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Vaccination is one of the most effective prophylactic public health interventions for the prevention of infectious diseases such as coronavirus disease (COVID-19). Considering the ongoing need for new COVID-19 vaccines, it is crucial to modify our approach and incorporate more conserved regions of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) to effectively address emerging viral variants. The nucleocapsid protein is a structural protein of SARS-CoV-2 that is involved in replication and immune responses. Furthermore, this protein offers significant advantages owing to the minimal accumulation of mutations over time and the inclusion of key T-cell epitopes critical for SARS-CoV-2 immunity. A novel strategy that may be suitable for the new generation of vaccines against COVID-19 is to use a combination of antigens, including the spike and nucleocapsid proteins, to elicit robust humoral and potent cellular immune responses, along with long-lasting immunity. The strategic use of multiple antigens aims to enhance vaccine efficacy and broaden protection against viruses, including their variants. The immune response against the nucleocapsid protein from other coronavirus is long-lasting, and it can persist up to 11 years post-infection. Thus, the incorporation of nucleocapsids (N) into vaccine design adds an important dimension to vaccination efforts and holds promise for bolstering the ability to combat COVID-19 effectively. In this review, we summarize the preclinical studies that evaluated the use of the nucleocapsid protein as antigen. This study discusses the use of nucleocapsid alone and its combination with spike protein or other proteins of SARS-CoV-2.
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Vacunas contra la COVID-19 , COVID-19 , Proteínas de la Nucleocápside de Coronavirus , SARS-CoV-2 , Humanos , Vacunas contra la COVID-19/inmunología , SARS-CoV-2/inmunología , COVID-19/prevención & control , COVID-19/inmunología , Proteínas de la Nucleocápside de Coronavirus/inmunología , Proteínas de la Nucleocápside de Coronavirus/genética , Inmunogenicidad Vacunal , Animales , Fosfoproteínas/inmunología , Glicoproteína de la Espiga del Coronavirus/inmunología , Glicoproteína de la Espiga del Coronavirus/genética , Epítopos de Linfocito T/inmunología , Anticuerpos Antivirales/inmunología , Proteínas de la Nucleocápside/inmunologíaRESUMEN
Viruses have a wide repertoire of molecular strategies that focus on their replication or the facilitation of different stages of the viral cycle. One of these strategies is mediated by the activity of viroporins, which are multifunctional viral proteins that, upon oligomerization, exhibit ion channel properties with mild ion selectivity. Viroporins facilitate multiple processes, such as the regulation of immune response and inflammasome activation through the induction of pore formation in various cell organelle membranes to facilitate the escape of ions and the alteration of intracellular homeostasis. Viroporins target diverse membranes (such as the cellular membrane), endoplasmic reticulum, and mitochondria. Cumulative data regarding the importance of mitochondria function in multiple processes, such as cellular metabolism, energy production, calcium homeostasis, apoptosis, and mitophagy, have been reported. The direct or indirect interaction of viroporins with mitochondria and how this interaction affects the functioning of mitochondrial cells in the innate immunity of host cells against viruses remains unclear. A better understanding of the viroporin-mitochondria interactions will provide insights into their role in affecting host immune signaling through the mitochondria. Thus, in this review, we mainly focus on descriptions of viroporins and studies that have provided insights into the role of viroporins in hijacked mitochondria.
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Proteínas Viroporinas , Virus , Proteínas Viroporinas/metabolismo , Proteínas Virales/metabolismo , Canales Iónicos/metabolismo , Inmunidad InnataRESUMEN
The limited regenerative capacity of the human heart contributes to high morbidity and mortality worldwide. In contrast, zebrafish exhibit robust regenerative capacity, providing a powerful model for studying how to overcome intrinsic epigenetic barriers maintaining cardiac homeostasis and initiate regeneration. Here, we present a comprehensive analysis of the histone modifications H3K4me1, H3K4me3, H3K27me3 and H3K27ac during various stages of zebrafish heart regeneration. We found a vast gain of repressive chromatin marks one day after myocardial injury, followed by the acquisition of active chromatin characteristics on day four and a transition to a repressive state on day 14, and identified distinct transcription factor ensembles associated with these events. The rapid transcriptional response involves the engagement of super-enhancers at genes implicated in extracellular matrix reorganization and TOR signaling, while H3K4me3 breadth highly correlates with transcriptional activity and dynamic changes at genes involved in proteolysis, cell cycle activity, and cell differentiation. Using loss- and gain-of-function approaches, we identified transcription factors in cardiomyocytes and endothelial cells influencing cardiomyocyte dedifferentiation or proliferation. Finally, we detected significant evolutionary conservation between regulatory regions that drive zebrafish and neonatal mouse heart regeneration, suggesting that reactivating transcriptional and epigenetic networks converging on these regulatory elements might unlock the regenerative potential of adult human hearts.
