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Movimiento Celular , Neutrófilos , Neutrófilos/inmunología , Humanos , Animales , Movimiento Celular/inmunología , RatonesRESUMEN
Neutrophil recruitment to sites of infection and inflammation is an essential process in the early innate immune response. Upon activation, a subset of neutrophils rapidly assembles the multiprotein complex known as the NLRP3 inflammasome. The NLRP3 inflammasome forms at the microtubule organizing center, which promotes the formation of interleukin (IL)-1ß and IL-18, essential cytokines in the immune response. We recently showed that mice deficient in NLRP3 (NLRP3-/-) have reduced neutrophil recruitment to the peritoneum in a model of thioglycolate-induced peritonitis. Here, we tested the hypothesis that this diminished recruitment could be, in part, the result of defects in neutrophil chemotaxis. We find that NLRP3-/- neutrophils show loss of cell polarization, as well as reduced directionality and velocity of migration toward increasing concentrations of leukotriene B4 (LTB4) in a chemotaxis assay in vitro, which was confirmed through intravital microscopy of neutrophil migration toward a laser-induced burn injury of the liver. Furthermore, pharmacologically blocking NLRP3 inflammasome assembly with MCC950 in vitro reduced directionality but preserved nondirectional movement, indicating that inflammasome assembly is specifically required for polarization and directional chemotaxis, but not cell motility per se. In support of this, pharmacological breakdown of the microtubule cytoskeleton via nocodazole treatment induced cell polarization and restored nondirectional cell migration in NLRP3-deficient neutrophils in the LTB4 gradient. Therefore, NLRP3 inflammasome assembly is required for establishment of cell polarity to guide the directional chemotactic migration of neutrophils.
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Quimiotaxis , Leucotrieno B4 , Proteína con Dominio Pirina 3 de la Familia NLR , Animales , Ratones , Inflamasomas , Leucotrieno B4/metabolismo , Neutrófilos , Proteína con Dominio Pirina 3 de la Familia NLR/metabolismoRESUMEN
Integral membrane proteins (IMPs) have crucial roles in many cellular processes. A novel intramembrane chaperone complex, recently elucidated by Chitwood and Hedge, provides mechanistic insight of IMP biogenesis and folding, illustrating how IMPs with multiple transmembrane domains (TMDs) are assembled within the endoplasmic reticulum (ER) membrane.
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Retículo Endoplásmico , Proteínas de la Membrana , Retículo Endoplásmico/metabolismo , Proteínas de la Membrana/metabolismo , Biosíntesis de Proteínas , Dominios Proteicos , Pliegue de ProteínaRESUMEN
Thymic stromal lymphopoietin (TSLP) is an epithelial-derived pleiotropic cytokine that regulates T-helper 2 (Th2) immune responses in the lung and plays a major role in severe uncontrolled asthma. Emerging evidence suggests a role for endoplasmic reticulum (ER) stress in the pathogenesis of asthma. In this study, we determined if ER stress and the unfolded protein response (UPR) signaling are involved in TSLP induction in the airway epithelium. For this, we treated human bronchial epithelial basal cells and differentiated primary bronchial epithelial cells with ER stress inducers and the TSLP mRNA and protein expression was determined. A series of siRNA gene knockdown experiments were conducted to determine the ER stress-induced TSLP signaling pathways. cDNA collected from asthmatic bronchial biopsies was used to determine the gene correlation between ER stress and TSLP. Our results show that ER stress signaling induces TSLP mRNA expression via the PERK-C/EBP homologous protein (CHOP) signaling pathway. AP-1 transcription factor is important in regulating this ER stress-induced TSLP mRNA induction, though ER stress alone cannot induce TSLP protein production. However, ER stress significantly enhances TLR3-induced TSLP protein secretion in the airway epithelium. TSLP and ER stress (PERK) mRNA expression positively correlates in bronchial biopsies from participants with asthma, particularly in neutrophilic asthma. In conclusion, these results suggest that ER stress primes TSLP that is then enhanced further upon TLR3 activation, which may induce severe asthma exacerbations. Targeting ER stress using pharmacological interventions may provide novel therapeutics for severe uncontrolled asthma.NEW & NOTEWORTHY TSLP is an epithelial-derived cytokine and a key regulator in the pathogenesis of severe uncontrolled asthma. We demonstrate a novel mechanism by which endoplasmic reticulum stress signaling upregulates airway epithelial TSLP mRNA expression via the PERK-CHOP signaling pathway and enhances TLR3-mediated TSLP protein secretion.
