RESUMEN
Rationale: The resolution of inflammation is an active process coordinated by mediators and immune cells to restore tissue homeostasis. However, the mechanisms for resolving eosinophilic allergic lung inflammation triggered by inhaled allergens have not been fully elucidated. Objectives: Our objectives were to investigate the cellular mechanism of tissue-resident macrophages involved in the resolution process of eosinophilic lung inflammation. Methods: For the study, we used the institutional review board-approved protocol for human subsegmental bronchoprovocation with allergen, mouse models for allergic lung inflammation, and novel transgenic mice, including a conditional CCL26 knockout. The samples were analyzed using mass cytometry, single-cell RNA sequencing, and biophysical and immunological analyses. Measurements and Main Results: We compared alveolar macrophage (AM) subsets in the BAL before and after allergen provocation. In response to provocation with inhaled allergens, the subsets of AMs are dynamically changed in humans and mice. In the steady state, the AM subset expressing CX3CR1 is a relatively small fraction in bronchoalveolar space and lung tissue but drastically increases after allergen challenges. This subset presents unique patterns of gene expression compared with classical AMs, expressing high C1q family genes. CX3CR1+ macrophages are activated by airway epithelial cell-derived CCL26 via a receptor-ligand interaction. The binding of CCL26 to the CX3CR1+ receptor induces CX3CR1+ macrophages to secrete C1q, subsequently facilitating the clearance of eosinophils. Furthermore, the depletion of CX3CR1 macrophages or CCL26 in airway epithelial cells delays the resolution of allergic lung inflammation displaying prolonged tissue eosinophilia. Conclusions: These findings indicate that the CCL26-CX3CR1 pathway is pivotal in resolving eosinophilic allergic lung inflammation.
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Alveolitis Alérgica Extrínseca , Hipersensibilidad , Neumonía , Eosinofilia Pulmonar , Humanos , Ratones , Animales , Complemento C1q/metabolismo , Pulmón/metabolismo , Macrófagos , Alérgenos , Inflamación/metabolismo , Neumonía/metabolismo , Quimiocina CCL26/metabolismoRESUMEN
BACKGROUND: Dendritic cells (DCs) are heterogeneous, comprising multiple subsets with unique functional specifications. Our previous work has demonstrated that the specific conventional type 2 DC subset, CSF1R+cDC2s, plays a critical role in sensing aeroallergens. OBJECTIVE: It remains to be understood how CSF1R+cDC2s recognize inhaled allergens. We sought to elucidate the transcriptomic programs and receptor-ligand interactions essential for function of this subset in allergen sensitization. METHODS: We applied single-cell RNA sequencing to mouse lung DCs. Conventional DC-selective knockout mouse models were employed, and mice were subjected to inhaled allergen sensitization with multiple readouts of asthma pathology. Under the clinical arm of this work, human lung transcriptomic data were integrated with mouse data, and bronchoalveolar lavage (BAL) specimens were collected from subjects undergoing allergen provocation, with samples assayed for C1q. RESULTS: We found that C1q is selectively enriched in lung CSF1R+cDC2s, but not in other lung cDC2 or cDC1 subsets. Depletion of C1q in conventional DCs significantly attenuates allergen sensing and features of asthma. Additionally, we found that C1q binds directly to human dust mite allergen, and the C1q receptor CD91 (LRP1) is required for lung CSF1R+cDC2s to recognize the C1q-allergen complex and induce allergic lung inflammation. Lastly, C1q is enriched in human BAL samples following subsegmental allergen challenge, and human RNA sequencing data demonstrate close homology between lung IGSF21+DCs and mouse CSF1R+cDC2s. CONCLUSIONS: C1q is secreted from the CSF1R+cDC2 subset among conventional DCs. Our data indicate that the C1q-LRP1 axis represents a candidate for translational therapeutics in the prevention and suppression of allergic lung inflammation.
