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Light-emitting diodes (LED)-derived lights have been widely used as a medical treatment in photobiomodulation (PBM). However, the PBM effects in ophthalmology are less well investigated. Herein, we explored the effect of LED-generated light on the tight-junction (TJ) formation in human corneal epithelial cells (HCEs). The HCEs were separately exposed to monochromatic LEDs at wavelengths of 365 nm (UVA), 420 nm (violet), 470 nm (blue), 530 nm (green), 590 nm (amber), 660 nm (deep red), and 740 nm (far red) at 10 J/cm2/day for 1 and 2 days. Long-term cultivation of HCEs without LED exposure for up to 14 days was established as a control. The effects of both LED wavelength and culture duration on cell morphology, cAMP-regulated proteins, TJ-associated proteins, and cell growth-associated proteins were also analyzed. Together with the increase in cell number during prolonged cultivation, cAMP, ZO-1, ZO-2, CLDN1, and CLDN4 all increased significantly during long-term cultivation without LED exposure. There was no difference in HCE viability after exposure to all monochromatic LEDs at an accumulated dose of 20 J/cm2. As determined by immunoblotting, UVA, violet, and blue light increased intracellular cAMP, ZO-1, ZO-2, CLDN1, and CLDN4 expression, respectively. UVA and violet, but not blue, light increased PKAreg-pS77 expression. However, none of the other treatments changed the expression of PKAcat-pT197, VASP-pS157, Bax, Bcl-2, or Bcl-xL. Immunofluorescence staining confirmed the formation of TJ structures. The expressions of ZO-1, ZO-2, CLDN1, and CLDN4 as well as TJ structures 2 days following UVA, violet, and blue exposure were similar to those of control cells after 9 days of cultivation. We conclude that short-wavelength LEDs at non-lethal exposure intensities accelerated the formation of TJ structure in HCEs via a cAMP-dependent regulatory cascade.
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Células Epiteliales , Uniones Estrechas , Humanos , Proteína de la Zonula Occludens-1/metabolismo , Uniones Estrechas/metabolismo , Células Epiteliales/metabolismo , Técnicas de Cultivo de CélulaRESUMEN
Objective: This study aims to investigate the variations in fatigue and sleep disturbances among female patients with advanced lung cancer (ALC) and advanced breast cancer (ABC) during chemotherapy. Methods: A total of 36 female patients with ALC and 36 with ABC, all of whom had completed their first cycle of chemotherapy, were included. Fatigue was assessed using the General Fatigue Scale (GFS), and sleep disturbances were evaluated using the Pittsburgh Sleep Quality Index (PSQI) at designated time points throughout the chemotherapy process. Results: Linear regression analysis indicated that variables such as age, education level, employment status, cancer type, clinical stage, and symptom distress had no significant correlation with either fatigue or sleep disturbances. The GFS significantly discriminated fatigue among the ALC, ABC, and combined groups, while the PSQI demonstrated a significant distinction in sleep disturbance only within the ALC and combined groups. Conclusions: In summary, when considering the findings of both assessments in this study, the GFS score exhibited greater sensitivity in detecting fatigue than the PSQI score did for identifying sleep disturbances in advanced cancer patients undergoing chemotherapy.
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INTRODUCTION: Previous studies have demonstrated varying sensitivity and specificity of computer-interpreted electrocardiography (CIE) in identifying ST-segment elevation myocardial infarction (STEMI). This study aims to evaluate the accuracy of contemporary computer software in recognizing electrocardiography (ECG) signs characteristic of STEMI compared to emergency physician overread in clinical practice. MATERIAL AND METHODS: In this retrospective observational single-center study, we reviewed the records of patients in the emergency department (ED) who underwent ECGs and troponin tests. Both the Philips DXL 16-Lead ECG. Algorithm and on-duty emergency physicians interpreted each standard 12lead ECG. The sensitivity and specificity of computer interpretation and physician overread ECGs for the definite diagnosis of STEMI were calculated and compared. RESULTS: Among the 9340 patients included in the final analysis, 133 were definitively diagnosed with STEMI. When "computer-reported infarct or injury" was used as the indicator, the sensitivity was 87.2% (95% CI 80.3% to 92.4%) and the specificity was 86.2% (95% CI 85.5% to 86.9%). When "physician-overread STEMI" was used as the indicator, the sensitivity was 88.0% (95% CI 81.2% to 93.0%) and the specificity was 99.9% (95% CI 99.8% to 99.9%). The area under the receiver operating characteristic curve for physician-overread STEMI and computer-reported infarct or injury were 0.939 (95% CI 0.907 to 0.972) and 0.867 (95% CI 0.834 to 0.900), respectively. CONCLUSIONS: This study reveals that while the sensitivity of the computer in recognizing ECG signs of STEMI is similar to that of physicians, physician overread of ECGs is more specific and, therefore, more accurate than CIE.