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Asma , Citocinas , Estrés del Retículo Endoplásmico , Células Epiteliales , Linfopoyetina del Estroma Tímico , Receptor Toll-Like 3 , Respuesta de Proteína Desplegada , Humanos , Citocinas/metabolismo , Receptor Toll-Like 3/metabolismo , Receptor Toll-Like 3/genética , Asma/metabolismo , Asma/patología , Asma/genética , Células Epiteliales/metabolismo , Células Epiteliales/patología , Factor de Transcripción CHOP/metabolismo , Factor de Transcripción CHOP/genética , Transducción de Señal , Mucosa Respiratoria/metabolismo , Mucosa Respiratoria/patología , Bronquios/metabolismo , Bronquios/patología , eIF-2 Quinasa/metabolismo , eIF-2 Quinasa/genética , Células Cultivadas , Femenino , ARN Mensajero/genética , ARN Mensajero/metabolismoRESUMEN
BACKGROUND: Non-neuronal cholinergic system (NNCS) contributes to various inflammatory airway diseases. However, the role of NNCS in severe asthma (SA) remains largely unexplored. OBJECTIVE: To explore airway NNCS in SA. METHODS: In this prospective cohort study based on the Australasian Severe Asthma Network in a real-world setting, patients with SA (n = 52) and non-SA (n = 104) underwent clinical assessment and sputum induction. The messenger RNA (mRNA) levels of NNCS components and proinflammatory cytokines in the sputum were detected using real-time quantitative polymerase chain reaction, and the concentrations of acetylcholine (Ach)-related metabolites were evaluated using liquid chromatography coupled with tandem mass spectrometry. Asthma exacerbations were prospectively investigated during the next 12 months. The association between NNCS and future asthma exacerbations was also analyzed. RESULTS: Patients with SA were less controlled and had worse airway obstruction, a lower bronchodilator response, higher doses of inhaled corticosteroids, and more add-on treatments. The sputum mRNA levels of NNCS components, such as muscarinic receptors M1R-M5R, OCT3, VACHT, and ACHE; proinflammatory cytokines; and Ach concentration in the SA group were significantly higher than those in the non-SA group. Furthermore, most NNCS components positively correlated with non-type (T) 2 inflammatory profiles, such as sputum neutrophils, IL8, and IL1B. In addition, the mRNA levels of sputum M2R, M3R, M4R, M5R, and VACHT were independently associated with an increased risk of moderate-to-severe asthma exacerbations. CONCLUSION: This study indicated that the NNCS was significantly activated in SA, leading to elevated Ach and was associated with clinical features, non-T2 inflammation, and future exacerbations of asthma, highlighting the potential role of the NNCS in the pathogenesis of SA. CLINICAL TRIAL REGISTRATION: ChiCTR-OOC-16009529 (http://www.chictr.org.cn).
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Asma , Citocinas , Sistema Colinérgico no Neuronal , Esputo , Adulto , Anciano , Femenino , Humanos , Masculino , Persona de Mediana Edad , Acetilcolina/metabolismo , Asma/inmunología , Asma/metabolismo , Citocinas/metabolismo , Progresión de la Enfermedad , Inflamación/metabolismo , Sistema Colinérgico no Neuronal/inmunología , Estudios Prospectivos , Índice de Severidad de la Enfermedad , Esputo/metabolismo , Esputo/inmunologíaRESUMEN
BACKGROUND: A better understanding of the association between chronic kidney disease (CKD) and glaucoma is required to optimize clinical outcomes. Therefore, this study aimed to investigate the association of chronic kidney disease (CKD) with new diagnoses of glaucoma over time from January 2009 to December 2019. METHOD: This retrospective propensity-matched cohort study utilizing Taiwanese electronic health records examined the incidence of newly diagnosed glaucoma in patients with and without chronic kidney disease (CKD). The exposure variable was the diagnosis of CKD, identified through diagnostic codes. The primary outcome was the incidence of new-onset glaucoma. Subgroup analyses on glaucoma risk included age, gender, comorbidities, glaucoma subtypes, and dialysis status. Statistical analyses included Kaplan-Meier analysis, Cox proportional hazards models, and Poisson regression models, with the associated hazard ratios and confidence intervals reported. RESULTS: Seven hundred twenty-three thousand two hundred sixteen patients with CKD (42.3% female; mean [SD] age at index, 66.3 [15.6] years) and 723,216 patients without CKD (42.3% female; mean [SD] age at index, 66.3 [15.7]) were recruited. We showed a significantly increased risk of glaucoma irrespective of subtypes in CKD patients compared to those without CKD (HR: 1.29 [CI: 1.26-1.32], p < 0.001). Kaplan-Meier curves revealed a significantly increased glaucoma risk in both the dialytic subtype and non-dialytic CKD patients when compared to their non-CKD counterparts (p < 0.001). We also showed that all genders (aHR 1.17 [CI: 1.13-1.21] for females vs. aHR 1.39 [CI:1.35-1.43] for males), all ages (< = 49: aHR 1.49 [CI: 1.37-1.62]; 50-59: aHR 1.48 [CI: 1.40-1.56]; 60-69: aHR 1.30 [CI: 1.25-1.6]; 70-79: aHR 1.21 [CI: 1.17-1.26]; > 80: aHR 1.29 [CI: 1.21-1.37]); all income brackets and all urbanization status were associated with significantly increased risk of glaucoma from among the CKD cohort when compared to their respective non-CKD cohort (p < 0.001). CONCLUSIONS: Our cohort study spanning 12 years showed an elevated glaucoma risk following a CKD diagnosis compared to a frequency-matched non-CKD cohort. Our findings have relevance for the clinical practice of at-risk CKD patients. TRIAL REGISTRATION: Due to the retrospective nature of the study, no registration was necessary.