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Asma , Neumonía , Animales , Humanos , Ratones , Alérgenos/metabolismo , Asma/metabolismo , Complemento C1q/metabolismo , Células Dendríticas , Ratones Noqueados , Neumonía/metabolismo , Proteínas Tirosina Quinasas Receptoras , Receptores del Factor Estimulante de Colonias/metabolismoRESUMEN
VEGFR2 signaling in endothelial cells (ECs) is regulated by reactive oxygen species (ROS) derived from NADPH oxidases (NOXs) and mitochondria, which plays an important role in postnatal angiogenesis. However, it remains unclear how highly diffusible ROS signal enhances VEGFR2 signaling and reparative angiogenesis. Protein disulfide isomerase A1 (PDIA1) functions as an oxidoreductase depending on the redox environment. We hypothesized that PDIA1 functions as a redox sensor to enhance angiogenesis. Here we showed that PDIA1 co-immunoprecipitated with VEGFR2 or colocalized with either VEGFR2 or an early endosome marker Rab5 at the perinuclear region upon stimulation of human ECs with VEGF. PDIA1 silencing significantly reduced VEGF-induced EC migration, proliferation and spheroid sprouting via inhibiting VEGFR2 signaling. Mechanistically, VEGF stimulation rapidly increased Cys-OH formation of PDIA1 via the NOX4-mitochondrial ROS axis. Overexpression of "redox-dead" mutant PDIA1 with replacement of the active four Cys residues with Ser significantly inhibited VEGF-induced PDIA1-CysOH formation and angiogenic responses via reducing VEGFR2 phosphorylation. Pdia1+/- mice showed impaired angiogenesis in developmental retina and Matrigel plug models as well as ex vivo aortic ring sprouting model. Study using hindlimb ischemia model revealed that PDIA1 expression was markedly increased in angiogenic ECs of ischemic muscles, and that ischemia-induced limb perfusion recovery and neovascularization were impaired in EC-specific Pdia1 conditional knockout mice. These results suggest that PDIA1 can sense VEGF-induced H2O2 signal via CysOH formation to promote VEGFR2 signaling and angiogenesis in ECs, thereby enhancing postnatal angiogenesis. The oxidized PDIA1 is a potential therapeutic target for treatment of ischemic vascular diseases.
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Células Endoteliales , Proteína Disulfuro Isomerasas , Ratones , Humanos , Animales , Células Endoteliales/metabolismo , Proteína Disulfuro Isomerasas/genética , Proteína Disulfuro Isomerasas/metabolismo , Especies Reactivas de Oxígeno/metabolismo , Factor A de Crecimiento Endotelial Vascular/metabolismo , Peróxido de Hidrógeno/metabolismo , Neovascularización Fisiológica , Oxidación-Reducción , Receptor 2 de Factores de Crecimiento Endotelial Vascular/genética , Receptor 2 de Factores de Crecimiento Endotelial Vascular/metabolismo , Isquemia/metabolismoRESUMEN
BACKGROUND: In recent years, researchers have been examining the impact of work-life balance (WLB) on mental health, considering it as a potential risk factor. However, it remains unclear whether the traditional understanding of WLB applies to older adults who worked for fewer hours before full retirement and whose children are likely to be independent adults. Therefore, this study aims to propose a modified form of WLB specifically for older adults. Within this context, we hypothesize that an optimum balance between working hours and social engagement protects against depressive symptoms among older adults. METHOD: We conducted an analysis using data on 5,751 Korean adults older than 55 years from the Korean Longitudinal Study of Aging 2016. Multivariate logistic regression analysis was used to evaluate the relationships among working hours, social engagement, and depressive symptoms. RESULTS: Older adults who worked fewer than 35 h per week were less likely to experience depressive symptoms than were non-working older adults and those working 35 h or more per week. Additionally, older adults with a high level of informal social participation, thus occurring almost every day or two to three times per week, were less likely to experience depressive symptoms than were those with a low level of such participation (once a month or less). Furthermore, depressive symptoms were less frequent among those who worked fewer than 35 h per week and engaged in a high level of informal social participation compared to non-working older individuals and those with a low level of informal social participation. CONCLUSIONS: Maintaining an optimal number of working hours and degree of social engagement are necessary to minimize the risk of depressive symptoms in older adults. Based on these findings, we suggest that fulfillment for work and life and their balance are important for older adults and propose work-life fulfillment balance.
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Depresión , Participación Social , Niño , Humanos , Anciano , Depresión/epidemiología , Depresión/psicología , Participación Social/psicología , Estudios Longitudinales , Equilibrio entre Vida Personal y Laboral , Envejecimiento/psicologíaRESUMEN
Pluripotent stem cells are known to shift their mitochondrial metabolism upon differentiation, but the mechanisms underlying such metabolic rewiring are not fully understood. We hypothesized that during differentiation of human induced pluripotent stem cells (hiPSCs), mitochondria undergo mitophagy and are then replenished by the biogenesis of new mitochondria adapted to the metabolic needs of the differentiated cell. To evaluate mitophagy during iPSC differentiation, we performed live cell imaging of mitochondria and lysosomes in hiPSCs differentiating into vascular endothelial cells using confocal microscopy. We observed a burst of mitophagy during the initial phases of hiPSC differentiation into the endothelial lineage, followed by subsequent mitochondrial biogenesis as assessed by the mitochondrial biogenesis biosensor MitoTimer. Furthermore, hiPSCs undergoing differentiation showed greater mitochondrial oxidation of fatty acids and an increase in ATP levels as assessed by an ATP biosensor. We also found that during mitophagy, the mitochondrial phosphatase PGAM5 is cleaved in hiPSC-derived endothelial progenitor cells and in turn activates ß-catenin-mediated transcription of the transcriptional coactivator PGC-1α, which upregulates mitochondrial biogenesis. These data suggest that mitophagy itself initiates the increase in mitochondrial biogenesis and oxidative metabolism through transcriptional changes during endothelial cell differentiation. In summary, these findings reveal a mitophagy-mediated mechanism for metabolic rewiring and maturation of differentiating cells via the ß-catenin signaling pathway. We propose that such mitochondrial-nuclear cross talk during hiPSC differentiation could be leveraged to enhance the metabolic maturation of differentiated cells.