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Servicios Médicos de Urgencia , Médicos , Infarto del Miocardio con Elevación del ST , Humanos , Infarto del Miocardio con Elevación del ST/diagnóstico , Estudios Retrospectivos , Electrocardiografía , ComputadoresRESUMEN
PURPOSE: Cardiovascular disease (CVD) is a major worldwide health burden. As the risk factors of CVD, hypertension, and hyperlipidemia are most mentioned. Early stage hypertension in the population with dyslipidemia is an important public health hazard. This study was the application of data-driven machine learning (ML), demonstrating complex relationships between risk factors and outcomes and promising predictive performance with vast amounts of medical data, aimed to investigate the association between dyslipidemia and the incidence of early stage hypertension in a large cohort with normal blood pressure at baseline. METHODS: This study analyzed annual health screening data for 71,108 people from 2005 to 2017, including data for 27 risk-related indicators, sourced from the MJ Group, a major health screening center in Taiwan. We used five machine learning (ML) methods-stochastic gradient boosting (SGB), multivariate adaptive regression splines (MARS), least absolute shrinkage and selection operator regression (Lasso), ridge regression (Ridge), and gradient boosting with categorical features support (CatBoost)-to develop a multi-stage ML algorithm-based prediction scheme and then evaluate important risk factors at the early stage of hypertension, especially for groups with high-density lipoprotein cholesterol (HDL-C) and low-density lipoprotein cholesterol (LDL-C) levels within or out of the reference range. RESULTS: Age, body mass index, waist circumference, waist-to-hip ratio, fasting plasma glucose, and C-reactive protein (CRP) were associated with hypertension. The hemoglobin level was also a positive contributor to blood pressure elevation and it appeared among the top three important risk factors in all LDL-C/HDL-C groups; therefore, these variables may be important in affecting blood pressure in the early stage of hypertension. A residual contribution to blood pressure elevation was found in groups with increased LDL-C. This suggests that LDL-C levels are associated with CPR levels, and that the LDL-C level may be an important factor for predicting the development of hypertension. CONCLUSION: The five prediction models provided similar classifications of risk factors. The results of this study show that an increase in LDL-C is more important than the start of a drop in HDL-C in health screening of sub-healthy adults. The findings of this study should be of value to health awareness raising about hypertension and further discussion and follow-up research.
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Corneal endothelial cell (CEC) dysfunction causes corneal edema and severe visual impairment that require transplantation to restore vision. To address the unmet need of organ shortage, descemetorhexis without endothelial keratoplasty has been specifically employed to treat early stage Fuchs endothelial corneal dystrophy, which is pathophysiologically related to oxidative stress and exhibits centrally located corneal guttae. After stripping off central Descemet's membrane, rho-associated protein kinase (ROCK) inhibitor has been found to facilitate CEC migration, an energy-demanding task, thereby achieving wound closure. However, the correlation between ROCK inhibition and the change in bioenergetic status of CECs remained to be elucidated. Through transcriptomic profiling, we found that the inhibition of ROCK activity by the selective inhibitor, ripasudil or Y27632, promoted enrichment of oxidative phosphorylation (OXPHOS) gene set in bovine CECs (BCECs). Functional analysis revealed that ripasudil, a clinically approved anti-glaucoma agent, enhanced mitochondrial respiration, increased spare respiratory capacity, and induced overexpression of electron transport chain components through upregulation of AMP-activated protein kinase (AMPK) pathway. Accelerated BCEC migration and in vitro wound healing by ripasudil were diminished by OXPHOS and AMPK inhibition, but not by glycolysis inhibition. Correspondingly, lamellipodial protrusion and actin assembly that were augmented by ripasudil became reduced with additional OXPHOS or AMPK inhibition. These results indicate that ROCK inhibition induces metabolic reprogramming toward OXPHOS to support migration of CECs.
