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Local renin-angiotensin signaling is present within various organ systems of the body. Angiotensin II signaling acts as a crucial mechanism of the renin-angiotensin signaling system. In this manuscript, we review current literature regarding local angiotensin II signaling in the urinary bladder. Our interest in this pathway is due to its potential role in interstitial cystitis/bladder pain syndrome and other bladder diseases, which are chronic in nature and negatively affect patients' quality of life. Current treatments for bladder diseases are generally ineffective due to our lack of understanding of their pathophysiology. We evaluate literature investigating angiotensin II signaling in the bladder across several perspectives, including local expression and production, functional properties, tissue morphology, and clinical implications. Further, we identify gaps in knowledge and suggest areas where we can improve our understanding of angiotensin II signaling in the bladder.
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Background: The renin-angiotensin system has been identified as a potential therapeutic target for posttraumatic stress disorder, although its mechanisms are not well understood. Brain angiotensin type 2 receptors (AT2Rs) are a subtype of angiotensin II receptors located in stress and anxiety-related regions, including the medial prefrontal cortex (mPFC), but their function and mechanism in the mPFC remain unexplored. Therefore, we used a combination of imaging, cre/lox, and behavioral methods to investigate mPFC-AT2R-expressing neurons in fear and stess related behavior. Methods: To characterize mPFC-AT2R-expressing neurons in the mPFC, AT2R-Cre/tdTomato male and female mice were used for immunohistochemistry. mPFC brain sections were stained with glutamatergic or interneuron markers, and density of AT2R+ cells and colocalization with each marker were quantified. To assess fear-related behaviors in AT2R-flox mice, we selectively deleted AT2R from mPFC neurons using a Cre-expressing adeno-associated virus. Mice then underwent Pavlovian auditory fear conditioning, elevated plus maze, and open field testing. Results: Immunohistochemistry results revealed that AT2R was densely expressed throughout the mPFC and primarily expressed in somatostatin interneurons in a sex-dependent manner. Following fear conditioning, mPFC-AT2R Cre-lox deletion impaired extinction and increased exploratory behavior in female but not male mice, while locomotion was unaltered by mPFC-AT2R deletion in both sexes. Conclusions: These results identify mPFC-AT2R+ neurons as a novel subgroup of somatostatin interneurons and reveal their role in regulating fear learning in a sex-dependent manner, potentially offering insights into novel therapeutic targets for posttraumatic stress disorder.
Posttraumatic stress disorder (PTSD) is a significant predictor of cardiovascular disease (CVD), although the underlying mechanisms are poorly understood. The brain renin-angiotensin system (RAS) is important for cardiovascular and emotional stress regulation and may better help understand the link between PTSD and CVD risk. Our research reveals that the brain angiotensin II type 2 receptor (AT2R) subtype is located on specific somatostatin (SOM+) interneurons in the medial prefrontal cortex (mPFC) and plays a role in fear memory extinction, particularly in females. These findings reveal a role for the mPFC-AT2R in fear-based learning and memory, offering potential insights into the mechanisms underlying the PTSD-CVD association and therapeutic strategies.
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Idiopathic pulmonary fibrosis (IPF) is a progressive and fatal fibrotic disease. Recent studies have highlighted the persistence of an intermediate state of alveolar stem cells in IPF lungs. In this study, we discovered a close correlation between the distribution pattern of intermediate alveolar stem cells and the progression of fibrotic changes. We showed that amphiregulin (AREG) expression is significantly elevated in intermediate alveolar stem cells of mouse fibrotic lungs and IPF patients. High levels of serum AREG correlate significantly with profound deteriorations in lung function in IPF patients. We demonstrated that AREG in alveolar stem cells is both required and sufficient for activating EGFR in fibroblasts, thereby driving lung fibrosis. Moreover, pharmacological inhibition of AREG using a neutralizing antibody effectively blocked the initiation and progression of lung fibrosis in mice. Our study underscores the therapeutic potential of anti-AREG antibodies in attenuating IPF progression, offering a promising strategy for treating fibrotic diseases.