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Reprogramación Celular , Células Endoteliales , Células Madre Pluripotentes Inducidas/metabolismo , Mitofagia , Humanos , Proteínas Mitocondriales/metabolismo , Fosforilación Oxidativa , Fosfoproteínas Fosfatasas/metabolismo , Transcripción Genética , beta Catenina/metabolismoRESUMEN
PURPOSE: Probiotics and prebiotics are commonly used to improve the gut microbiota. Since prebiotics can support the growth of probiotics, co-administration of these is called synbiotics. It has been demonstrated that obesity-induced gut dysbiosis can worsen inflammatory bowel disease symptoms. This study evaluated how modulation of gut microbiota with Schizophyllum commune-derived ß-glucan (BG), probiotics (PRO), and synbiotics containing both BG and PRO (SYN) could improve the symptoms of obesity-associated colitis and hepatic manifestation. METHODS: Mice were fed a normal diet (ND), high-fat diet (HFD), and HFD with different additives (BG, PRO, and SYN) for 12 weeks, followed by 5 days of colitis induction. Mice were sacrificed before and after colitis induction. During the experiment, body weight, food and water consumption, and rectal bleeding were monitored. Proteins from the colon were subjected to western blotting, and serum biomarkers such as alanine transaminase, alkaline phosphatase, triglycerides, and total cholesterol were analyzed. Colon and liver samples were sectioned for histological analysis. The fecal microbiota was analyzed based on partial 16S rRNA gene sequences. RESULTS: Although BG and PRO secured intestinal tight junctions, these two treatments did not modulate inflammatory cell infiltration and inflammatory markers (i.e., IL-6 and TNF-α). In contrast, SYN demonstrated stronger and broader effects in reducing colonic inflammation. While BG treatment increased the abundance of indigenous Lactobacillus, PRO treatment decreased bacterial diversity by suppressing the growth of several species of bacteria. SYN treatment groups, however, supported the growth of both indigenous and supplemented bacteria while maintaining bacterial diversity. CONCLUSION: Obesity-associated colitis can be improved by modulating gut bacteria with ß-glucan and probiotics. The co-administration of both outperformed ß-glucan and probiotic treatment alone by fostering both indigenous and supplemented probiotic strains.
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Colitis , Probióticos , Simbióticos , beta-Glucanos , Animales , Colitis/metabolismo , Hígado/metabolismo , Ratones , Ratones Endogámicos C57BL , Obesidad/metabolismo , Probióticos/farmacología , ARN Ribosómico 16S/genética , beta-Glucanos/farmacologíaRESUMEN
PURPOSE: Western diet, rich in carbohydrates and fat, is said to be a major factor underlying metabolic syndrome. Interventions with prebiotics, the key modulators of the gut microbiota, have paramount impact on host-associated metabolic disorders. Herein, we investigated the effect of fungus-derived (1,3)/(1,6)-ß-glucan, a highly soluble dietary fiber, on high-fat diet (HFD)-induced metabolic distress. METHODS: Male C57BL/6 J mice were fed with different diet groups (n = 11): control diet, HFD, 3 g/kg or 5 g/kg of ß-glucan-incorporated HFD. At the end of experimental study period (12th week), body weight, feces weight and fecal moisture content were observed. Further, colonic motility was measured using activated charcoal meal study. Proteins extracted from liver and intestine tissues were subjected to western blot technique. Paraffin-embedded intestinal tissues were sectioned for histochemical [Periodic acid-Schiff (PAS) and Alcian blue (AB) staining] analysis. Fecal microbiota analysis was performed using MOTHUR bioinformatic software. RESULTS: ß-glucan consumption exhibited anti-obesity property in mice groups fed with HFD. In addition, ß-glucan ameliorated HFD-induced hepatic stress, colonic motility and intestinal atrophy (reduction in colon length, goblet cells, and mucosal layer thickness). Further, ß-glucan incorporation shifted bacterial community by increasing butyrate-producing bacteria such as Anaerostipes, Coprobacillus, and Roseburia and decreasing reportedly obesity-associated bacteria such as Parabacteroides and Lactococcus. CONCLUSION: Altogether, the outcomes of this present pre-clinical animal study show ß-glucan to be a promising therapeutic candidate in the treatment of HFD-induced metabolic distress. Further comprehensive research has to be conducted to brace its clinical relevance, reproducibility and efficacy for aiding human health.