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Glaucoma , Insuficiencia Renal Crónica , Humanos , Masculino , Femenino , Insuficiencia Renal Crónica/epidemiología , Persona de Mediana Edad , Taiwán/epidemiología , Estudios Retrospectivos , Anciano , Glaucoma/epidemiología , Incidencia , Estudios de Cohortes , Factores de Riesgo , Medición de Riesgo/métodos , Anciano de 80 o más Años , Modelos de Riesgos Proporcionales , Comorbilidad , AdultoRESUMEN
Background and Objectives: The risks of uveitis development among pediatric patients with Down syndrome (DS) remain unclear. Therefore, we aimed to determine the risk of uveitis following a diagnosis of DS. Materials and Methods: This multi-institutional retrospective cohort study utilized the TriNetX database to identify individuals aged 18 years and younger with and without a diagnosis of DS between 1 January 2000 and 31 December 2023. The non-DS cohort consisted of randomly selected control patients matched by selected variables. This included gender, age, ethnicity, and certain comorbidities. The main outcome is the incidence of new-onset uveitis. Statistical analysis of the uveitis risk was reported using hazard ratios (HRs) and 95% confidence intervals (CIs). Separate analyses of the uveitis risk among DS patients based on age groups and gender were also performed. Results: A total of 53,993 individuals with DS (46.83% female, 58.26% white, mean age at index 5.21 ± 5.76 years) and 53,993 non-DS individuals (45.56% female, 58.28% white, mean age at index 5.21 ± 5.76 years) were recruited from the TriNetX database. Our analysis also showed no overall increased risk of uveitis among DS patients (HR: 1.33 [CI: 0.89-1.99]) compared to the non-DS cohort across the 23-year study period. Subgroup analyses based on different age groups showed that those aged 0-1 year (HR: 1.36 [CI: 0.68-2.72]), 0-5 years (HR: 1.34 [CI: 0.75-2.39]), and 6-18 years (HR: 1.15 [CI: 0.67-1.96]) were found to have no association with uveitis risk compared to their respective non-DS comparators. There was also no increased risk of uveitis among females (HR: 1.49 [CI: 0.87-2.56]) or males (HR: 0.82 [CI: 0.48-1.41]) with DS compared to their respective non-DS comparators. Conclusions: Our study found no overall increased risk of uveitis following a diagnosis of DS compared to a matched control population.