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Anfirregulina , Progresión de la Enfermedad , Fibrosis Pulmonar Idiopática , Anfirregulina/metabolismo , Animales , Humanos , Fibrosis Pulmonar Idiopática/patología , Fibrosis Pulmonar Idiopática/metabolismo , Ratones , Ratones Endogámicos C57BL , Células Madre/metabolismo , Receptores ErbB/metabolismo , Fibroblastos/metabolismo , Fibroblastos/patología , Células Epiteliales Alveolares/metabolismo , Células Epiteliales Alveolares/patología , Masculino , Alveolos Pulmonares/patología , Alveolos Pulmonares/metabolismo , Anticuerpos Neutralizantes/farmacología , FemeninoRESUMEN
Background: Gestational intermittent hypoxia (GIH), a hallmark of maternal obstructive sleep apnea, sex-differentially causes hypertension and endothelial dysfunction in adult male offspring but not in females. This study investigated whether the GIH-exposed female offspring, a "protected" group against the hypertensive effects of maternal GIH exposure, exhibit increased susceptibility to hypertension and cardiovascular dysfunction when fed a high-fat high-sucrose (HFHS) diet and whether this effect could be reversed by pharmacological intervention activating the angiotensin II type 2 receptor (AT2R). Methods: Female offspring of control and GIH-exposed (10.5% O2, 8 h/d, E10-21) dams were assigned either an HFHS diet or a standard diet from 12 weeks of age. Blood pressure was monitored. At 28 weeks, a systemic CGP42112 (AT2R agonist) or saline infusion was administered through the osmotic pump. At 30 weeks, the heart was weighed and collected for H&E staining, mesenteric arteries for vascular reactivity assessment and protein analysis, and plasma for ELISA. Results: The HFHS diet induced similar increases in body weight gain and blood pressure in control and GIH female offspring. HFHS feeding did not affect heart structure, but impaired endothelial-dependent vascular relaxation with associated decreased AT2R and eNOS expression and reduced plasma bradykinin levels in both control and GIH offspring. CGP42112 administration effectively mitigated HFHS-induced hypertension and endothelial dysfunction only in control offspring, accompanied by restored AT2R, eNOS, and bradykinin levels, but not in the GIH counterparts. Conclusion: These findings suggest that GIH induces endothelial dysfunction and AT2R insensitivity in female offspring exposed to an HFHS diet.
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Idiopathic pulmonary fibrosis (IPF) is a long-term condition with an unidentified cause, and currently there are no specific treatment options available. Alveolar epithelial type II cells (AT2) constitute a heterogeneous population crucial for secreting and regenerative functions in the alveolus, essential for maintaining lung homeostasis. However, a comprehensive investigation into their cellular diversity, molecular features, and clinical implications is currently lacking. In this study, we conducted a comprehensive examination of single-cell RNA sequencing data from both normal and fibrotic lung tissues. We analyzed alterations in cellular composition between IPF and normal tissue and investigated differentially expressed genes across each cell population. This analysis revealed the presence of two distinct subpopulations of IPF-related alveolar epithelial type II cells (IR_AT2). Subsequently, three unique gene co-expression modules associated with the IR_AT2 subtype were identified through the use of hdWGCNA. Furthermore, we refined and identified IPF-related AT2-related gene (IARG) signatures using various machine learning algorithms. Our analysis demonstrated a significant association between high IARG scores in IPF patients and shorter survival times (p-value < 0.01). Additionally, we observed a negative correlation between the percent predicted diffusing capacity for lung carbon monoxide (% DLCO) and increased IARG scores (cor = -0.44, p-value < 0.05). The cross-validation findings demonstrated a high level of accuracy (AUC > 0.85, p-value < 0.01) in the prognostication of patients with IPF utilizing the identified IARG signatures. Our study has identified distinct molecular and biological features among AT2 subpopulations, specifically highlighting the unique characteristics of IPF-related AT2 cells. Importantly, our findings underscore the prognostic relevance of specific genes associated with IPF-related AT2 cells, offering valuable insights into the advancement of IPF.
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Células Epiteliales Alveolares , Biomarcadores , Fibrosis Pulmonar Idiopática , Aprendizaje Automático , Análisis de la Célula Individual , Humanos , Fibrosis Pulmonar Idiopática/metabolismo , Fibrosis Pulmonar Idiopática/genética , Fibrosis Pulmonar Idiopática/diagnóstico , Fibrosis Pulmonar Idiopática/patología , Análisis de la Célula Individual/métodos , Células Epiteliales Alveolares/metabolismo , Células Epiteliales Alveolares/patología , Perfilación de la Expresión Génica , Transcriptoma , Femenino , Masculino , PronósticoRESUMEN
Fibrosing interstitial lung diseases (FILDs), e.g., due to idiopathic pulmonary fibrosis (IPF), are chronic progressive diseases with a poor prognosis. The management of these diseases is challenging and focuses mainly on the suppression of progression with anti-fibrotic drugs. Therefore, novel FILD treatments are needed. In recent years, cell-based therapy with various stem cells has been investigated for FILD, and the use of mesenchymal stem cells (MSCs) has been widely reported and clinical studies are also ongoing. Induced pluripotent stem cells (iPSCs) have also been reported to have an anti-fibrotic effect in FILD; however, these have not been as well studied as MSCs in terms of the mechanisms and side effects. While MSCs show a potent anti-fibrotic effect, the possibility of quality differences between donors and a stable supply in the case of donor shortage or reduced proliferative capacity after cell passaging needs to be considered. The application of iPSC-derived cells has the potential to overcome these problems and may lead to consistent quality of the cell product and stable product supply. This review provides an overview of iPSCs and FILD, followed by the current status of cell-based therapy for FILD, and then discusses the possibilities and perspectives of FILD therapy with iPSC-derived cells.