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Endotelio Corneal , Distrofia Endotelial de Fuchs , Proteínas Quinasas Activadas por AMP/metabolismo , Animales , Bovinos , Células Endoteliales/metabolismo , Endotelio Corneal/metabolismo , Distrofia Endotelial de Fuchs/genética , Distrofia Endotelial de Fuchs/metabolismo , Distrofia Endotelial de Fuchs/cirugía , Fosforilación Oxidativa , Quinasas Asociadas a rho/metabolismoRESUMEN
Brainstem encephalitis, a manifestation of severe enterovirus 71 (EV71) infection, is an acute excessive inflammatory response. The mechanisms underlying its development remain poorly understood. Usually neurotropic viruses trigger acute host immune response by engaging cell surface or intracellular receptors. Here, we show that EV71 engagement with intracellular receptor TLR9 elicits IL-12p40-iNOS signaling causing encephalitis in mice. We identified IL-12p40 to be the only prominent cytokine-induced at the early infection stage in the brainstem of mice subjected to a lethal dose of EV71. The upregulated IL-12p40 proteins were expressed in glial cells but not neuronal cells. To better understand the role of IL-12p40 in severe EV71 infection, we treated the EV71-infected mice with an antibody against IL-12p40 and found the mortality rate, brainstem inflammation, and gliosis to be markedly reduced, suggesting that the acute IL-12p40 response plays a critical role in the pathogenesis of brainstem encephalitis. Mechanistically, intracellular TLR9 was found essential to the activation of the IL-12p40 response. Blocking TLR9 signaling with CpG-ODN antagonist ameliorated IL-12p40 response, brainstem inflammation, and limb paralysis in mice with EV71-induced encephalitis. We further found the glial IL-12p40 response might damage neurons by inducing excess production of neurotoxic NO by iNOS. Overall, EV71 engagement with intracellular TLR9 was found to elicit a neurotoxic glial response via IL12p40-iNOS signaling contributing to the neurological manifestation of EV71 infection. This pathway could potentially be targeted for the treatment of brainstem encephalitis.
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Encefalitis , Enterovirus Humano A , Infecciones por Enterovirus , Subunidad p40 de la Interleucina-12 , Receptor Toll-Like 9 , Animales , Encefalitis/inmunología , Encefalitis/virología , Infecciones por Enterovirus/inmunología , Inflamación , Subunidad p40 de la Interleucina-12/metabolismo , Ratones , Óxido Nítrico Sintasa de Tipo II/metabolismo , Receptor Toll-Like 9/metabolismoRESUMEN
Transgenic proteins can be routinely expressed in various mammalian cell types via different transgenic systems, but the efficiency of transgene expression is constrained by the complex interplay among factors such as the temporal consistency of expression and compatibility with specific cell types, including ocular cells. Here, we report a more efficient way to express an enhanced green fluorescent protein (EGFP) in human corneal fibroblasts, corneal epithelial cells, and conjunctival epithelial cells through a lentiviral expression system. The relative transducing unit criterion for EGFP-expressing pseudovirions was first determined in HEK-293T cells. Homogeneous populations of EGFP-positive and EGFP-negative cells could be isolated by cell sorting. The half-maximal inhibitory concentration (IC50) value for puromycin was calculated according to viability curves for each cell type. The results revealed that cell types differed with respect to EGFP expression efficiency after transduction with the same amount of EGFP-encoding pseudovirions. Using a cell sorter, the homogeneity of EGFP-positive cells reached >95%. In the initial sorting stage, however, the efficiency of EGFP expression in the sorted cells was noticeably reduced after two rounds of sequential culture, but repeated sorting for up to four rounds yielded homogeneous EGFP-positive human corneal fibroblasts that could be maintained in continuous culture in vitro. The sorted EGFP-positive cells retained their proper morphology and cell type-specific protein expression patterns. Puromycin resistance was found to depend on cell type, indicating that the IC50 for puromycin must be determined for each cell type to ensure the isolation of homogeneous EGFP-positive cells. Taken together, repeated cell sorting is an efficient means of obtaining homogeneous populations of ocular cells expressing a transgenic protein during continuous culture without the potential confounding effects of antibiotics.