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Cromatina , Redes Reguladoras de Genes , Corazón , Animales , Humanos , Ratones , Diferenciación Celular , Cromatina/metabolismo , Cromatina/genética , Epigénesis Genética , Código de Histonas , Histonas/metabolismo , Miocitos Cardíacos/metabolismo , Miocitos Cardíacos/citología , Regeneración/genética , Factores de Transcripción/metabolismo , Factores de Transcripción/genética , Pez Cebra/genéticaRESUMEN
Fibrosis is a hallmark of chronic disease. Although fibroblasts are involved, it is unclear to what extent endothelial cells also might contribute. We detected increased expression of the transcription factor Sox9 in endothelial cells in several different mouse fibrosis models. These models included systolic heart failure induced by pressure overload, diastolic heart failure induced by high-fat diet and nitric oxide synthase inhibition, pulmonary fibrosis induced by bleomycin treatment, and liver fibrosis due to a choline-deficient diet. We also observed up-regulation of endothelial SOX9 in cardiac tissue from patients with heart failure. To test whether SOX9 induction was sufficient to cause disease, we generated mice with endothelial cell-specific overexpression of Sox9, which promoted fibrosis in multiple organs and resulted in signs of heart failure. Endothelial Sox9 deletion prevented fibrosis and organ dysfunction in the two mouse models of heart failure as well as in the lung and liver fibrosis mouse models. Bulk and single-cell RNA sequencing of mouse endothelial cells across multiple vascular beds revealed that SOX9 induced extracellular matrix, growth factor, and inflammatory gene expression, leading to matrix deposition by endothelial cells. Moreover, mouse endothelial cells activated neighboring fibroblasts that then migrated and deposited matrix in response to SOX9, a process partly mediated by the secreted growth factor CCN2, a direct SOX9 target; endothelial cell-specific Sox9 deletion reversed these changes. These findings suggest a role for endothelial SOX9 as a fibrosis-promoting factor in different mouse organs during disease and imply that endothelial cells are an important regulator of fibrosis.
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Insuficiencia Cardíaca , Factores de Transcripción , Animales , Humanos , Ratones , Modelos Animales de Enfermedad , Células Endoteliales , Fibrosis , Péptidos y Proteínas de Señalización Intercelular , Cirrosis Hepática/complicaciones , Factor de Transcripción SOX9/genéticaRESUMEN
Once regarded as inert organelles with limited and ill-defined roles, lipid droplets (LDs) have emerged as dynamic entities with multifaceted functions within the cell. Recent research has illuminated their pivotal role as primary energy reservoirs in the form of lipids, capable of being metabolized to meet cellular energy demands. Their high dynamism is underscored by their ability to interact with numerous cellular organelles, notably the endoplasmic reticulum (the site of LD genesis) and mitochondria, which utilize small LDs for energy production. Beyond their contribution to cellular bioenergetics, LDs have been associated with viral infections. Evidence suggests that viruses can co-opt LDs to facilitate their infection cycle. Furthermore, recent discoveries highlight the role of LDs in modulating the host's immune response. Observations of altered LD levels during viral infections suggest their involvement in disease pathophysiology, potentially through production of proinflammatory mediators using LD lipids as precursors. This review explores these intriguing aspects of LDs, shedding light on their multifaceted nature and implications in viral interactions and disease development.