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Tratamiento Basado en Trasplante de Células y Tejidos , Células Madre Pluripotentes Inducidas , Enfermedades Pulmonares Intersticiales , Humanos , Células Madre Pluripotentes Inducidas/citología , Enfermedades Pulmonares Intersticiales/terapia , Enfermedades Pulmonares Intersticiales/patología , Tratamiento Basado en Trasplante de Células y Tejidos/métodos , Animales , Fibrosis Pulmonar Idiopática/terapia , Fibrosis Pulmonar Idiopática/patologíaRESUMEN
Lung adenocarcinoma (LUAD), a type of non-small cell lung cancer (NSCLC), originates from not only bronchial epithelial cells but also alveolar type 2 (AT2) cells, which could differentiate into AT2-like cells. AT2-like cells function as cancer stem cells (CSCs) of LUAD tumorigenesis to give rise to adenocarcinoma. However, the mechanism underlying AT2 cell differentiation into AT2-like cells in LUAD remains unknown. We analyze genes differentially expressed and genes with significantly different survival curves in LUAD, and the combination of these two analyses yields 147 differential genes, in which 14 differentially expressed genes were enriched in cell cycle pathway. We next analyze the protein levels of these genes in LUAD and find that Cyclin-A2 (CCNA2) is closely associated with LUAD tumorigenesis. Unexpectedly, high CCNA2 expression in LUAD is restrictedly associated with smoking and independent of other driver mutations. Single-cell sequencing analyses reveal that CCNA2 is predominantly involved in AT2-like cell differentiation, while inhibition of CCNA2 significantly reverses smoking-induced AT2-like cell differentiation. Mechanistically, CCNA2 binding to CDK2 phosphorylates the AXIN1 complex, which in turn induces ubiquitination-dependent degradation of ß-catenin and inhibits the WNT signaling pathway, thereby failing AT2 cell maintenance. These results uncover smoking-induced CCNA2 overexpression and subsequent WNT/ß-catenin signaling inactivation as a hitherto uncharacterized mechanism controlling AT2 cell differentiation and LUAD tumorigenesis.
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Adenocarcinoma del Pulmón , Carcinogénesis , Diferenciación Celular , Ciclina A2 , Neoplasias Pulmonares , Fumar , Animales , Femenino , Humanos , Masculino , Ratones , Adenocarcinoma del Pulmón/genética , Adenocarcinoma del Pulmón/patología , Adenocarcinoma del Pulmón/metabolismo , Células Epiteliales Alveolares/metabolismo , Células Epiteliales Alveolares/patología , beta Catenina/metabolismo , beta Catenina/genética , Carcinogénesis/genética , Línea Celular Tumoral , Ciclina A2/genética , Ciclina A2/metabolismo , Quinasa 2 Dependiente de la Ciclina/genética , Quinasa 2 Dependiente de la Ciclina/metabolismo , Regulación Neoplásica de la Expresión Génica , Neoplasias Pulmonares/patología , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/metabolismo , Células Madre Neoplásicas/patología , Células Madre Neoplásicas/metabolismo , Fumar/efectos adversos , Vía de Señalización Wnt/genética , RatasRESUMEN
Glycine-12 mutations in the GTPase KRAS (KRASG12) are an initiating event for development of lung adenocarcinoma (LUAD). KRASG12 mutations promote cell-intrinsic rewiring of alveolar type-II progenitor (AT2) cells, but to what extent such changes interplay with lung homeostasis and cell fate pathways is unclear. Here, we generated single-cell RNA-seq (scRNA-seq) profiles from AT2-mesenchyme organoid co-cultures, mice, and stage-IA LUAD patients, identifying conserved regulators of AT2 transcriptional dynamics and defining the impact of KRASG12D mutation with temporal resolution. In AT2WT organoids, we found a transient injury/plasticity state preceding AT2 self-renewal and AT1 differentiation. Early-stage AT2KRAS cells exhibited perturbed gene expression dynamics, most notably retention of the injury/plasticity state. The injury state in AT2KRAS cells of patients, mice, and organoids was distinguishable from AT2WT states via altered receptor expression, including co-expression of ITGA3 and SRC. The combination of clinically relevant KRASG12D and SRC inhibitors impaired AT2KRAS organoid growth. Together, our data show that an injury/plasticity state essential for lung repair is co-opted during AT2 self-renewal and LUAD initiation, suggesting that early-stage LUAD may be susceptible to interventions that target specifically the oncogenic nature of this cell state.