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Microbioma Gastrointestinal , beta-Glucanos , Animales , Dieta Alta en Grasa/efectos adversos , Hongos , Masculino , Ratones , Ratones Endogámicos C57BL , Prebióticos , Reproducibilidad de los ResultadosRESUMEN
BACKGROUND: Work-life conflict (WLC) has a critical effect on employee mental health. However, research on occupational health has neglected the family domain. Furthermore, although it is reasonable to assume that the effect of WLC on health may differ according to socioeconomic circumstances, there is little empirical evidence for differences in the impact of WLC by socioeconomic status (SES). The purpose of this study was to assess the role of SES as an effect modifier, while examining whether the SES level affects the relationship between WLC and mental health. METHOD: We analyzed data from the nationally representative South Korean Working Conditions Survey of 2014, including 49 401 workers. Logistic regression analyses, stratified by sexes, were performed to identify sex differences, and interaction terms including WLC and SES were also incorporated. RESULTS: WLC (men: OR = 1.24; women: OR = 1.18) and domestic demands (men: OR = 1.16; women: OR = 1.22) were significantly associated with mental health. WLC exhibited a stronger association with mental health for individuals with high SES, both in terms of education (men: OR = 1.61 vs 1.51; women: OR = 1.52 vs 1.24) and income (men: OR = 1.44 vs 1.10; women: OR = 1.48 vs 1.20). CONCLUSIONS: Our data suggest that future efforts for health promotion should consider workers' family demands and SES as important modifying factors of psychological health in the workplace.
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Trastornos Mentales/epidemiología , Enfermedades Profesionales/epidemiología , Clase Social , Equilibrio entre Vida Personal y Laboral/estadística & datos numéricos , Lugar de Trabajo/psicología , Adulto , Estudios Transversales , Femenino , Humanos , Modelos Logísticos , Masculino , Trastornos Mentales/psicología , Persona de Mediana Edad , Enfermedades Profesionales/psicología , Prevalencia , República de Corea/epidemiología , Adulto JovenRESUMEN
PURPOSE: The purpose of this paper is to identify the relationships between structural empowerment and patient identification behaviors of nurses. DESIGN/METHODOLOGY/APPROACH: The present study was a descriptive survey using a self-reported questionnaire, following a quality improvement project at a hospital in South Korea. The participants included 984 registered nurses, who administer medication and transfusions to patients in the hospital. Data were analyzed using the t-test, ANOVA, Scheffé's test, Pearson correlation coefficients and multiple regression analysis. FINDINGS: The patient identification behaviors of nurses were significantly correlated with opportunity, support, information, resources, formal power and informal power of structural empowerment. The support, information and informal power of structural empowerment, as well as the age and gender of the participants explained 10.7 percent of the variance in the patient identification behaviors of nurses. RESEARCH LIMITATIONS/IMPLICATIONS: The present study has some limitations. Although the data collected by the cross-sectional survey were analyzed, causal analysis could not have been conducted. Nursing managers can promote safety by creating a work environment that facilitates access to the support, information and resources needed for nurses to perform their duties effectively; providing opportunities for nurses to learn and develop professionally; acknowledging the achievements of nurses; and expanding their duties, so that nurses can demonstrate greater work flexibility. Future studies should investigate structural empowerment in multiple nursing organizations, and particularly the organizational characteristics that affect structural empowerment. ORIGINALITY/VALUE: The present study confirms that structural empowerment influences the patient identification behaviors of nurses.
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Empoderamiento , Satisfacción en el Trabajo , Personal de Enfermería en Hospital , Adulto , Estudios Transversales , Femenino , Humanos , Masculino , Errores de Medicación/prevención & control , Autoinforme , Adulto JovenRESUMEN
Cutaneous wound repair is an intricate process whereby the skin reprograms itself after injury. In the mid-phase of wound repair, the proliferation, migration, and differentiation of cells are the major mechanisms to lead remodeling. We investigated the effect of BMM ((1E,2E)-1,2-bis((6-bromo-2H-chromen-3-yl)methylene)hydrazine), a novel synthetic material, on the migration and viability of keratinocytes or fibroblasts using the in vitro scratch woundhealing, electric cell-substrate imedance sensing (ECIS), invasion, and MTT assays. Cell migration-related factors were analyzed using western blot, and we found that treatment with BMM stimulated the EMT pathway and focal adhesion kinase (FAK)/Src signaling. Differentiation of HaCaT keratinocyte and fibroblast cells was also stimulated by BMM and specifically, NOX2/4 contributed to the activation of fibroblasts for wound healing. Furthermore, BMM treated HaCaT keratinocyte and fibroblast-co-cultured cells increased migration and differentiation. TGF-ß and Cyr61 were also secreted to a greater extent than in single cultured cells. In vivo experiments showed that treatment with BMM promotes wound closure by promoting re-epithelialization. In this study, we demonstrated that a novel synthetic material, BMM, is capable of promoting wound healing via the stimulation of re-epithelialization in the epidermis and the activation of fibroblasts in the dermis, in particular, via the acceleration of the interaction between the epidermis and dermis.