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Síndrome de Down , Uveítis , Humanos , Síndrome de Down/complicaciones , Masculino , Femenino , Uveítis/epidemiología , Uveítis/diagnóstico , Uveítis/etiología , Niño , Estudios Retrospectivos , Preescolar , Adolescente , Lactante , Bases de Datos Factuales , Incidencia , Estudios de Cohortes , Factores de Riesgo , Medición de Riesgo/métodos , Medición de Riesgo/estadística & datos numéricosRESUMEN
Respiratory virus infections initiate and transmit from the upper respiratory tract (URT). Coronaviruses, including OC43, are a major cause of respiratory infection and disease. Failure to mount an effective antiviral immune response in the nasal mucosa increases the risk of severe disease and person-to-person transmission, highlighting the need for URT infection models to support the development of nasal treatments that improve coronavirus antiviral immunity. We aimed to determine if OC43 productively infected the mouse URT and would therefore be a suitable model to assess the efficacy and mechanism of action of nasal-targeting immune-modifying treatments. We administered OC43 via intranasal inoculation to wild-type Balb/c mice and assessed virus airway tropism (by comparing total respiratory tract vs. URT-only virus exposure) and characterized infection-induced immunity by quantifying specific antiviral cytokines and performing gene array assessment of immune genes. We then assessed the effect of immune-modulating therapies, including an immune-stimulating TLR2/6 agonist (INNA-X) and the immune-suppressing corticosteroid fluticasone propionate (FP). OC43 replicated in nasal respiratory epithelial cells, with peak viral RNA observed 2 days after infection. Prophylactic treatment with INNA-X accelerated expression of virus-induced IFN-λ and IFN-stimulated genes. In contrast, intranasal FP treatment increased nasal viral load by 2.4 fold and inhibited virus-induced IFN and IFN-stimulated gene expression. Prior INNA-X treatment reduced the immune-suppressive effect of FP. We demonstrate that the mouse nasal epithelium is permissive to OC43 infection and strengthen the evidence that TLR2 activation is a ß-coronavirus innate immune determinant and therapeutic target.
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Infecciones del Sistema Respiratorio , Receptor Toll-Like 2 , Humanos , Animales , Ratones , Infecciones del Sistema Respiratorio/tratamiento farmacológico , Citocinas/metabolismo , Mucosa Nasal/metabolismo , Interferón lambdaRESUMEN
Influenza A virus (IAV) infections are increased during pregnancy especially with asthma as a comorbidity, leading to asthma exacerbations, secondary bacterial infections, intensive care unit admissions, and mortality. We aimed to define the processes involved in increased susceptibility and severity of IAV infections during pregnancy, especially with asthma. We sensitized mice to house dust mite (HDM), induced pregnancy, and challenged with HDM to induce allergic airway disease (AAD). At midpregnancy, we induced IAV infection. We assessed viral titers, airway inflammation, lung antiviral responses, mucus hypersecretion, and airway hyperresponsiveness (AHR). During early IAV infection, pregnant mice with AAD had increased mRNA expression of the inflammatory markers Il13 and IL17 and reduced mRNA expression of the neutrophil chemoattractant marker Kc. These mice had increased mucous hyperplasia and increased AHR. miR155, miR574, miR223, and miR1187 were also reduced during early infection, as was mRNA expression of the antiviral ß-defensins, Bd1, Bd2, and Spd and IFNs, Ifnα, Ifnß, and Ifnλ. During late infection, Il17 was still increased as was eosinophil infiltration in the lungs. mRNA expression of Kc was reduced, as was neutrophil infiltration and mRNA expression of the antiviral markers Ifnß, Ifnλ, and Ifnγ and Ip10, Tlr3, Tlr9, Pkr, and Mx1. Mucous hyperplasia was still significantly increased as was AHR. Early phase IAV infection in pregnancy with asthma heightens underlying inflammatory asthmatic phenotype and reduces antiviral responses.NEW & NOTEWORTHY Influenza A virus (IAV) infection during pregnancy with asthma is a major health concern leading to increased morbidity for both mother and baby. Using murine models, we show that IAV infection in pregnancy with allergic airway disease is associated with impaired global antiviral and antimicrobial responses, increased lung inflammation, mucus hypersecretion, and airway hyperresponsiveness (AHR). Targeting specific ß-defensins or microRNAs (miRNAs) may prove useful in future treatments for IAV infection during pregnancy.
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Asma , Virus de la Influenza A , Gripe Humana , Trastornos Respiratorios , Hipersensibilidad Respiratoria , beta-Defensinas , Embarazo , Femenino , Ratones , Animales , Humanos , Citocinas/metabolismo , Hiperplasia/patología , Asma/patología , Pulmón/metabolismo , Hipersensibilidad Respiratoria/patología , Gripe Humana/patología , Antivirales/uso terapéutico , ARN Mensajero , Pyroglyphidae , Modelos Animales de EnfermedadRESUMEN
CRISPR/Cas9-based tissue-specific knockout techniques are essential for probing the functions of genes in embryonic development and disease using zebrafish. However, the lack of capacity to perform gene-specific rescue or live imaging in the tissue-specific knockout background has limited the utility of this approach. Here, we report a robust and flexible gateway system for tissue-specific gene inactivation in neutrophils. Using a transgenic fish line with neutrophil-restricted expression of Cas9 and ubiquitous expression of single guide (sg)RNAs targeting rac2, specific disruption of the rac2 gene in neutrophils is achieved. Transient expression of sgRNAs targeting rac2 or cdk2 in the neutrophil-restricted Cas9 line also results in significantly decreased cell motility. Re-expressing sgRNA-resistant rac2 or cdk2 genes restores neutrophil motility in the corresponding knockout background. Moreover, active Rac and force-bearing F-actins localize to both the cell front and the contracting tail during neutrophil interstitial migration in an oscillating fashion that is disrupted when rac2 is knocked out. Together, our work provides a potent tool that can be used to advance the utility of zebrafish in identifying and characterizing gene functions in a tissue-specific manner.