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Neoplasias Pulmonares , Organoides , Proteínas Proto-Oncogénicas p21(ras) , Animales , Humanos , Ratones , Adenocarcinoma del Pulmón/genética , Adenocarcinoma del Pulmón/patología , Adenocarcinoma del Pulmón/metabolismo , Diferenciación Celular , Regulación Neoplásica de la Expresión Génica , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/patología , Neoplasias Pulmonares/metabolismo , Mutación , Organoides/metabolismo , Organoides/patología , Proteínas Proto-Oncogénicas p21(ras)/genética , Proteínas Proto-Oncogénicas p21(ras)/metabolismo , Familia-src Quinasas/metabolismo , Familia-src Quinasas/genéticaRESUMEN
BACKGROUND: We have documented profound release of nitric oxide (NO) and endothelium-derived hyperpolarization factor (EDHF) by angiotensin II (ANGII) receptor 1 (AT1) blocker (ARB) losartan and its unique metabolite EXP3179, a pleiotropic effect that may help rationalize the protective properties of ARBs. Since blood pressure (BP) lowering by ARBs likely require an ANGII-dependent switch from AT1 to ANGII receptor 2 (AT2) signaling, a receptor known to stimulate endothelial NO release, we investigated the contribution of AT1 and AT2 to losartan and EXP3179's endothelial function-activating properties. EXPERIMENTAL APPROACH: Two AT1 ligands were used in an attempt to block the AT1-dependent endothelium-enhancing effects of EXP3179. AT2-null mice were used to evaluate the acute ex vivo and chronic in vivo effects of EXP3179 (20µM) and losartan (0.6 g/l), respectively, on endothelial function, BP and aortic stiffness. KEY RESULTS: Ex vivo blockade of AT1 receptors did not attenuate EXP3179's effects on NO and EDHF-dependent endothelial function activation. We observed significant reductions in PE-induced contractility with EXP3179 in both WT and AT2 knockout (KO) aortic rings. In vivo, a 1-month chronic treatment with losartan did not affect pulse wave velocity (PWV) but decreased PE-induced contraction by 74.9 % in WT (p < 0.0001) and 47.3 % in AT2 KO (p < 0.05). Presence of AT2 was critical to losartan's BP lowering activity. CONCLUSION: In contrast to BP lowering, the endothelial function-enhancing effects of losartan and EXP3179 are mostly independent of the classic ANGII/AT1/AT2 pathway, which sheds light on ARB pleiotropism.
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Presión Sanguínea , Endotelio Vascular , Losartán , Ratones Noqueados , Receptor de Angiotensina Tipo 2 , Animales , Losartán/farmacología , Presión Sanguínea/efectos de los fármacos , Endotelio Vascular/efectos de los fármacos , Endotelio Vascular/metabolismo , Ratones , Receptor de Angiotensina Tipo 2/metabolismo , Receptor de Angiotensina Tipo 2/genética , Masculino , Óxido Nítrico/metabolismo , Receptor de Angiotensina Tipo 1/metabolismo , Receptor de Angiotensina Tipo 1/genética , Imidazoles/farmacología , Ratones Endogámicos C57BL , Bloqueadores del Receptor Tipo 1 de Angiotensina II/farmacología , Rigidez Vascular/efectos de los fármacos , Sulfonamidas , TiofenosRESUMEN
BACKGROUND: Wnt/ß-catenin signaling is critical for lung development and AT2 stem cell maintenance in adults, but excessive pathway activation has been associated with pulmonary fibrosis, both in animal models and human diseases such as idiopathic pulmonary fibrosis (IPF). IPF is a detrimental interstitial lung disease, and although two approved drugs limit functional decline, transplantation is the only treatment that extends survival, highlighting the need for regenerative therapies. METHODS: Using our antibody-based platform of Wnt/ß-catenin modulators, we investigated the ability of a pathway antagonist and pathway activators to reduce pulmonary fibrosis in the acute bleomycin model, and we tested the ability of a WNT mimetic to affect alveolar organoid cultures. RESULTS: A WNT mimetic agonist with broad FZD-binding specificity (FZD1,2,5,7,8) potently expanded alveolar organoids. Upon therapeutic dosing, a broad FZD-binding specific Wnt mimetic decreased pulmonary inflammation and fibrosis and increased lung function in the bleomycin model, and it impacted multiple lung cell types in vivo. CONCLUSIONS: Our results highlight the unexpected capacity of a WNT mimetic to effect tissue repair after lung damage and support the continued development of Wnt/ß-catenin pathway modulation for the treatment of pulmonary fibrosis.
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Fibrosis Pulmonar Idiopática , beta Catenina , Adulto , Animales , Humanos , beta Catenina/metabolismo , Pulmón/metabolismo , Fibrosis Pulmonar Idiopática/inducido químicamente , Fibrosis Pulmonar Idiopática/tratamiento farmacológico , Fibrosis Pulmonar Idiopática/metabolismo , Vía de Señalización Wnt , Bleomicina/toxicidadRESUMEN
Alveolar type 2 (AT2) cells are stem cells of the alveolar epithelia. Previous genetic lineage tracing studies reported multiple cellular origins for AT2 cells after injury. However, conventional lineage tracing based on Cre-loxP has the limitation of non-specific labeling. Here, we introduced a dual recombinase-mediated intersectional genetic lineage tracing approach, enabling precise investigation of AT2 cellular origins during lung homeostasis, injury, and repair. We found AT1 cells, being terminally differentiated, did not contribute to AT2 cells after lung injury and repair. Distinctive yet simultaneous labeling of club cells, bronchioalveolar stem cells (BASCs), and existing AT2 cells revealed the exact contribution of each to AT2 cells post-injury. Mechanistically, Notch signaling inhibition promotes BASCs but impairs club cells' ability to generate AT2 cells during lung repair. This intersectional genetic lineage tracing strategy with enhanced precision allowed us to elucidate the physiological role of various epithelial cell types in alveolar regeneration following injury.