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Vascular networks form, remodel, and mature under the influence of both fluid shear stress (FSS) and soluble factors. Physiological FSS promotes and maintains vascular stability via synergy with bone morphogenic proteins 9 and 10 (BMP9 and BMP10). Conversely, mutation of the BMP receptors activin-like kinase 1 (ALK1), endoglin (ENG), or the downstream effector, SMAD family member 4 (SMAD4) leads to hereditary hemorrhagic telangiectasia (HHT), characterized by fragile and leaky arterial-venous malformations (AVMs). How endothelial cells (ECs) integrate FSS and BMP signals in vascular development and homeostasis and how mutations give rise to vascular malformations is not well understood. Here, we aimed to elucidate the mechanism of synergy between FSS and SMAD signaling in vascular stability and how disruption of this synergy leads to AVMs. We found that loss of Smad4 increased the sensitivity of ECs to flow by lowering the FSS set point, with resulting AVMs exhibiting features of excessive flow-mediated morphological responses. Mechanistically, loss of SMAD4 disinhibits flow-mediated KLF4-TIE2-PI3K/Akt signaling, leading to cell cycle progression-mediated loss of arterial identity due to KLF4-mediated repression of cyclin dependent Kinase (CDK) inhibitors CDKN2A and CDKN2B. Thus, AVMs caused by Smad4 deletion are characterized by chronic high flow remodeling with excessive EC proliferation and loss of arterial identity as triggering events.
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Malformaciones Arteriovenosas , Telangiectasia Hemorrágica Hereditaria , Ratones , Animales , Malformaciones Arteriovenosas/genética , Malformaciones Arteriovenosas/metabolismo , Células Endoteliales/metabolismo , Fosfatidilinositol 3-Quinasas/metabolismo , Ratones Noqueados , Telangiectasia Hemorrágica Hereditaria/genética , Proteínas Morfogenéticas Óseas/genéticaRESUMEN
The nuclear factor erythroid 2-related factor 2 (NRF2) is a transcription factor that plays a critical role in the xenobiotic and stress responses. During viral infection, NRF2 can modulate the host metabolism and innate immunity; however, the most common activity of NRF2 in viral diseases is controlling reactive oxygen species (ROS). The Zika virus (ZIKV) is involved in a vertical infection in pregnancy, with reported fetal health consequences. However, the possibility that ZIKV regulates NRF2 expression in placental trophoblasts has not been investigated. In this report, we evaluated the upregulation of NRF2 and antioxidant enzymes in a trophoblast-like cell. These findings could help us understand the antioxidant mechanism underlying the ZIKV infection in the placenta during pregnancy.
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Infección por el Virus Zika , Virus Zika , Femenino , Humanos , Embarazo , Antioxidantes/metabolismo , Factor 2 Relacionado con NF-E2/genética , Factor 2 Relacionado con NF-E2/metabolismo , Placenta , Trofoblastos/metabolismo , Virus Zika/genéticaRESUMEN
Background: Small cell lung cancer (SCLC) is an extremely aggressive cancer type with a patient median survival of 6-12 months. Epidermal growth factor (EGF) signaling plays an important role in triggering SCLC. In addition, growth factor-dependent signals and alpha-, beta-integrin (ITGA, ITGB) heterodimer receptors functionally cooperate and integrate their signaling pathways. However, the precise role of integrins in EGF receptor (EGFR) activation in SCLC remains elusive. Methods: We analyzed human precision-cut lung slices (hPCLS), retrospectively collected human lung tissue samples and cell lines by classical methods of molecular biology and biochemistry. In addition, we performed RNA-sequencing-based transcriptomic analysis in human lung cancer cells and human lung tissue samples, as well as high-resolution mass spectrometric analysis of the protein cargo from extracellular vesicles (EVs) that were isolated from human lung cancer cells. Results: Our results demonstrate that non-canonical ITGB2 signaling activates EGFR and RAS/MAPK/ERK signaling in SCLC. Further, we identified a novel SCLC gene expression signature consisting of 93 transcripts that were induced by ITGB2, which may be used for stratification of SCLC patients and prognosis prediction of LC patients. We also found a cell-cell communication mechanism based on EVs containing ITGB2, which were secreted by SCLC cells and induced in control human lung tissue RAS/MAPK/ERK signaling and SCLC markers. Conclusions: We uncovered a mechanism of ITGB2-mediated EGFR activation in SCLC that explains EGFR-inhibitor resistance independently of EGFR mutations, suggesting the development of therapies targeting ITGB2 for patients with this extremely aggressive lung cancer type.