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Benzopiranos/farmacología , Fibroblastos/efectos de los fármacos , Hidrazinas/farmacología , Repitelización , Animales , Benzopiranos/química , Línea Celular , Movimiento Celular , Proliferación Celular , Células Cultivadas , Proteína 61 Rica en Cisteína/metabolismo , Fibroblastos/metabolismo , Fibroblastos/fisiología , Proteína-Tirosina Quinasas de Adhesión Focal/metabolismo , Humanos , Hidrazinas/química , Queratinocitos/efectos de los fármacos , Queratinocitos/metabolismo , Queratinocitos/fisiología , Masculino , Ratones , Ratones Endogámicos ICR , NADPH Oxidasas/metabolismo , Transducción de Señal , Factor de Crecimiento Transformador beta/metabolismo , Familia-src Quinasas/metabolismoRESUMEN
Reactive oxygen species (ROS) derived from NADPH oxidase (NOX) and mitochondria play a critical role in growth factor-induced switch from a quiescent to an angiogenic phenotype in endothelial cells (ECs). However, how highly diffusible ROS produced from different sources can coordinate to stimulate VEGF signaling and drive the angiogenic process remains unknown. Using the cytosol- and mitochondria-targeted redox-sensitive RoGFP biosensors with real-time imaging, here we show that VEGF stimulation in human ECs rapidly increases cytosolic RoGFP oxidation within 1 min, followed by mitochondrial RoGFP oxidation within 5 min, which continues at least for 60 min. Silencing of Nox4 or Nox2 or overexpression of mitochondria-targeted catalase significantly inhibits VEGF-induced tyrosine phosphorylation of VEGF receptor type 2 (VEGFR2-pY), EC migration and proliferation at the similar extent. Exogenous hydrogen peroxide (H2O2) or overexpression of Nox4, which produces H2O2, increases mitochondrial ROS (mtROS), which is prevented by Nox2 siRNA, suggesting that Nox2 senses Nox4-derived H2O2 to promote mtROS production. Mechanistically, H2O2 increases S36 phosphorylation of p66Shc, a key mtROS regulator, which is inhibited by siNox2, but not by siNox4. Moreover, Nox2 or Nox4 knockdown or overexpression of S36 phosphorylation-defective mutant p66Shc(S36A) inhibits VEGF-induced mtROS, VEGFR2-pY, EC migration, and proliferation. In summary, Nox4-derived H2O2 in part activates Nox2 to increase mtROS via pSer36-p66Shc, thereby enhancing VEGFR2 signaling and angiogenesis in ECs. This may represent a novel feed-forward mechanism of ROS-induced ROS release orchestrated by the Nox4/Nox2/pSer36-p66Shc/mtROS axis, which drives sustained activation of angiogenesis signaling program.
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Retroalimentación Fisiológica , Peróxido de Hidrógeno/metabolismo , Glicoproteínas de Membrana/genética , Mitocondrias/metabolismo , NADPH Oxidasas/genética , Transducción de Señal , Técnicas Biosensibles , Catalasa/genética , Catalasa/metabolismo , Movimiento Celular/efectos de los fármacos , Proliferación Celular/efectos de los fármacos , Regulación de la Expresión Génica , Células Endoteliales de la Vena Umbilical Humana , Humanos , Glicoproteínas de Membrana/antagonistas & inhibidores , Glicoproteínas de Membrana/metabolismo , Microscopía Fluorescente , Mitocondrias/efectos de los fármacos , NADPH Oxidasa 2 , NADPH Oxidasa 4 , NADPH Oxidasas/antagonistas & inhibidores , NADPH Oxidasas/metabolismo , Neovascularización Fisiológica/efectos de los fármacos , Neovascularización Fisiológica/genética , Oxidación-Reducción , Fosforilación/efectos de los fármacos , ARN Interferente Pequeño/genética , ARN Interferente Pequeño/metabolismo , Proteína Transformadora 1 que Contiene Dominios de Homología 2 de Src/genética , Proteína Transformadora 1 que Contiene Dominios de Homología 2 de Src/metabolismo , Imagen de Lapso de Tiempo , Factor A de Crecimiento Endotelial Vascular/farmacología , Receptor 2 de Factores de Crecimiento Endotelial Vascular/genética , Receptor 2 de Factores de Crecimiento Endotelial Vascular/metabolismoRESUMEN
The transmembrane NS4B protein of dengue virus (DENV) is a validated antiviral target that plays important roles in viral replication and invasion of innate immune response. The first 125 amino acids of DENV NS4B are sufficient for inhibition of alpha/beta interferon signaling. Resistance mutations to NS4B inhibitors are all mapped to the first 125 amino acids. In this study, we expressed and purified a protein representing the first 125 amino acids of NS4B (NS4B(1-125)). This recombinant NS4B(1-125) protein was reconstituted into detergent micelles. Solution NMR spectroscopy demonstrated that there are five helices (α1 to α5) present in NS4B(1-125). Dynamic studies, together with a paramagnetic relaxation enhancement experiment demonstrated that four helices, α2, α3, α4, and α5 are embedded in the detergent micelles. Comparison of wild type and V63I mutant (a mutation that confers resistance to NS4B inhibitor) NS4B(1-125) proteins demonstrated that V63I mutation did not cause significant conformational changes, however, V63 may have a molecular interaction with residues in the α5 transmembrane domain under certain conditions. The structural and dynamic information obtained in study is helpful to understand the structure and function of NS4B.