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Células Epiteliales Alveolares , Pulmón , Células Madre , Animales , Ratones , Células Epiteliales Alveolares/metabolismo , Células Epiteliales Alveolares/citología , Diferenciación Celular , Linaje de la Célula , Pulmón/citología , Pulmón/metabolismo , Pulmón/fisiología , Lesión Pulmonar/patología , Ratones Endogámicos C57BL , Alveolos Pulmonares/citología , Alveolos Pulmonares/metabolismo , Receptores Notch/metabolismo , Regeneración , Transducción de Señal , Células Madre/metabolismo , Células Madre/citologíaRESUMEN
Alveoli serve as the functional units of the lungs, responsible for the critical task of blood-gas exchange. Comprising type I (AT1) and type II (AT2) cells, the alveolar epithelium is continuously subject to external aggressors like pathogens and airborne particles. As such, preserving lung function requires both the homeostatic renewal and reparative regeneration of this epithelial layer. Dysfunctions in these processes contribute to various lung diseases. Recent research has pinpointed specific cell subgroups that act as potential stem or progenitor cells for the alveolar epithelium during both homeostasis and regeneration. Additionally, endothelial cells, fibroblasts, and immune cells synergistically establish a nurturing microenvironment-or "niche"-that modulates these epithelial stem cells. This review aims to consolidate the latest findings on the identities of these stem cells and the components of their niche, as well as the molecular mechanisms that govern them. Additionally, this article highlights diseases that arise due to perturbations in stem cell-niche interactions. We also discuss recent technical innovations that have catalyzed these discoveries. Specifically, this review underscores the heterogeneity, plasticity, and dynamic regulation of these stem cell-niche systems. It is our aspiration that a deeper understanding of the fundamental cellular and molecular mechanisms underlying alveolar homeostasis and regeneration will open avenues for identifying novel therapeutic targets for conditions such as chronic obstructive pulmonary disease (COPD), fibrosis, coronavirus disease 2019 (COVID-19), and lung cancer.
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The renin-angiotensin system (RAS) plays a key role in blood pressure regulation. The RAS is a complex interconnected system composed of two axes with opposite effects. The pressor arm, represented by angiotensin (Ang) II and the AT1 receptor (AT1R), mediates the vasoconstrictor, proliferative, hypertensive, oxidative, and pro-inflammatory effects of the RAS, while the depressor/protective arm, represented by Ang-(1-7), its Mas receptor (MasR) and the AT2 receptor (AT2R), opposes the actions elicited by the pressor arm. The AT1R, AT2R, and MasR belong to the G-protein-coupled receptor (GPCR) family. GPCRs operate not only as monomers, but they can also function in dimeric (homo and hetero) or higher-order oligomeric states. Due to the interaction with other receptors, GPCR properties may change: receptor affinity, trafficking, signaling, and its biological function may be altered. Thus, heteromerization provides a newly recognized means of modulation of receptor function, as well as crosstalk between GPCRs. This review is focused on angiotensin receptors, and how their properties are influenced by crosstalk with other receptors, adding more complexity to an already complex system and potentially opening up new therapeutic approaches.
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Receptores Acoplados a Proteínas G , Sistema Renina-Angiotensina , Humanos , Sistema Renina-Angiotensina/fisiología , Animales , Receptores Acoplados a Proteínas G/metabolismo , Transducción de Señal/fisiología , Receptor Cross-Talk/fisiología , Receptores de Angiotensina/metabolismo , Receptor de Angiotensina Tipo 1/metabolismo , Presión Sanguínea/fisiología , Receptor de Angiotensina Tipo 2/metabolismoRESUMEN
Lung aging triggers the onset of various chronic lung diseases, with alveolar repair being a key focus for alleviating pulmonary conditions. The regeneration of epithelial structures, particularly the differentiation from type II alveolar epithelial (AT2) cells to type I alveolar epithelial (AT1) cells, serves as a prominent indicator of alveolar repair. Nonetheless, the precise role of aging in impeding alveolar regeneration and its underlying mechanism remain to be fully elucidated. Our study employed histological methods to examine lung aging effects on structural integrity and pathology. Lung aging led to alveolar collapse, disrupted epithelial structures, and inflammation. Additionally, a relative quantification analysis revealed age-related decline in AT1 and AT2 cells, along with reduced proliferation and differentiation capacities of AT2 cells. To elucidate the mechanisms underlying AT2 cell functional decline, we employed transcriptomic techniques and revealed a correlation between inflammatory factors and genes regulating proliferation and differentiation. Furthermore, a D-galactose-induced senescence model in A549 cells corroborated our omics experiments and confirmed inflammation-induced cell cycle arrest and a >30% reduction in proliferation/differentiation. Physiological aging-induced chronic inflammation impairs AT2 cell functions, hindering tissue repair and promoting lung disease progression. This study offers novel insights into chronic inflammation's impact on stem cell-mediated alveolar regeneration.