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Neoplasias Pulmonares , Carcinoma Pulmonar de Células Pequeñas , Humanos , Carcinoma Pulmonar de Células Pequeñas/genética , Estudios Retrospectivos , Receptores ErbB/metabolismo , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/metabolismo , Integrinas/genética , MutaciónRESUMEN
Despite all successful efforts to develop a COVID-19 vaccine, the need to evaluate alternative antigens to produce next-generation vaccines is imperative to target emerging variants. Thus, the second generation of COVID-19 vaccines employ more than one antigen from severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) to induce an effective and lasting immune response. Here, we analyzed the combination of two SARS-CoV-2 viral antigens that could elicit a more durable immune response in both T- and B-cells. The nucleocapsid (N) protein, Spike protein S1 domain, and receptor binding domain (RBD) of the SARS-CoV-2 spike surface glycoproteins were expressed and purified in a mammalian expression system, taking into consideration the posttranscriptional modifications and structural characteristics. The immunogenicity of these combined proteins was evaluated in a murine model. Immunization combining S1 or RBD with the N protein induced higher levels of IgG antibodies, increased the percentage of neutralization, and elevated the production of cytokines TNF-α, IFN-γ, and IL-2 compared to the administration of a single antigen. Furthermore, sera from immunized mice recognized alpha and beta variants of SARS-CoV-2, which supports ongoing clinical results on partial protection in vaccinated populations, despite mutations. This study identifies potential antigens for second-generation COVID-19 vaccines.
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(1) Background: Diabetic cardiomyopathy is a major health problem worldwide. CTRP9, a secreted glycoprotein, is mainly expressed in cardiac endothelial cells and becomes downregulated in mouse models of diabetes mellitus; (2) Methods: In this study, we investigated the impact of CTRP9 on early stages of diabetic cardiomyopathy induced by 12 weeks of high-fat diet; (3) Results: While the lack of CTRP9 in knock-out mice aggravated insulin resistance and triggered diastolic left ventricular dysfunction, AAV9-mediated cardiac CTRP9 overexpression ameliorated cardiomyopathy under these conditions. At this early disease state upon high-fat diet, no fibrosis, no oxidative damage and no lipid deposition were identified in the myocardium of any of the experimental groups. Mechanistically, we found that CTRP9 is required for insulin-dependent signaling, cardiac glucose uptake in vivo and oxidative energy production in cardiomyocytes. Extensive RNA sequencing from myocardial tissue of CTRP9-overexpressing and knock-out as well as respective control mice revealed that CTRP9 acts as an anti-inflammatory mediator in the myocardium. Hence, CTRP9 knock-out exerted more, while CTRP9-overexpressing mice showed less leukocytes accumulation in the heart during high-fat diet; (4) Conclusions: In summary, endothelial-derived CTRP9 plays a prominent paracrine role to protect against diabetic cardiomyopathy and might constitute a therapeutic target.
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Diabetes Mellitus , Cardiomiopatías Diabéticas , Resistencia a la Insulina , Ratones , Animales , Cardiomiopatías Diabéticas/metabolismo , Complemento C1q/metabolismo , Células Endoteliales/metabolismo , Adiponectina/metabolismo , Factor de Necrosis Tumoral alfa/metabolismo , Miocitos Cardíacos/metabolismo , Inflamación/patología , Ratones Noqueados , Diabetes Mellitus/metabolismo , Glicoproteínas/genética , Glicoproteínas/metabolismoRESUMEN
Our understanding of how active learning affects different groups of students is still developing. One group often overlooked in higher education research is students with disabilities. Two of the most commonly occurring disabilities on college campuses are attention-deficit/hyperactivity disorder (ADHD) and specific learning disorders (SLD). We investigated how the incorporation of active-learning practices influences the learning and self-advocacy experiences of students with ADHD and/or SLD (ADHD/SLD) in undergraduate science, technology, engineering, and mathematics (STEM) courses. Semistructured interviews were conducted with 25 STEM majors with ADHD/SLD registered with a campus disability resource center at a single university, and data were analyzed using qualitative methods. Participants described how they perceived active learning in their STEM courses to support or hinder their learning and how active learning affected their self-advocacy. Many of the active-learning barriers could be attributed to issues related to fidelity of implementation of a particular active-learning strategy and limited awareness of universal design for learning. Active learning was also reported to influence self-advocacy for some participants, and examples of self-advocacy in active-learning STEM courses were identified. Defining the supports and barriers perceived by students with ADHD/SLD is a crucial first step in developing more-inclusive active-learning STEM courses. Suggestions for research and teaching are provided.