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Mamíferos , Animales , Animales Modificados Genéticamente , Separación Celular , Citometría de Flujo , Proteínas Fluorescentes Verdes/metabolismo , Humanos , Mamíferos/metabolismo , Puromicina/farmacología , TransgenesRESUMEN
Moxifloxacin (MOX) suppresses cell movement in human corneal fibroblasts (HCFs). Zonula occludens-1 (ZO-1) is localized to the leading edge of migrating HCFs. This study explored the role of ZO-1 in MOX-suppressed cell migration in HCFs. A single-cell trajectory analysis revealed that MOX negatively regulated the migratory properties of HCFs including migration distance, migration velocity, and directionality (P < 0.001, P < 0.001, and P = 0.018, respectively). MOX increased endogenous ZO-1 in HCFs in a concentration-dependent manner (P = 0.083, P = 0.005, and P = 0.001 at 10, 50, and 100 µg/ml, respectively), but decreased the phosphorylation of endogenous ZO-1 at serines, threonines, and tyrosines. In contrast, MOX did not alter the expression of protein kinase C epsilon (PKCε), Rac-1, Cdc42, and MRCKß. However, MOX did also reduce the phosphorylation level of PKCε at serines and threonines (P < 0.001 at 100 µg/ml). In addition, MOX increased the phosphorylation level of Rac-1 in a concentration-dependent manner (P < 0.001 at 100 µg/ml). Compared with the mock cells, the directionality of cell movement increased significantly in ZO-1-expressing HCFs (P = 0.012) and decreased significantly in ZO-1-silenced HCFs (P = 0.002). The directionality did not change significantly in Rac-1-silenced HCFs. ZO-1-expressing HCFs moved faster than mock cells. PKCε, Cdc42, Rac-1, and phosphorylated Rac-1 were decreased in ZO-1-overexpressing HCFs, but increased in ZO-1-silenced HCFs. Finally, silencing ZO-1 blocked MOX hyperactivation of Rac-1. These suggest that MOX might trigger random migration in human corneal stromal cells through PKCε-modulated ZO-1 inactivation and Rac-1 hyperactivation.
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Córnea/citología , Córnea/metabolismo , Fibroblastos/metabolismo , Moxifloxacino/farmacología , Proteína Quinasa C-epsilon/metabolismo , Transducción de Señal/efectos de los fármacos , Proteína de la Zonula Occludens-1/metabolismo , Movimiento Celular/efectos de los fármacos , Movimiento Celular/genética , Proliferación Celular/efectos de los fármacos , Proliferación Celular/genética , Células Cultivadas , Córnea/efectos de los fármacos , Fibroblastos/efectos de los fármacos , Humanos , Fosforilación/efectos de los fármacos , Proteína de la Zonula Occludens-1/genética , Proteína de Unión al GTP rac1/genética , Proteína de Unión al GTP rac1/metabolismoRESUMEN
The fungus Antrodia cinnamomea has been used as a folk medicine for various diseases, especially cancer. When A. cinnamomea is cultured on the original host, an endangered woody plant Cinnamomum kanehirai Hayata, the fungus produces more active ingredients, but its growth is slow. Here, C. kanehirai leaf ethanol extract (KLEE) was used as a substitute for C. kanehirai wood to culture A. cinnamomea on solid medium to shorten the culture period and produce active metabolites en masse. The antioxidant activities of methanol extracts from A. cinnamomea cultured on KLEE (MEAC-KLEE) were evaluated by 2,2-diphenyl-1-picrylhydrazyl (DPPH) radical-scavenging effect, reducing power, and ferrous ion-chelating effect, and the effective concentration (EC50) values were 0.27, 0.74, and 0.37 mg mL-1, respectively. MEAC-KLEE exhibited specific anti-proliferative activity against a non-small-cell lung cancer cell line (A549) by Annexin V assay. A secondary metabolite (2,4-dimethoxy-6-methylbenzene-1,3-diol, DMMB) present in the extract (MEAC-KLEE) was purified by high-performance liquid chromatography (HPLC) and identified by nuclear magnetic resonance (NMR) spectra. DMMB exhibited moderate antioxidant activity against DPPH radicals and reducing power, with EC50 values of 12.97 and 25.59 µg mL-1, respectively, and also induced apoptosis in A549 cells. Our results provide valuable insight into the development of DMMB for nutraceutical biotechnology.
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Mitomycin C (MMC) is often used to prevent postoperative corneal haze and subconjunctival fibrosis in ocular surgery. It also affects the motility and viability of the residual ocular cells, including corneal stromal cells. Extracellular matrix metalloproteinase-9 (MMP-9) contributes to the promotion of cell movement in macrophage and cancer cells, but the intracellular role of MMP-9 remained unclear. Herein, we illustrated the novel role of intracellular MMP-9 in MMC-suppressed cell migration using isolated human corneal fibroblasts (HCFs). In HCFs, MMC enhanced intracellular MMP-9 at transcriptional and protein levels. Using co-immunoprecipitation analysis, we confirmed that MMC enhanced the association between intracellular MMP-9 and inactive FAK/paxillin (PXN) complexes, i.e. PXN without phospho-tyrosine 118 (pY118) and FAK without phospho-tyrosine 397 (pY397). To verify the role of intracellular MMP-9 in migration, its gene was directly isolated from HCFs and highly expressed in HCFs by a lentivirus-based pseudovirus system with encephalomyocarditis virus (EMCV)-driven enhanced green fluorescent protein (GFP) as the MMP-9-IG-versus IG-expressing cells. Compared with the IG-expressing cells, higher intracellular MMP-9 expression in the MMP-9-IG-expressing HCFs proliferated and migrated more slowly. Phosphorylation of FAK at Y397 and PXN at both Y31 and Y118 were significantly less in the MMP-9-IG-expressing HCFs. These suggested that MMC-upregulated intracellular MMP-9 clutched inactive FAK/PXN complexes at focal adhesion sites to form a new "inactive" trimer, prohibited FAK/PXN complexes phosphorylation and retarded corneal fibroblast migration.