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Células Epiteliales Alveolares , Pulmón , Humanos , Células Epiteliales Alveolares/metabolismo , Células Cultivadas , Pulmón/metabolismo , Diferenciación Celular/fisiología , Inflamación/metabolismoRESUMEN
A large portion of the heterogeneity in coronavirus disease 2019 (COVID-19) susceptibility and severity of illness (SOI) remains poorly understood. Recent evidence suggests that SARS-CoV-2 infection-associated damage to alveolar epithelial type 2 cells (AT2s) in the distal lung may directly contribute to disease severity and poor prognosis in COVID-19 patients. Our in vitro modeling of SARS-CoV-2 infection in induced pluripotent stem cell (iPSC)-derived AT2s from 10 different individuals showed interindividual variability in infection susceptibility and the postinfection cellular viral load. To understand the underlying mechanism of the AT2's capacity to regulate SARS-CoV-2 infection and cellular viral load, a genome-wide differential gene expression analysis between the mock and SARS-CoV-2 infection-challenged AT2s was performed. The 1393 genes, which were significantly (one-way ANOVA FDR-corrected p ≤ 0.05; FC abs ≥ 2.0) differentially expressed (DE), suggest significant upregulation of viral infection-related cellular innate immune response pathways (p-value ≤ 0.05; activation z-score ≥ 3.5), and significant downregulation of the cholesterol- and xenobiotic-related metabolic pathways (p-value ≤ 0.05; activation z-score ≤ -3.5). Whilst the effect of post-SARS-CoV-2 infection response on the infection susceptibility and postinfection viral load in AT2s is not clear, interestingly, pre-infection (mock-challenged) expression of 238 DE genes showed a high correlation with the postinfection SARS-CoV-2 viral load (FDR-corrected p-value ≤ 0.05 and r2-absolute ≥ 0.57). The 85 genes whose expression was negatively correlated with the viral load showed significant enrichment in viral recognition and cytokine-mediated innate immune GO biological processes (p-value range: 4.65 × 10-10 to 2.24 × 10-6). The 153 genes whose expression was positively correlated with the viral load showed significant enrichment in cholesterol homeostasis, extracellular matrix, and MAPK/ERK pathway-related GO biological processes (p-value range: 5.06 × 10-5 to 6.53 × 10-4). Overall, our results strongly suggest that AT2s' pre-infection innate immunity and metabolic state affect their susceptibility to SARS-CoV-2 infection and viral load.
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COVID-19 , Células Madre Pluripotentes Inducidas , Humanos , SARS-CoV-2 , Carga Viral , Inmunidad Innata , ColesterolRESUMEN
O sistema renina angiotensina (SRA) é de grande importância para o equilíbrio hídrico e regulação da pressão arterial do organismo, além de estar associado ao estimulo de vias próinflamatórias. Seu principal peptídeo é a angiotensina II, que interage principalmente com os receptores do tipo 1 (AT1) e do tipo 2 (AT2). Foi encontrado interrelação entre as doenças cardiovasculares e a periodontite. Este estudo teve como objetivo avaliar os aspectos moleculares em camundongos submetidos a um modelo experimental de periodontite, observando a influência dos receptores de Ang II tipo 1 (AT1(-)) e Ang II tipo 2 (AT2(-)) na periodontite. Métodos: A periodontite experimental foi induzida colocando-se uma ligadura com fio de nylon 5.0 ao redor do segundo molar superior esquerdo de camundongos knockoutAT1(-), AT2(-) e selvagem (WT), subdivididos 2 grupos para cada linhagem: sem ligadura e ligadura, totalizando seis grupos: três controles e três experimentais. Após 15 dias da indução da doença os animais foram submetidos à eutanásia. Com o intuito de avaliar se as variações genéticas teriam influência sobre a periodontite foram realizadas as análises de citocinas, peptídeos e enzimas foram analisados a partir de tecidos gengivais por ELISA e RT-PCR. Resultados: Os animais WT e AT2(-) apresentaram resultados semelhantes em relação às citocinas IL-1ß, IL-6, TNF-α, com aumento dos níveis em relação aos saudáveis (p < 0,001). Houve diferenças significativas em IL-ß entre os grupo AT1(-)-L e WT-L (p < 0,05), e em IL-6 e TNF-α os grupos AT1(-)-L apresentaram diferenças significativas (p < 0,001) tanto quando comparado aos grupo WT-L quanto aos grupos AT2(-)-L. Os níveis de IL-10 foram maiores em WT-L (p < 0,01), enquanto os grupos AT2(-) e AT1(-) não apresentaram alterações significativas em relação a essa citocina. Houve diferenças significativas em Angiotensina II entre os grupos AT2(-)-NL e AT2(-)-L (p < 0,01); e em Angiotensina 1-7 entre os grupos AT1(-)-L e AT2(-)-L (p < 0,05). Para TLR2 houve diferenças entre os grupos WT-NL/WT-L (p < 0,05); AT1(-)-NL/AT1(-)-L (p < 0,01) e AT2(-)-NL/AT2(-) - L (p < 0,01). Para o receptor MAS houve diferenças entre os grupos WT-NL/WT-L (p < 0,001) e AT2(-)-NL/AT2(-)-L (p < 0,001), e também em relação ao grupo WT-L/AT1(-)-L (p < 0,001) e AT1(-)-L/AT2(-)-L (p < 0,001). Para a expressão dos peptídeos ECA e ECA2, houve diferença estatística apenas para ECA entre os tipos de grupos WT-NL/WT-L (p < 0,001). Conclusão: Os animais do grupo AT1(-) apresentaram menor inflamação que as demais linhagens doentes, assim como uma menor expressão do receptor Mas e Ang 1-7. Além disso os animais dos grupos WT e AT2(-) demonstraram resultados próximos em diversas análises, evidenciando que o bloqueio do receptor AT1, sobre os efeitos moleculares, é mais positiva (AU).
