daño por Reperfusión Miocárdica/prevención & control , Administración por Inhalación , Animales , Modelos Animales de Enfermedad , Ecocardiografía , Humanos , Inflamasomas/antagonistas & inhibidores , Inflamasomas/metabolismo , Masculino , Infarto del Miocardio/complicaciones , Infarto del Miocardio/inmunología , Infarto del Miocardio/patología , Daño por Reperfusión Miocárdica/diagnóstico , Daño por Reperfusión Miocárdica/inmunología , Daño por Reperfusión Miocárdica/patología , Miocitos Cardíacos/efectos de los fármacos , Miocitos Cardíacos/inmunología , Miocitos Cardíacos/patología , Proteína con Dominio Pirina 3 de la Familia NLR/antagonistas & inhibidores , Proteína con Dominio Pirina 3 de la Familia NLR/metabolismo , Estrés Oxidativo/efectos de los fármacos , Estrés Oxidativo/inmunología , Piroptosis/efectos de los fármacos , Piroptosis/inmunología , Ratas , Especies Reactivas de Oxígeno/metabolismoRESUMEN
Adiponectin is a small peptide secreted and a key component of the endocrine system and immune system. Although globular adiponectin protects myocardial ischemia/reperfusion-induced cardiomyocyte injury, the protective mechanisms remain largely unresolved. Using a neonatal rat ventricular myocyte hypoxia/reoxygenation model, we investigated the role of its potential mechanisms of necroptosis in globular adiponectin-mediated protection in hypoxia/reoxygenation-induced cardiomyocyte injury as compared to apoptosis. We found that globular adiponectin treatment attenuated cardiomyocyte injury as indicated by increased cell viability and reduced lactate dehydrogenase release following hypoxia/reoxygenation. Immunofluorescence staining and Western blotting demonstrated that both necroptosis and apoptosis were triggered by hypoxia/reoxygenation and diminished by globular adiponectin. Necrostatin-1 (RIP1-specific inhibitor) and Z-VAD-FMK (pan-caspase inhibitor) only mimicked the inhibition of necroptosis and apoptosis, respectively, by globular adiponectin in hypoxia/reoxygenation-treated cardiomyocytes. Globular adiponectin attenuated reactive oxygen species production, oxidative damage, and p38MAPK and NF-κB signaling, all important for necroptosis and apoptosis. Collectively, our study suggests that globular adiponectin inhibits hypoxia/reoxygenation-induced necroptosis and apoptosis in cardiomyocytes probably by reducing oxidative stress and interrupting p38MAPK signaling.