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Movimiento Celular/efectos de los fármacos , Córnea/citología , Fibroblastos/efectos de los fármacos , Metaloproteinasa 9 de la Matriz/metabolismo , Mitomicina/farmacología , Proliferación Celular/efectos de los fármacos , Células Cultivadas , Fibroblastos/metabolismo , Fibroblastos/fisiología , Humanos , Metaloproteinasa 9 de la Matriz/genéticaRESUMEN
PURPOSE: To evaluate the safety and feasibility of patent blue (PB) as the vital dye in Descemet membrane endothelial keratoplasty (DMEK). METHODS: Bovine corneal endothelial cells were incubated with different concentrations (0.02%-2.5%) of PB. The cell viability, which was assessed by Cell Counting Kit-8 assay, was compared with that of untreated control and 0.06% to 0.4% trypan blue. The dyes were also used for graft preparation and implantation in the porcine eye model to evaluate stain quality, dye retention, and the feasibility of using PB in DMEK surgery. RESULTS: No obvious increase in cytotoxicity was detected for 0.06% to 0.4% trypan blue and PB at concentrations up to 1.0%, but the cell viability after incubating with 1.5% to 2.5% PB was significantly reduced. PB at 0.5% to 1.0% generated good staining quality that can be used to facilitate graft implantation. Although the staining quality of 0.5% to 1.0% PB faded to an intermediate level after a 30-minute wash in phosphate-buffered saline, dye retention persisted for up to 24 hours. CONCLUSIONS: PB at 0.5% to 1.0% is biocompatible and can stain the graft sufficiently, making it an alternative for DMEK surgery.
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Colorantes/toxicidad , Pérdida de Celulas Endoteliales de la Córnea/patología , Queratoplastia Endotelial de la Lámina Limitante Posterior/métodos , Células Endoteliales/efectos de los fármacos , Endotelio Corneal/efectos de los fármacos , Colorantes de Rosanilina/toxicidad , Coloración y Etiquetado/métodos , Animales , Bovinos , Supervivencia Celular/efectos de los fármacos , Células Cultivadas , Porcinos , Azul de Tripano/toxicidadRESUMEN
Tartrate-resistant acid phosphatase 5a (TRACP5a) is mainly secreted by activated macrophages in chronic inflammation. Serum TRACP5a is associated with symptom distress in lung cancer patients during chemotherapy. Therefore, this study aimed to investigate whether chemotherapy drugs modulate TRACP5a as an inducible marker for symptom distress in lung cancer patients during chemotherapy. In clinical analysis, lung cancer participants completely received the six-cycle chemotherapy process (nâ¯=â¯42). Clinical determinations for TRACP5a, C-reactive protein (CRP), interleukin-6 (IL-6), white blood cells, monocytes, and hemoglobin were analyzed at six time points: BL, C1d8, C2d1, C4d1, C4d8, and Ed28. Meanwhile, five questionnaires for fatigue, sleep disturbance, pain, depression, and confusion were finished before drug treatment. For monocyte-to-macrophage differentiation, THP-1 cells were treated with phorbol 12-myristate 13-acetate (PMA). TRACP5a secretion in THP-1 cells was determined at the following days up to 6 days after 1-day incubation of chemotherapy drugs by dot blotting. Clinical analysis revealed that TRACP5a significantly increased at C1d8 and C4d8, but dropped at C2d1 and Ed28. CRP and IL-6 displayed a broad-range variation, resulting in no significant difference among the assessment time points. In contrast, monocytes decreased at C1d8 and C4d8, but rose again at C2d1 and Ed28. In symptom distress, the changes only in fatigue and sleep disturbance were positively associated with the trend in TRACP5a. In PMA-treated THP-1 cells, TRACP5a significantly increased after stimulation with gemcitabine and paclitaxel. Taken together, induction of TRACP5a by chemotherapy drugs might be generated from monocyte-differentiated macrophages, further causing clinical symptom distress in lung cancer patients.