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Neutrófilos , Pez Cebra , Animales , Animales Modificados Genéticamente , Sistemas CRISPR-Cas/genética , Repeticiones Palindrómicas Cortas Agrupadas y Regularmente Espaciadas/genética , Neutrófilos/metabolismo , Pez Cebra/genética , Pez Cebra/metabolismo , Proteínas de Pez Cebra/genética , Proteínas de Pez Cebra/metabolismo , Proteínas de Unión al GTP rac/metabolismoRESUMEN
Reports on uveitis after COVID-19 have been limited. Our objective was to examine the risk of uveitis among COVID-19 patients. This was a retrospective cohort study based on the TriNetX platform. The exposure group was patients with positive laboratory test result for SARS-CoV-2 and the comparison group was those tested negative for COVID-19 throughout the study period. The endpoint is the new diagnoses of uveitis. This study composed of 2 105 424 patients diagnosed with COVID-19 (55.4% female; 62.5% white; mean age at index 40.7 years) and 2 105 424 patients (55.4% female; 62.4% white; mean age at index 40.7 years) who never had COVID-19. There was significantly increased risk of new diagnosis of uveitis since the first month after diagnosis of COVID-19 compared with matched controls (HR: 1.18, 95% CI: 1.03-1.34) up to 24 months (HR: 1.16, 95% CI: 1.09-1.22). Our findings strengthen those previously raised by case series with a larger and multicenter study. We found that uveitis was significantly associated with COVID-19 infection. Our findings reiterate the need for careful investigation as well as increased awareness from ophthalmologists in considering the possibility of COVID-19 in vulnerable patients with new presentation of uveitis.
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COVID-19 , Uveítis , Humanos , Femenino , Adulto , Masculino , COVID-19/complicaciones , COVID-19/diagnóstico , Estudios Retrospectivos , SARS-CoV-2 , Uveítis/diagnóstico , Uveítis/epidemiología , Uveítis/etiología , Medición de RiesgoRESUMEN
Background and Objectives: The identification of possible biomarkers that can predict treatment response among DME eyes is important for the individualization of treatment plans. We investigated optical coherence tomography (OCT)-based biomarkers that may predict the one-year real-life outcomes among diabetic macular edema (DME) eyes following treatment by intravitreal ranibizumab (IVR) injections. Materials and Methods: A total of 65 eyes from 35 treatment-naïve patients with DME treated with ranibizumab injection were recruited. Best-corrected visual acuity (BCVA), central retinal thickness (CRT), intraocular pressure (IOP), and OCT scans were retrospectively recorded at baseline before treatment and at 3 months, 6 months, and 12 months after treatment. The OCT scans were evaluated for biomarkers of interest, which included central retinal thickness (CRT), amount and locations of hyperreflective foci (HRF), subretinal fluid (SRF), intraretinal cysts (IRC), large outer nuclear layer cyst (LONLC), ellipsoid zone disruption (EZD), disorganization of retinal inner layers (DRIL), hard exudates (HE), epiretinal membrane (ERM), and vitreomacular interface (VMI). Correlations between these OCT biomarkers and outcome measures (visual and structural) were statistically analyzed. Results: A total of 65 eyes from 35 patients with DME were enrolled. The mean age was 64.2 ± 10.9 years old. Significant improvement in terms of mean BCVA (p < 0.005) and mean CRT was seen at final follow-up compared to baseline. The biomarkers of DRIL, LONLC, and SRF were found to be predictive for at least 50 µm CRT reduction after treatment (with odds ratio of 8.69, 8.5, and 17.58, respectively). The biomarkers of IRC, LONLC, and SRF were predictive for significant improvement in terms of BCVA and CRT after treatment. Finally, the number of HRF was predictive for both BCVA improvement and a CRT reduction of less than 100 µm after treatment. No serious complications were reported during the study. Conclusion: Our study demonstrated the utility of OCT biomarkers as therapeutic predictors of ranibizumab treatment among DME eyes.