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Virus del Dengue/química , Proteínas no Estructurales Virales/química , Dicroismo Circular , Mutación , Resonancia Magnética Nuclear Biomolecular , Estructura Secundaria de Proteína , Proteínas no Estructurales Virales/genéticaRESUMEN
Flavonoids are plant secondary compounds with various pharmacological properties. We previously showed that one flavonoid, trimethoxyisoflavone (TMF), could promote wound healing by inducing keratinocyte migration. Here, we screened TMF derivatives for enhanced activity and identified one compound, 2',6 Dichloro-7-methoxyisoflavone (DCMF), as most effective at promoting migration in a scratch wound assay. Using the HaCaT keratinocyte cell line, we found DCMF treatment induced phosphorylation of both FAK and Src, and increased keratinocyte migration. DCMF-induced Src kinase could promote activation of ERK, AKT, and p38 signaling pathways, and DCMF-induced secretion of matrix metalloproteinase (MMP)-2 and MMP-9 and partial epithelial-mesenchymal transition (EMT), whereas Src inhibition abolished DCMF-induced EMT. Using an in vivo excisional wound model, we observed improved wound closure and re-epithelialization in DCMF-treated mice, as compared to controls. Collectively, our data demonstrate that DCMF induces cell migration and promotes wound healing through activation of Src/FAK, ERK, AKT, and p38 MAPK signaling.
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Movimiento Celular/fisiología , Quinasa 1 de Adhesión Focal/metabolismo , Isoflavonas/administración & dosificación , Queratinocitos/fisiología , Cicatrización de Heridas/efectos de los fármacos , Familia-src Quinasas/metabolismo , Animales , Línea Celular , Movimiento Celular/efectos de los fármacos , Relación Dosis-Respuesta a Droga , Activación Enzimática/efectos de los fármacos , Humanos , Isoflavonas/síntesis química , Queratinocitos/efectos de los fármacos , Masculino , Ratones , Ratones Endogámicos ICR , Transducción de Señal/efectos de los fármacos , Transducción de Señal/fisiología , Cicatrización de Heridas/fisiologíaRESUMEN
AIM: This empirical research aimed to identify relationships between nurses' unit tenure, nursing unit tenure diversity and medication errors. BACKGROUND: Research examining medication errors has paid little attention to the effects of multilevel precursors. METHOD: In total, 567 registered nurses (from 36 nursing units) completed a survey questionnaire at a university hospital during September 2012. Of these, 334 (completed by nurses from 22 nursing units) were eligible for multilevel analysis. RESULTS: The average frequency of self-reported medication errors per registered nurse in the preceding 6 months was 0.98. Multilevel analysis showed that medication errors were significantly negatively associated with nurses' unit tenure at individual level (B = -0.64, P = 0.002) and nursing unit tenure diversity at unit level (B = -0.69, P < 0.001). Furthermore, nursing unit tenure diversity moderated the relationship between nurses' unit tenure and medication errors (B = 0.48, P = 0.012). CONCLUSION: This study provided evidence indicating that novice nurses made a higher number of medication errors relative to experienced nurses, and that including a mixture of novice and experienced nurses in a nursing unit attenuated novice nurses' medication errors. IMPLICATIONS FOR NURSING MANAGEMENT: The development of staffing strategies that enhance nursing unit tenure diversity is required.