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Trastorno por Déficit de Atención con Hiperactividad , Discapacidades para el Aprendizaje , Humanos , Aprendizaje Basado en Problemas , Estudiantes , Matemática , Tecnología/educaciónRESUMEN
Tight control of cell fate choices is crucial for normal development. Here we show that lamin A/C plays a key role in chromatin organization in embryonic stem cells (ESCs), which safeguards naïve pluripotency and ensures proper cell fate choices during cardiogenesis. We report changes in chromatin compaction and localization of cardiac genes in Lmna-/- ESCs resulting in precocious activation of a transcriptional program promoting cardiomyocyte versus endothelial cell fate. This is accompanied by premature cardiomyocyte differentiation, cell cycle withdrawal and abnormal contractility. Gata4 is activated by lamin A/C loss and Gata4 silencing or haploinsufficiency rescues the aberrant cardiovascular cell fate choices induced by lamin A/C deficiency. We uncover divergent functions of lamin A/C in naïve pluripotent stem cells and cardiomyocytes, which have distinct contributions to the transcriptional alterations of patients with LMNA-associated cardiomyopathy. We conclude that disruption of lamin A/C-dependent chromatin architecture in ESCs is a primary event in LMNA loss-of-function cardiomyopathy.
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Cromatina , Lamina Tipo A , Humanos , Lamina Tipo A/metabolismo , Cromatina/metabolismo , Diferenciación Celular/genética , Células Madre Embrionarias/metabolismo , Miocitos Cardíacos/metabolismoRESUMEN
Dengue and Zika viruses cocirculate annually in endemic areas of Mexico, causing outbreaks of different magnitude and severity every year, suggesting a continuous selection of Flavivirus variants with variable phenotypes of transmissibility and virulence. To evaluate if Flavivirus variants with different phenotypes cocirculate during outbreaks, we isolated dengue and Zika viruses from blood samples of febrile patients from Oaxaca City during the 2016 and 2019 epidemic years. We compared their replication kinetics in human cells, susceptibility to type I interferon antiviral response, and the accumulation of subgenomic RNA on infected cells. We observed correlations between type I interferon susceptibility and subgenomic RNA accumulation, with high hematocrit percentage and thrombocytopenia. Our results suggest that Flaviviruses that cocirculate in Oaxaca, Mexico, have variable sensitivity to the antiviral activity of type I interferons, and this phenotypic trait correlates with the severity of the disease.
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Dengue , Flavivirus , Interferón Tipo I , Infección por el Virus Zika , Virus Zika , Antivirales , Flavivirus/genética , Humanos , México/epidemiología , ARN Viral/genética , Índice de Severidad de la Enfermedad , Replicación Viral , Virus Zika/genéticaRESUMEN
To identify cellular mechanisms responsible for pressure overload triggered heart failure, we isolated cardiomyocytes, endothelial cells, and fibroblasts as most abundant cell types from mouse hearts in the subacute and chronic stages after transverse aortic constriction (TAC) and performed RNA-sequencing. We detected highly cell-type specific transcriptional responses with characteristic time courses and active intercellular communication. Cardiomyocytes after TAC exerted an early and sustained upregulation of inflammatory and matrix genes and a concomitant suppression of metabolic and ion channel genes. Fibroblasts, in contrast, showed transient early upregulation of inflammatory and matrix genes and downregulation of angiogenesis genes, but sustained induction of cell cycle and ion channel genes during TAC. Endothelial cells transiently induced cell cycle and extracellular matrix genes early after TAC, but exerted a long-lasting upregulation of inflammatory genes. As we found that matrix production by multiple cell types triggers pathological cellular responses, it might serve as a future therapeutic target.