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Diabetes Mellitus , Retinopatía Diabética , Edema Macular , Humanos , Persona de Mediana Edad , Anciano , Ranibizumab/uso terapéutico , Edema Macular/tratamiento farmacológico , Edema Macular/etiología , Tomografía de Coherencia Óptica/métodos , Estudios Retrospectivos , Inhibidores de la Angiogénesis/uso terapéutico , Estudios de Seguimiento , Resultado del Tratamiento , Biomarcadores , Diabetes Mellitus/tratamiento farmacológicoRESUMEN
The Golgi apparatus (GA) is the hub of intracellular trafficking, but selectively targeting GA remains a challenge. We show an unconventional types of peptide thioesters, consisting of an aminoethyl thioester and acting as substrates of thioesterases, for instantly targeting the GA of cells. The peptide thioesters, above or below their critical micelle concentrations, enter cells mainly via caveolin-mediated endocytosis or macropinocytosis, respectively. After being hydrolyzed by GA-associated thioesterases, the resulting thiopeptides form dimers and accumulate in the GA. After saturating the GA, the thiopeptides are enriched in the endoplasmic reticulum (ER). Their buildup in ER and GA disrupts protein trafficking, thus leading to cell death via multiple pathways. The peptide thioesters target the GA of a wide variety of cells, including human, murine, and Drosophila cells. Changing d-diphenylalanine to l-diphenylalanine in the peptide maintains the GA-targeting ability. In addition, targeting GA redirects protein (e.g., NRAS) distribution. This work illustrates a thioesterase-responsive and redox-active molecular platform for targeting the GA and controlling cell fates.
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Retículo Endoplásmico , Aparato de Golgi , Animales , Drosophila , Retículo Endoplásmico/metabolismo , Aparato de Golgi/metabolismo , Ratones , Péptidos/metabolismo , Fenilalanina/metabolismoRESUMEN
Neutrophils rely on glycolysis for energy production. How mitochondria regulate neutrophil function is not fully understood. Here, we report that mitochondrial outer membrane protein Mitofusin 2 (MFN2) regulates neutrophil homeostasis and chemotaxis in vivoMfn2-deficient neutrophils are released from the hematopoietic tissue, trapped in the vasculature in zebrafish embryos, and not capable of chemotaxis. Consistent with this, human neutrophil-like cells that are deficient for MFN2 fail to arrest on activated endothelium under sheer stress or perform chemotaxis on 2D surfaces. Deletion of MFN2 results in a significant reduction of neutrophil infiltration to the inflamed peritoneal cavity in mice. Mechanistically, MFN2-deficient neutrophil-like cells display disrupted mitochondria-ER interaction, heightened intracellular Ca2+ levels and elevated Rac activation after chemokine stimulation. Restoring a mitochondria-ER tether rescues the abnormal Ca2+ levels, Rac hyperactivation and chemotaxis defect resulting from MFN2 depletion. Finally, inhibition of Rac activation restores chemotaxis in MFN2-deficient neutrophils. Taken together, we have identified that MFN2 regulates neutrophil migration via maintaining the mitochondria-ER interaction to suppress Rac activation, and uncovered a previously unrecognized role of MFN2 in regulating cell migration and the actin cytoskeleton.This article has an associated First Person interview with the first authors of the paper.
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Neutrófilos , Pez Cebra , Citoesqueleto de Actina , Adhesivos , Animales , Movimiento Celular , RatonesRESUMEN
INTRODUCTION: The significance of endoplasmic reticulum (ER) stress in asthma is unclear. Here, we demonstrate that ER stress and the unfolded protein response (UPR) are related to disease severity and inflammatory phenotype. METHODS: Induced sputum (n=47), bronchial lavage (n=23) and endobronchial biopsies (n=40) were collected from participants with asthma with varying disease severity, inflammatory phenotypes and from healthy controls. Markers for ER stress and UPR were assessed. These markers were also assessed in established eosinophilic and neutrophilic murine models of asthma. RESULTS: Our results demonstrate increased ER stress and UPR pathways in asthma and these are related to clinical severity and inflammatory phenotypes. Genes associated with ER protein chaperone (BiP, CANX, CALR), ER-associated protein degradation (EDEM1, DERL1) and ER stress-induced apoptosis (DDIT3, PPP1R15A) were dysregulated in participants with asthma and are associated with impaired lung function (forced expiratory volume in 1 s) and active eosinophilic and neutrophilic inflammation. ER stress genes also displayed a significant correlation with classic Th2 (interleukin-4, IL-4/13) genes, Th17 (IL-17F/CXCL1) genes, proinflammatory (IL-1b, tumour necrosis factor α, IL-8) genes and inflammasome activation (NLRP3) in sputum from asthmatic participants. Mice with allergic airway disease (AAD) and severe steroid insensitive AAD also showed increased ER stress signalling in their lungs. CONCLUSION: Heightened ER stress is associated with severe eosinophilic and neutrophilic inflammation in asthma and may play a crucial role in the pathogenesis of asthma.