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Errores de Medicación/enfermería , Enfermeras y Enfermeros/normas , Habitaciones de Pacientes , Factores de Tiempo , Adulto , Femenino , Hospitales Universitarios/organización & administración , Humanos , Relaciones Interprofesionales , Masculino , Persona de Mediana Edad , Análisis Multinivel , Cultura Organizacional , Habitaciones de Pacientes/normas , Autoinforme , Encuestas y Cuestionarios , Recursos HumanosRESUMEN
BACKGROUND: Wound healing is an intricate process whereby the skin repairs itself after injury. The epithelial-mesenchymal transition (EMT) is associated with wound healing and tissue regeneration. Naphthochalcone derivatives have various pharmaceutical properties. We investigated the effect of a novel naphthochalcone derivative, 2-(5-(2,4,6-trimethoxyphenyl)-4,5-dihydro-1H-pyrazol-3-yl)naphthalen-1-ol (TDPN), on dermal wound healing in vivo and the migration of keratinocytes in vitro. RESULT: We investigated the effect of TDPN on signaling pathway and epithelial-mesenchymal transition through protein and transcriptional expression. The TDPN treatment accelerated dermal closure about 3 days and remodeling of dermis. We found that treatment with TDPN induced the migration of keratinocytes but not cytotoxicity. TDPN induced the phosphorylation of ERK and AKT. TDPN-treated cells showed loss of adherence protein and showed induction of the transcriptional factor Slug, mesenchymal marker, and fibronectin. Moreover, TDPN treatment induced the expression of matrix metalloproteinase-1 (MMP-1), which degrades specific components of the extracellular matrix, thereby providing new substrates that facilitate migration and invasion. MMP expression is considered to be one of the major attributes acquired by cells after EMT. CONCLUSION: We propose that a novel naphthochalcone derivative TDPN is capable of promoting keratinocyte migration via the induction of EMT resulting acceleration of wound closure and matrix remodeling.
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Transición Epitelial-Mesenquimal/efectos de los fármacos , Queratinocitos/efectos de los fármacos , Metaloproteinasa 1 de la Matriz/genética , Naftoles/administración & dosificación , Pirazoles/administración & dosificación , Cicatrización de Heridas/efectos de los fármacos , Animales , Movimiento Celular/efectos de los fármacos , Matriz Extracelular/efectos de los fármacos , Matriz Extracelular/metabolismo , Regulación de la Expresión Génica/efectos de los fármacos , Humanos , Queratinocitos/metabolismo , Queratinocitos/patología , Metaloproteinasa 1 de la Matriz/biosíntesis , Naftalenos/administración & dosificación , Fosforilación , Ratas , Transducción de Señal/efectos de los fármacos , Piel/efectos de los fármacos , Piel/patología , Cicatrización de Heridas/genéticaRESUMEN
Glycitin is a soy isoflavone that exhibits antioxidant, antiallergic, and anti-osteoporosis activities. We investigated the effects of glycitin on dermal fibroblast proliferation and migration. Treatment of primary dermal fibroblasts with glycitin increased cell proliferation and migration. In addition, treatment with 20 µM glycitin for 24 h induced the synthesis of collagen type I and type III at both the mRNA and protein levels. Fibronectin was also increased by 20% after treatment. Matrix metalloproteinase-1 collagenase was decreased in the media after 24-h incubation with glycitin, and the synthesis of transforming growth factor-beta (TGF-ß) mRNA increased approximately twofold in cells following glycitin treatment. Phosphorylation of Smad2 and Smad3 increased after 1 h of glycitin treatment, and phosphorylation continued for 24 h. Furthermore, the phosphorylated form of AKT was increased in glycitin-treated cells after 3 h and remained higher for 24 h. Thus, glycitin treatment produces anti-aging effects including increased total collagen in the culture media, decreased elastase, and decreased ß-galactosidase. Together, these results indicate that glycitin stimulates TGF-ß secretion, and the subsequent autocrine actions of TGF-ß induce proliferation of fibroblasts, ultimately protecting skin cells from aging and wrinkling.
Asunto(s)
Fibroblastos/efectos de los fármacos , Isoflavonas/farmacología , Factor de Crecimiento Transformador beta/metabolismo , Movimiento Celular/efectos de los fármacos , Proliferación Celular/efectos de los fármacos , Células Cultivadas , Niño , Colágeno Tipo I/metabolismo , Colágeno Tipo III/metabolismo , Fibroblastos/citología , Fibronectinas/metabolismo , Humanos , Masculino , Metaloproteinasa 1 de la Matriz/metabolismo , Fosforilación , ARN Mensajero/metabolismo , Transducción de Señal/efectos de los fármacos , Proteína Smad2/metabolismo , Proteína smad3/metabolismo , Glycine max/química , beta-Galactosidasa/metabolismoRESUMEN
Metastasis is the leading cause of death in patients with breast, lung, and head and neck cancers. However, the molecular mechanisms underlying metastases in these cancers remain unclear. We found that the p90 ribosomal S6 kinase 2 (RSK2)-cAMP response element-binding protein (CREB) pathway is commonly activated in diverse metastatic human cancer cells, leading to up-regulation of a CREB transcription target Fascin-1. We also observed that the protein expression patterns of RSK2 and Fascin-1 correlate in primary human tumor tissue samples from head and neck squamous cell carcinoma patients. Moreover, knockdown of RSK2 disrupts filopodia formation and bundling in highly invasive cancer cells, leading to attenuated cancer cell invasion in vitro and tumor metastasis in vivo, whereas expression of Fascin-1 significantly rescues these phenotypes. Furthermore, targeting RSK2 with the small molecule RSK inhibitor FMK-MEA effectively attenuated the invasive and metastatic potential of cancer cells in vitro and in vivo, respectively. Taken together, our findings for the first time link RSK2-CREB signaling to filopodia formation and bundling through the up-regulation of Fascin-1, providing a proinvasive and prometastatic advantage to human cancers. Therefore, protein effectors of the RSK2-CREB-Fascin-1 pathway represent promising biomarkers and therapeutic targets in the clinical prognosis and treatment of metastatic human cancers.