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Antineoplásicos/efectos adversos , Carcinoma de Pulmón de Células no Pequeñas/tratamiento farmacológico , Neoplasias Pulmonares/tratamiento farmacológico , Macrófagos/metabolismo , Fosfatasa Ácida Tartratorresistente/metabolismo , Anciano , Antineoplásicos/uso terapéutico , Biomarcadores/metabolismo , Proteína C-Reactiva/metabolismo , Carcinoma de Pulmón de Células no Pequeñas/fisiopatología , Carcinoma de Pulmón de Células no Pequeñas/psicología , Diferenciación Celular , Confusión/inducido químicamente , Confusión/metabolismo , Depresión/inducido químicamente , Depresión/metabolismo , Fatiga/inducido químicamente , Fatiga/metabolismo , Femenino , Hemoglobinas/efectos de los fármacos , Humanos , Interleucina-6/metabolismo , Leucocitos/efectos de los fármacos , Neoplasias Pulmonares/fisiopatología , Neoplasias Pulmonares/psicología , Macrófagos/efectos de los fármacos , Masculino , Persona de Mediana Edad , Monocitos/efectos de los fármacos , Trastornos del Sueño-Vigilia/inducido químicamente , Trastornos del Sueño-Vigilia/metabolismo , Evaluación de Síntomas , Células THP-1 , Fosfatasa Ácida Tartratorresistente/sangre , Acetato de Tetradecanoilforbol/uso terapéuticoRESUMEN
Purpose: To explore the variability in interleukin-12 (IL-12) secretion in human corneal fibroblasts (HCFs), the effects of IL-12 on HCF viability and migration, and the effects of recombinant human transforming growth factor-ß1 (rhTGF-ß1) and moxifloxacin (MOX) on IL-12 expression. Methods: IL-12 secretion in 27 primary HCFs incubated with rhTGF-ß1 and/or MOX was analyzed using immunoblotting. Viability and migration were assessed using WST-1 assay and culture inserts, respectively. The effects of IL-12 on HCFs were verified via adding recombinant human IL-12 (rhIL-12) or silencing IL-12. The effects of rhTGF-ß1 and MOX on IL-12 secretion were also verified. Results: There were 25.9% HCFs that did not secrete IL-12 and 74.1% HCFs that secreted IL-12 were divided into low- (25.9%), medium- (37.0%), and high-secretion (11.1%) subgroups. After 3-day incubation, rhTGF-ß1 stimulated IL-12 by 1.90 ± 0.84 folds (P = 0.026), with no difference in folds of induction among subgroups (P = 0.870). MOX suppressed IL-12 secretion concentration-dependently, more noticeably in the presence of rhTGF-ß1. Exogenous rhIL-12 enhanced HCF proliferation (1.29 folds, P < 0.001) and migration. In contrast, rhTGF-ß1 only enhanced proliferation (1.24 folds, P < 0.001). In IL-12-silenced HCFs, rhTGF-ß1 failed to enhance IL-12 secretion and resulted in reduced cell proliferation and migration. Conclusions: IL-12 enhanced HCF proliferation and migration. TGF-ß1 per se enhanced HCF proliferation but not migration. However, it promoted migration indirectly through upregulating IL-12 secretion. MOX-inhibited HCF migration via suppressing IL-12 secretion. The interindividual variation in IL-12 secretion might contribute to disparity in HCF proliferation and migration after corneal wounding among individuals.
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Córnea/metabolismo , Fluoroquinolonas/farmacología , Interleucina-12/metabolismo , Factor de Crecimiento Transformador beta1/farmacología , Adulto , Anciano , Antibacterianos/farmacología , Movimiento Celular , Proliferación Celular , Células Cultivadas , Córnea/citología , Córnea/efectos de los fármacos , Fibroblastos/citología , Fibroblastos/efectos de los fármacos , Fibroblastos/metabolismo , Humanos , Immunoblotting , Interleucina-12/genética , Persona de Mediana Edad , Moxifloxacino , ARN/genética , Reacción en Cadena en Tiempo Real de la Polimerasa , Proteínas Recombinantes/farmacología , Adulto JovenRESUMEN
BACKGROUND: Symptom distress often occurs in lung cancer patients undergoing chemotherapy. However, a biomarker has not been identified to reflect the severity of their symptom distress. OBJECTIVE: The aim of this study was to investigate the relationship between symptom distress and serum inflammatory biomarkers in lung cancer patients undergoing chemotherapy. METHODS: A longitudinal, repeated-measures design was used to assess subjective symptoms (fatigue, sleep disturbance, pain, depression, and confusion), serum biomarkers (tartrate-resistant acid phosphatase 5a [TRACP5a], interleukin 6 [IL-6], IL-8, and C-reactive protein), and white blood cells in 62 lung cancer patients recruited from a single medical center at 3 time points: T1 was the baseline, T2 was the eighth day after the first chemotherapy cycle, and T3 was prior to the second cycle. Symptom distress was measured individually by 5 questionnaires (General Fatigue Scale, Pittsburgh Sleep Quality Index, Brief Pain Inventory, Profile of Mood States-Depressive, and Confusion). RESULTS: The trend of TRACP5a was positively correlated to the trend of the patients' symptom distress. However, the trends of IL-6 and IL-8 did not correlate. CONCLUSIONS: Serum TRACP5a was associated with symptom distress in lung cancer patients. Therefore, TRACP5a might be a potential biomarker to assess symptom distress of lung cancer patients undergoing chemotherapy. IMPLICATIONS FOR PRACTICE: Oncology nurses may be able to apply TRACP5a expression to predict or monitor multiple distress symptoms in lung cancer patients undergoing chemotherapy. Furthermore, nurses can use these study findings to better understand the patients who need more attention to improve their quality of life.