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Hyperactivity of the sympathetic nervous system is a major driver of cardiac remodeling, exerting its effects through both α-, and ß-adrenoceptors (α-, ß-ARs). As the relative contribution of subtype α1-AR to cardiac stress responses remains poorly investigated, we subjected mice to either subcutaneous perfusion with the ß-AR agonist isoprenaline (ISO, 30 mg/kg × day) or to a combination of ISO and the stable α1-AR agonist phenylephrine (ISO/PE, 30 mg/kg × day each). Telemetry analysis revealed similar hemodynamic responses under both ISO and ISO/PE treatment i.e., permanently increased heart rates and only transient decreases in mean blood pressure during the first 24 h. Echocardiography and single cell analysis after 1 week of exposure showed that ISO/PE-, but not ISO-treated animals established α1-AR-mediated inotropic responsiveness to acute adrenergic stimulation. Morphologically, additional PE perfusion limited concentric cardiomyocyte growth and enhanced cardiac collagen deposition during 7 days of treatment. Time-course analysis demonstrated a diverging development in transcriptional patterns at day 4 of treatment i.e., increased expression of selected marker genes Xirp2, Nppa, Tgfb1, Col1a1, Postn under chronic ISO/PE treatment which was either less pronounced or absent in the ISO group. Transcriptome analyses at day 4 via RNA sequencing demonstrated that additional PE treatment caused a marked upregulation of genes allocated to extracellular matrix and fiber organization along with a more pronounced downregulation of genes involved in metabolic processes, muscle adaptation and cardiac electrophysiology. Consistently, transcriptome changes under ISO/PE challenge more effectively recapitulated early transcriptional alterations in pressure overload-induced experimental heart failure and in human hypertrophic cardiomyopathy.
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Corazón , Receptores Adrenérgicos alfa 1 , Animales , Isoproterenol/farmacología , Ratones , Fenilefrina/farmacología , Receptores Adrenérgicos alfa 1/genética , Receptores Adrenérgicos alfa 1/metabolismo , Receptores Adrenérgicos betaRESUMEN
Cachexia is associated with poor prognosis in chronic heart failure patients, but the underlying mechanisms of cachexia triggered disease progression remain poorly understood. Here, we investigate whether the dysregulation of myokine expression from wasting skeletal muscle exaggerates heart failure. RNA sequencing from wasting skeletal muscles of mice with heart failure reveals a reduced expression of Ostn, which encodes the secreted myokine Musclin, previously implicated in the enhancement of natriuretic peptide signaling. By generating skeletal muscle specific Ostn knock-out and overexpressing mice, we demonstrate that reduced skeletal muscle Musclin levels exaggerate, while its overexpression in muscle attenuates cardiac dysfunction and myocardial fibrosis during pressure overload. Mechanistically, Musclin enhances the abundance of C-type natriuretic peptide (CNP), thereby promoting cardiomyocyte contractility through protein kinase A and inhibiting fibroblast activation through protein kinase G signaling. Because we also find reduced OSTN expression in skeletal muscle of heart failure patients, augmentation of Musclin might serve as therapeutic strategy.