Asunto(s)
Biomarcadores de Tumor/metabolismo , Proteína de Unión a CREB/metabolismo , Proteínas Portadoras/biosíntesis , Regulación Neoplásica de la Expresión Génica , Proteínas de Microfilamentos/biosíntesis , Proteínas de Neoplasias/metabolismo , Neoplasias/metabolismo , Proteínas Quinasas S6 Ribosómicas 90-kDa/metabolismo , Transducción de Señal , Animales , Biomarcadores de Tumor/genética , Proteína de Unión a CREB/genética , Proteínas Portadoras/genética , Línea Celular Tumoral , Técnicas de Silenciamiento del Gen , Humanos , Ratones , Ratones Desnudos , Proteínas de Microfilamentos/genética , Metástasis de la Neoplasia , Proteínas de Neoplasias/genética , Neoplasias/genética , Neoplasias/patología , Seudópodos/genética , Seudópodos/metabolismo , Seudópodos/patología , Proteínas Quinasas S6 Ribosómicas 90-kDa/genética , Regulación hacia Arriba/genéticaRESUMEN
The dengue virus (DENV) is a mosquito-borne pathogen responsible for an estimated 100 million human infections annually. The viral genome encodes a two-component trypsin-like protease that contains the cofactor region from the nonstructural protein NS2B and the protease domain from NS3 (NS3pro). The NS2B-NS3pro complex plays a crucial role in viral maturation and has been identified as a potential drug target. Using a DENV protease construct containing NS2B covalently linked to NS3pro via a Gly4-Ser-Gly4 linker ("linked protease"), previous x-ray crystal structures show that the C-terminal fragment of NS2B is remote from NS3pro and exists in an open state in the absence of an inhibitor; however, in the presence of an inhibitor, NS2B complexes with NS3pro to form a closed state. This linked enzyme produced NMR spectra with severe signal overlap and line broadening. To obtain a protease construct with a resolved NMR spectrum, we expressed and purified an unlinked protease complex containing a 50-residue segment of the NS2B cofactor region and NS3pro without the glycine linker using a coexpression system. This unlinked protease complex was catalytically active at neutral pH in the absence of glycerol and produced dispersed cross-peaks in a (1)H-(15)N heteronuclear single quantum correlation spectrum that enabled us to conduct backbone assignments using conventional techniques. In addition, titration with an active-site peptide aldehyde inhibitor and paramagnetic relaxation enhancement studies demonstrated that the unlinked DENV protease exists predominantly in a closed conformation in solution. This protease complex can serve as a useful tool for drug discovery against DENV.
Asunto(s)
Virus del Dengue/enzimología , Complejos Multienzimáticos/química , Proteínas no Estructurales Virales/química , Cristalografía por Rayos X , Virus del Dengue/genética , Humanos , Espectroscopía de Resonancia Magnética , Complejos Multienzimáticos/genética , Resonancia Magnética Nuclear Biomolecular , Estructura Cuaternaria de Proteína , Estructura Secundaria de Proteína , ARN Helicasas/química , ARN Helicasas/genética , Serina Endopeptidasas/química , Serina Endopeptidasas/genética , Proteínas no Estructurales Virales/genéticaRESUMEN
Wound healing plays an important role in protecting the human body from external infection. Cell migration and proliferation of keratinocytes and dermal fibroblasts are essential for proper wound healing. Recently, several studies have demonstrated that secondary compounds produced in plants could affect skin cells migration and proliferation. In this study, we identified a novel compound DK223 ([1E,2E-1,2-bis(6-methoxy-2H-chromen-3-yl)methylene]hydrazine) that concomitantly induced human keratinocyte migration and dermal fibroblast proliferation. We evaluated the regulation of epithelial and mesenchymal protein markers, such as E-cadherin and Vimentin, in human keratinocytes, as well as extracellular matrix (ECM) secretion and metalloproteinase families in dermal fibroblasts. DK223 upregulated keratinocyte migration and significantly increased the epithelial marker E-cadherin in a time-dependent manner. We also found that reactive oxygen species (ROS) increased significantly in keratinocytes after 2 h of DK223 exposure, returning to normal levels after 24 h, which indicated that DK223 had an early shock effect on ROS production. DK223 also stimulated fibroblast proliferation, and induced significant secretion of ECM proteins, such as collagen I, III, and fibronectin. In dermal fibroblasts, DK223 treatment induced TGF-ß1, which is involved in a signaling pathway that mediates proliferation. In conclusion, DK223 simultaneously induced both keratinocyte migration via ROS production and fibroblast proliferation via TGF-ß1 induction.