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Proteína C-Reactiva/análisis , Interleucina-6/sangre , Interleucina-8/sangre , Neoplasias Pulmonares/tratamiento farmacológico , Neoplasias Pulmonares/psicología , Estrés Psicológico/sangre , Fosfatasa Ácida Tartratorresistente/sangre , Anciano , Biomarcadores/sangre , Confusión/sangre , Confusión/etiología , Depresión/sangre , Depresión/etiología , Fatiga/sangre , Fatiga/etiología , Femenino , Humanos , Estudios Longitudinales , Masculino , Persona de Mediana Edad , Dolor/sangre , Dolor/etiología , Trastornos del Sueño-Vigilia/sangre , Trastornos del Sueño-Vigilia/etiología , Encuestas y CuestionariosRESUMEN
Topical fluoroquinolones are widely used to prevent ocular infections after ophthalmic surgery. However, they have been shown to affect the corneal cell motility, whose mechanism remains indefinite. The purpose of this study was to investigate how fluoroquinolones affect corneal stromal cell motility. Human corneal fibroblasts (HCFs) were incubated in ciprofloxacin (CIP), levofloxacin (LEV), or moxifloxacin (MOX) at 0, 10, 50, and 100 µg/ml for up to 3 days. Effect of CIP, LEV, or MOX on HCF migration was monitored using migration assay. HCF viability was determined by WST-1 assay. Expression of focal adhesion kinase (FAK), paxillin (PXN), and their phosphorylated forms were analyzed by immunoblotting. Binding affinity between FAK and PXN was determined by co-immunoprecipitation. Our results revealed that CIP and MOX, but not LEV, noticeably retarded HCF migration. HCF proliferation was significantly reduced by CIP (38.2%), LEV (29.5%), and MOX (21.3%), respectively (p = 0.002). CIP and MOX suppressed the phosphorylation of PXN at tyrosines (10.2 ± 4.3%, p < 0.001; 11.7 ± 2.4%, p < 0.001, respectively), including tyrosine 118 (33.3 ± 5.2%, p < 0.001; 34.0 ± 4.4%, p < 0.001, respectively). CIP and MOX diminished the binding affinity between FAK and PXN (8.2 ± 1.8%, p < 0.001; 9.0 ± 4.5%, p < 0.001, respectively). Nevertheless, tyrosine dephosphorylation and FAK dissociation of PXN were not found in LEV-treated HCFs. None of these fluoroquinolones affect phosphorylation of FAK-Y397. We conclude that CIP and MOX, but not LEV, might delay corneal fibroblast migration via interfering with recruitment of PXN to focal adhesions and dephosphorylation of PXN at the tyrosines.