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Asma , Animales , Asma/metabolismo , Retículo Endoplásmico/metabolismo , Retículo Endoplásmico/patología , Humanos , Inflamación/metabolismo , Ratones , Neutrófilos/metabolismo , Transducción de Señal , Respuesta de Proteína DesplegadaRESUMEN
Neutrophil migration is essential for inflammatory responses to kill pathogens; however, excessive neutrophilic inflammation also leads to tissue injury and adverse effects. To discover novel therapeutic targets that modulate neutrophil migration, we performed a neutrophil-specific microRNA (miRNA) overexpression screen in zebrafish and identified 8 miRNAs as potent suppressors of neutrophil migration. Among those, miR-199 decreases neutrophil chemotaxis in zebrafish and human neutrophil-like cells. Intriguingly, in terminally differentiated neutrophils, miR-199 alters the cell cycle-related pathways and directly suppresses cyclin-dependent kinase 2 (Cdk2), whose known activity is restricted to cell cycle progression and cell differentiation. Inhibiting Cdk2, but not DNA replication, disrupts cell polarity and chemotaxis of zebrafish neutrophils without inducing cell death. Human neutrophil-like cells deficient in CDK2 fail to polarize and display altered signaling downstream of the formyl peptide receptor. Chemotaxis of primary human neutrophils is also reduced upon CDK2 inhibition. Furthermore, miR-199 overexpression or CDK2 inhibition significantly improves the outcome of lethal systemic inflammation challenges in zebrafish. Our results therefore reveal previously unknown functions of miR-199 and CDK2 in regulating neutrophil migration and provide directions in alleviating systemic inflammation.
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Quimiotaxis de Leucocito/genética , Quinasa 2 Dependiente de la Ciclina/genética , MicroARNs/metabolismo , Neutrófilos/inmunología , Síndrome de Respuesta Inflamatoria Sistémica/inmunología , Animales , Animales Modificados Genéticamente , Línea Celular Tumoral , Quimiotaxis de Leucocito/efectos de los fármacos , Quimiotaxis de Leucocito/inmunología , Quinasa 2 Dependiente de la Ciclina/antagonistas & inhibidores , Quinasa 2 Dependiente de la Ciclina/inmunología , Modelos Animales de Enfermedad , Regulación hacia Abajo/inmunología , Técnicas de Silenciamiento del Gen , Humanos , Larva , Cultivo Primario de Células , Inhibidores de Proteínas Quinasas/farmacología , Transducción de Señal/genética , Transducción de Señal/inmunología , Síndrome de Respuesta Inflamatoria Sistémica/genética , Pez CebraRESUMEN
Background and objectives: To report the initial response to a single intravitreal brolucizumab (IVI-B) injection in wet age-related macular degeneration (wAMD) or polypoidal choroidopathy (PCV) complicated with either persistent subretinal fluid (SRF) or pigment epithelial detachment refractory to previous anti-vascular endothelial growth factor (anti-VEGF) therapy. Material and methods: In this retrospective study, all eyes received a single IVI-B (6 mg/0.05 mL) for wAMD or PCV with treatment-resistant SRF or PED. Outcome measures included assessment in central retinal thickness (CRT), visual acuity, and evaluation for changes in the SRF or PED on OCT. Follow-up was prior to the first brolucizumab injection, then at 1 week and 5 weeks afterwards. Results: In total, 10 eyes of 10 patients (6 women [60%]) were enrolled. Five patients had wAMD and five patients had PCV. Average age of participants was 67.6 years. All patients received one IVI-B. All patients were not treatment-naïve to anti-VEGF agents. At the first week and fifth week following the first IVI-B, seven out of seven patients (100%) had resolved SRF. However, seven out of nine patients (78%) had no improvement of their PED at 5 weeks follow-up. Mean PED height and width before the first IVI-B was 339.77 µm and 2233.44 µm, respectively. Mean PED height and width at the fifthweek following the first IVI-B was 328.125 µm and 2129.5 µm, respectively. Overall mean visual acuity before the first IVI-B was 0.224; and 5 weeks following the first IVI-B was 0.38. Conclusions: Treatment with brolucizumab resulted in anatomical improvement for all patients with persistent SRF. Limited efficacy was seen for persistent PED. Brolucizumab appears to be a safe and effective option for treatment-resistant SRF. Future multicenter collaborative studies are warranted.