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Caquexia/genética , Fibrosis Endomiocárdica/genética , Insuficiencia Cardíaca/genética , Proteínas Musculares/genética , Músculo Esquelético/metabolismo , Atrofia Muscular/genética , Factores de Transcripción/genética , 2',3'-Nucleótido Cíclico 3'-Fosfodiesterasa/genética , 2',3'-Nucleótido Cíclico 3'-Fosfodiesterasa/metabolismo , Anciano , Anciano de 80 o más Años , Animales , Caquexia/metabolismo , Caquexia/fisiopatología , Caquexia/prevención & control , Estudios de Casos y Controles , Proteínas Quinasas Dependientes de AMP Cíclico/genética , Proteínas Quinasas Dependientes de AMP Cíclico/metabolismo , Proteínas Quinasas Dependientes de GMP Cíclico/genética , Proteínas Quinasas Dependientes de GMP Cíclico/metabolismo , Modelos Animales de Enfermedad , Fibrosis Endomiocárdica/metabolismo , Fibrosis Endomiocárdica/fisiopatología , Fibrosis Endomiocárdica/prevención & control , Femenino , Regulación de la Expresión Génica , Insuficiencia Cardíaca/metabolismo , Insuficiencia Cardíaca/fisiopatología , Insuficiencia Cardíaca/prevención & control , Pruebas de Función Cardíaca , Humanos , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Proteínas Musculares/agonistas , Proteínas Musculares/antagonistas & inhibidores , Proteínas Musculares/deficiencia , Atrofia Muscular/metabolismo , Atrofia Muscular/fisiopatología , Atrofia Muscular/prevención & control , Miocardio/metabolismo , Miocardio/patología , Miocitos Cardíacos/metabolismo , Miocitos Cardíacos/patología , ARN Interferente Pequeño/genética , ARN Interferente Pequeño/metabolismo , Transducción de Señal , Factores de Transcripción/agonistas , Factores de Transcripción/antagonistas & inhibidores , Factores de Transcripción/deficienciaRESUMEN
Severe coronavirus disease 2019 (COVID-19) is characterized by lung injury, cytokine storm, and increased neutrophil-to-lymphocyte ratio (NLR). Current therapies focus on reducing viral replication and inflammatory responses, but no specific treatment exists to prevent the development of severe COVID-19 in infected individuals. Angiotensin-converting enzyme-2 (ACE2) is the receptor for SARS-CoV-2, the virus causing COVID-19, but it is also critical for maintaining the correct functionality of lung epithelium and endothelium. Coronaviruses induce activation of a disintegrin and metalloprotease 17 (ADAM17) and shedding of ACE2 from the cell surface resulting in exacerbated inflammatory responses. Thus, we hypothesized that ADAM17 inhibition ameliorates COVID-19-related lung inflammation. We employed a preclinical mouse model using intratracheal instillation of a combination of polyinosinic:polycytidylic acid (poly(I:C)) and the receptor-binding domain of the SARS-CoV-2 spike protein (RBD-S) to mimic lung damage associated with COVID-19. Histologic analysis of inflamed mice confirmed the expected signs of lung injury including edema, fibrosis, vascular congestion, and leukocyte infiltration. Moreover, inflamed mice also showed an increased NLR as observed in critically ill COVID-19 patients. Administration of the ADAM17/MMP inhibitors apratastat and TMI-1 significantly improved lung histology and prevented leukocyte infiltration. Reduced leukocyte recruitment could be explained by reduced production of proinflammatory cytokines and lower levels of the endothelial adhesion molecules ICAM-1 and VCAM-1. Additionally, the NLR was significantly reduced by ADAM17/MMP inhibition. Thus, we propose inhibition of ADAM17/MMP as a novel promising treatment strategy in SARS-CoV-2-infected individuals to prevent the progression toward severe COVID-19.
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Tratamiento Farmacológico de COVID-19 , Lesión Pulmonar , Proteína ADAM17 , Enzima Convertidora de Angiotensina 2 , Animales , Modelos Animales de Enfermedad , Humanos , Lesión Pulmonar/etiología , Lesión Pulmonar/prevención & control , Metaloproteinasas de la Matriz , Ratones , SARS-CoV-2 , Glicoproteína de la Espiga del CoronavirusRESUMEN
The coronavirus disease 2019 (COVID-19) pandemic has reached an unprecedented level. There is a strong demand for diagnostic and serological supplies worldwide, making it necessary for countries to establish their own technologies to produce high-quality biomolecules. The two main viral antigens used for the diagnostics for severe acute respiratory syndrome coronavirus (SARS-CoV-2) are the structural proteins spike (S) protein and nucleocapsid (N) protein. The spike protein of SARS-CoV-2 is cleaved into S1 and S2, in which the S1 subunit has the receptor-binding domain (RBD), which induces the production of neutralizing antibodies, whereas nucleocapsid is an ideal target for viral antigen-based detection. In this study, we designed plasmids, pcDNA3.1/S1 and pcDNA3.1/N, and optimized their expression of the recombinant S1 and N proteins from SARS-CoV-2 in a mammalian system. The RBD was used as a control. The antigens were successfully purified from Expi293 cells, with high yields of the S1, N, and RBD proteins. The immunogenic abilities of these proteins were demonstrated in a mouse model. Further, enzyme-linked immunosorbent assays with human serum samples showed that the SARS-CoV-2 antigens are a suitable alternative for serological assays to identify patients infected with COVID-19.