The renin angiotensin system (RAS) is of great importance for water balance and regulation of blood pressure in the body, in addition to being associated with the stimulation of proinflammatory pathways. Its main peptide is angiotensin II, which interacts mainly with type 1 (AT1) and type 2 (AT2) receptors. An interrelationship was found between cardiovascular diseases and periodontitis. This study aimed to evaluate the molecular aspects in mice subjected to an experimental model of periodontal disease, observing the influence of Ang II type 1 (AT1(-)) and Ang II type 2 (AT2(-)) receptors on periodontitis. Methods: Experimental periodontitis was induced by placing a ligature with 5.0 nylon thread around the upper left second molar of AT1(-), AT2(-) and wild-type (WT) knockout mice, subdivided into 2 groups for each strain: without ligation and ligation, totaling six groups: three controls and three experimental. After 15 days of disease induction, the animals were euthanized. In order to evaluate whether genetic variations would have an influence on periodontal disease, analyzes of cytokines were carried out, peptides and enzymes were analyzed from gingival tissues by ELISA and RT-PCR. Results: WT and AT2(-) animals showed similar results in relation to the cytokines IL-1ß, IL-6, TNF-α, with increased levels compared to healthy ones (p < 0.001). There were significant differences in IL-ß between the AT1(-)-L and WT-L groups (p < 0.05), and in IL-6 and TNF-α the AT1(-)-L groups showed significant differences (p < 0.001) both when compared to the WT-L and AT2(-)-L groups. IL-10 levels were higher in WT-L (p < 0.01), while the AT2(-) and AT1(-) groups did not show significant changes in relation to this cytokine. There were significant differences in Angiotensin II between the AT2(-)-NL and AT2(-)-L groups (p < 0.01); and in Angiotensin 1-7 between the AT1(-)-L and AT2(-)-L groups (p < 0.05). For TLR2 there were differences between the WT-NL/WT-L groups (p < 0.05); AT1(-)-NL/AT1(-)-L (p < 0.01) and AT2(-)-NL/AT2(-)-L (p < 0.01). For the MAS receptor there were differences between the WT-NL/WT-L (p < 0.001) and AT2(-)-NL/AT2(- )-L (p < 0.001) groups, and also in relation to the WT-L group /AT1(-)-L (p < 0.001) and AT1(-)-L/AT2(-)-L (p < 0.001). For the expression of ACE and ACE2 peptides, there was a statistical difference only for ACE between the types of WT-NL/WT-L groups (p < 0.001). Conclusion: The animals in the AT1(-) group showed less inflammation than the other diseased lines, as well as a lower expression of the Mas and Ang 1-7 receptor. Furthermore, animals from the WT and AT2(-) groups demonstrated similar results in several analyses, showing that the blockade of the AT1 receptor, on molecular effects, is more positive (AU).
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Animales , Ratones , Enfermedades Periodontales/patología , Angiotensinas , Receptor de Angiotensina Tipo 1/efectos de los fármacos , Receptor de Angiotensina Tipo 2/efectos de los fármacos , Técnicas In Vitro/métodos , Epidemiología Descriptiva , Análisis de Varianza , Estadísticas no ParamétricasRESUMEN
The angiotensin AT2 receptor (AT2R), an important member of the "protective arm" of the renin-angiotensin system (RAS), has been recently defined as a therapeutic target in different pathological conditions. The AT2R activates complex signalling pathways linked to cellular proliferation, differentiation, anti-inflammation, antifibrosis, and induction or inhibition of apoptosis. The anti-inflammatory effect of AT2R activation is commonly associated with reduced fibrosis in different models. Current discoveries demonstrated a direct impact of AT2Rs on the regulation of cytokines, transforming growth factor beta1 (TGF-beta1), matrix metalloproteases (MMPs), and synthesis of the extracellular matrix components. This review article summarizes current knowledge on the AT2R in regard to immunity, inflammation and fibrosis in the heart and blood vessels. In particular, the differential influence of the AT2R on cardiovascular remodeling in preclinical models of myocardial infarction, heart failure and aneurysm formation are discussed. Overall, these studies demonstrate that AT2R stimulation represents a promising therapeutic approach to counteract myocardial and aortic damage in cardiovascular diseases.