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Inhibidores de la Angiogénesis , Degeneración Macular Húmeda , Anciano , Femenino , Humanos , Inhibidores de la Angiogénesis/uso terapéutico , Factores de Crecimiento Endotelial , Estudios Retrospectivos , Tomografía de Coherencia Óptica/métodos , Degeneración Macular Húmeda/tratamiento farmacológico , MasculinoRESUMEN
BACKGROUND AND OBJECTIVE: Influenza virus (FLU), rhinovirus (RV) and respiratory syncytial virus (RSV) are the most common acute respiratory infections worldwide. Infection can cause severe health outcomes, while therapeutic options are limited, primarily relieving symptoms without attenuating the development of lesions or impaired lung function. We therefore examined the inflammatory response to these infections with the intent to identify common components that are critical drivers of immunopathogenesis and thus represent potential therapeutic targets. METHODS: BALB/c mice were infected with FLU, RV or RSV, and lung function, airway inflammation and immunohistopathology were measured over a 10-day period. Anti-IL-17A mAb was administered to determine the impact of attenuating this cytokine's function on the development and severity of disease. RESULTS: All three viruses induced severe airway constriction and inflammation at 2 days post-infection (dpi). However, only FLU induced prolonged inflammation till 10 dpi. Increased IL-17A expression was correlated with the alterations in lung function and its persistence. Neutralization of IL-17A did not affect the viral replication but led to the resolution of airway hyperresponsiveness. Furthermore, anti-IL-17A treatment resulted in reduced infiltration of neutrophils (in RV- and FLU-infected mice at 2 dpi) and lymphocytes (in RSV-infected mice at 2 dpi and FLU-infected mice at 10 dpi), and attenuated the severity of immunopathology. CONCLUSION: IL-17A is a common pathogenic molecule regulating disease induced by three prevalent respiratory viruses. Targeting the IL-17A pathway may provide a unified approach to the treatment of these respiratory infections alleviating both inflammation-induced lesions and difficulties in breathing.
Asunto(s)
Interleucina-17/inmunología , Infecciones por Orthomyxoviridae/inmunología , Infecciones por Picornaviridae/inmunología , Infecciones por Virus Sincitial Respiratorio , Animales , Pulmón/fisiopatología , Ratones , Ratones Endogámicos BALB C , Orthomyxoviridae , Virus Sincitiales Respiratorios/inmunología , RhinovirusRESUMEN
BACKGROUND AND OBJECTIVE: COVID-19 is complicated by acute lung injury, and death in some individuals. It is caused by SARS-CoV-2 that requires the ACE2 receptor and serine proteases to enter AEC. We determined what factors are associated with ACE2 expression particularly in patients with asthma and COPD. METHODS: We obtained lower AEC from 145 people from two independent cohorts, aged 2-89 years, Newcastle (n = 115) and Perth (n = 30), Australia. The Newcastle cohort was enriched with people with asthma (n = 37) and COPD (n = 38). Gene expression for ACE2 and other genes potentially associated with SARS-CoV-2 cell entry was assessed by qPCR, and protein expression was confirmed with immunohistochemistry on endobronchial biopsies and cultured AEC. RESULTS: Increased gene expression of ACE2 was associated with older age (P = 0.03) and male sex (P = 0.03), but not with pack-years smoked. When we compared gene expression between adults with asthma, COPD and healthy controls, mean ACE2 expression was lower in asthma patients (P = 0.01). Gene expression of furin, a protease that facilitates viral endocytosis, was also lower in patients with asthma (P = 0.02), while ADAM-17, a disintegrin that cleaves ACE2 from the surface, was increased (P = 0.02). ACE2 protein expression was also reduced in endobronchial biopsies from asthma patients. CONCLUSION: Increased ACE2 expression occurs in older people and males. Asthma patients have reduced expression. Altered ACE2 expression in the lower airway may be an important factor in virus tropism and may in part explain susceptibility factors and why asthma patients are not over-represented in those with COVID-19 complications.