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Infarto del Miocardio , Sistema Renina-Angiotensina , Humanos , Infarto del Miocardio/metabolismo , Inflamación/tratamiento farmacológico , Inflamación/metabolismo , Receptor de Angiotensina Tipo 2/metabolismo , Miocardio/metabolismo , Angiotensinas/metabolismo , Fibrosis , Receptor de Angiotensina Tipo 1/metabolismoRESUMEN
The renin-angiotensin system is the most important peptide hormone system in the regulation of cardiovascular homeostasis. Its classical arm consists of the enzymes, renin, and angiotensin-converting enzyme, generating angiotensin II from angiotensinogen, which activates its AT1 receptor, thereby increasing blood pressure, retaining salt and water, and inducing cardiovascular hypertrophy and fibrosis. However, angiotensin II can also activate a second receptor, the AT2 receptor. Moreover, the removal of the C-terminal phenylalanine from angiotensin II by ACE2 (angiotensin-converting enzyme 2) yields angiotensin-(1-7), and this peptide interacts with its receptor Mas. When the aminoterminal Asp of angiotensin-(1-7) is decarboxylated, alamandine is generated, which activates the Mas-related G-protein-coupled receptor D, MrgD (Mas-related G-protein-coupled receptor type D). Since Mas, MrgD, and the AT2 receptor have opposing effects to the classical AT1 receptor, they and the enzymes and peptides activating them are called the alternative or protective arm of the renin-angiotensin system. This review will cover the historical aspects and the current standing of this recent addition to the biology of the renin-angiotensin system.
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Angiotensina II , Sistema Renina-Angiotensina , Angiotensina I/metabolismo , Fragmentos de Péptidos/metabolismo , Péptidos , Peptidil-Dipeptidasa A/metabolismo , Receptores Acoplados a Proteínas G/metabolismo , Renina , Sistema Renina-Angiotensina/fisiología , HumanosRESUMEN
This study investigated the effects of calcitriol on polyinosinic-polycytidylic acid (poly(I:C))-induced acute lung injury (ALI) and its association with Toll-like receptor 3 (TLR3) and renin-angiotensin system (RAS) signal pathways in obese mice. Normal mice were fed a high-fat diet to induce obesity. Obese mice were divided into four groups: SS group, intratracheally instilled with saline and intravenous (IV) saline injection via tail vein; SD group, instilled with saline and IV calcitriol injection; PS group, instilled with poly(I:C) and IV saline injection; and PD group, instilled with poly(I:C) and IV calcitriol injection. All mice were sacrificed 12 or 24 h after poly(I:C) stimulation. The results showed that poly(I:C) instillation led to increased production of systemic inflammatory cytokines. In the lungs, the population of macrophages decreased, while more neutrophils were recruited. TLR3-associated genes including IRF3, nuclear factor-κB, interferon-ß and phosphorylated IRF3 expression levels, were upregulated. The RAS-associated AT1R and ACE2 protein levels increased, whereas AT2R, Ang(1-7), and MasR levels decreased. Also, reduced tight junction (TJ) proteins and elevated lipid peroxide levels were observed 24 h after poly(I:C) stimulation. Compared to the PS group, the PD group exhibited reduced systemic and lung inflammatory cytokine levels, increased macrophage while decreased neutrophil percentages, downregulated TLR3-associated genes and phosphorylated IRF3, and polarized toward the RAS-AT2R/Ang(1-7)/MasR pathway in the lungs. Higher lung TJ levels and lower injury scores were also noted. These findings suggest that calcitriol treatment after poly(I:C) instillation alleviated ALI in obese mice possibly by downregulating TLR3 expression and tending toward the RAS-associated anti-inflammatory pathway.
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Lesión Pulmonar Aguda , Sistema Renina-Angiotensina , Ratones , Animales , Receptor Toll-Like 3/metabolismo , Calcitriol , Ratones Obesos , Poli I-C/metabolismo , Transducción de Señal , Lesión Pulmonar Aguda/metabolismo , Citocinas/metabolismoRESUMEN
Two series of N-(heteroaryl)thiophene sulfonamides, encompassing either a methylene imidazole group or a tert-butylimidazolylacetyl group in the meta position of the benzene ring, have been synthesized. An AT2R selective ligand with a Ki of 42 nM was identified in the first series and in the second series, six AT2R selective ligands with significantly improved binding affinities and Ki values of <5 nM were discovered. The binding modes to AT2R were explored by docking calculations combined with molecular dynamics simulations. Although some of the high affinity ligands exhibited fair stability in human liver microsomes, comparable to that observed with C21 undergoing clinical trials, most ligands displayed a very low metabolic stability with t½ of less than 10 min in human liver microsomes. The most promising ligand, with an AT2R Ki value of 4.9 nM and with intermediate stability in human hepatocytes (t½ = 77 min) caused a concentration-dependent vasorelaxation of pre-contracted mouse aorta.