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Úlcera de la Córnea/tratamiento farmacológico , Epitelio Corneal/metabolismo , Fluoroquinolonas/administración & dosificación , Proteína-Tirosina Quinasas de Adhesión Focal/biosíntesis , Paxillin/biosíntesis , Adulto , Anciano , Antibacterianos/administración & dosificación , Recuento de Células , Movimiento Celular/efectos de los fármacos , Úlcera de la Córnea/metabolismo , Úlcera de la Córnea/patología , Epitelio Corneal/efectos de los fármacos , Epitelio Corneal/patología , Femenino , Fibroblastos/efectos de los fármacos , Fibroblastos/metabolismo , Fibroblastos/patología , Proteína-Tirosina Quinasas de Adhesión Focal/efectos de los fármacos , Humanos , Immunoblotting , Masculino , Persona de Mediana Edad , Soluciones Oftálmicas , Paxillin/efectos de los fármacos , Adulto JovenRESUMEN
Intraoperative mitomycin C (MMC) is widely used to prevent pterygium recurrence and glaucoma filtering bleb failure, but it has been shown to induce corneal inflammation and cell death. Postoperative dexamethasone (DEX) is advocated to reduce MMC-related inflammation and cell death in corneal fibroblasts. Nevertheless, its long-term regulation mechanism in Tenon's capsule remains to be explored. The purpose of this study was to investigate how DEX modifies MMC's effects in human Tenon's capsule fibroblasts (HTFs). HTFs isolated from the pterygium surgical patients (n = 6) were treated with MMC at 0, 0.1, 0.2, and 0.4 mg/ml for 5 min and incubated in DEX at 10 µM for 0, 1, 2, and 3 days. Recombinant interleukin-8 (IL-8) was used to verify the effect of MMC-related IL-8 secretion. Cell proliferation of all the treated cells was analyzed by WST-1 assay. The amount of IL-8 secretion in HTFs was determined by enzyme-linked immunosorbent assay. Immunoblotting assay was used to analyze the expression of peroxisome-proliferator-activated receptor gamma (PPARγ) and B-cell lymphoma-extra large (Bcl-xL). Our results revealed that MMC significantly reduced the HTF cell proliferation rate. Additionally, MMC significantly upregulated IL-8 secretion in HTFs concentration-dependently. At 3 days post treatment (dpt), 5-min exposures to 0.1, 0.2, and 0.4 mg/ml MMC resulted in 1.4-fold (p = 0.012), 1.6-fold (p = 0.012), and 2.5-fold (p = 0.001) increases of IL-8 secretion. In contrast, DEX reversed the MMC-retarded cell proliferation rate (p = 0.036) and repressed MMC-related IL-8 secretion by 33.5% at 3 dpt (p = 0.003). Addition of recombinant IL-8 noticeably suppressed HTF cell proliferation in a concentration-dependent manner. DEX stimulated upregulation of both PPARγ and Bcl-xL at 1 dpt in normal HTFs and at 2 dpt in MMC-treated HTFs. PPARγ silencing reduced expression of PPARγ and Bcl-xL, but enhanced IL-8 secretion (p < 0.001). On the other hand, Bcl-xL silencing enhanced IL-8 secretion (p < 0.001), but did not affect PPARγ expression. These revealed that IL-8 secretion in HTFs is modulated by PPARγ-dependent Bcl-xL signaling. We conclude that DEX reversed the MMC-inhibited HTF cell proliferation via diminishing the MMC-induced IL-8 secretion, which resulted from a late-phase upregulation of the PPARγ and Bcl-xL. These long-term effects suggest a beneficial postoperative DEX treatment following intraoperative MMC application.
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Dexametasona/farmacología , Fibroblastos/metabolismo , Interleucina-8/metabolismo , Pterigion/tratamiento farmacológico , Cápsula de Tenon/metabolismo , Apoptosis , Western Blotting , Proliferación Celular/efectos de los fármacos , Células Cultivadas , Ensayo de Inmunoadsorción Enzimática , Fibroblastos/efectos de los fármacos , Fibroblastos/patología , Regulación de la Expresión Génica/efectos de los fármacos , Glucocorticoides/farmacología , Humanos , Interleucina-8/efectos de los fármacos , Mitomicina/farmacología , PPAR gamma/biosíntesis , PPAR gamma/genética , Pterigion/metabolismo , Pterigion/patología , ARN/genética , Cápsula de Tenon/patología , Proteína bcl-X/biosíntesis , Proteína bcl-X/genéticaRESUMEN
White spot syndrome virus (WSSV) is an enveloped, large dsDNA virus that mainly infects penaeid shrimp, causing serious damage to the shrimp aquaculture industry. Like other animal viruses, WSSV infection induces apoptosis. Although this occurs even in by-stander cells that are free of WSSV virions, apoptosis is generally regarded as a kind of antiviral immune response. To counter this response, WSSV has evolved several different strategies. From the presently available literature, we construct a model of how the host and virus both attempt to regulate apoptosis to their respective advantage. The basic sequence of events is as follows: first, when a WSSV infection occurs, cellular sensors detect the invading virus, and activate signaling pathways that lead to (1) the expression of pro-apoptosis proteins, including PmCasp (an effecter caspase), MjCaspase (an initiator caspase) and voltage-dependent anion channel (VDAC); and (2) mitochondrial changes, including the induction of mitochondrial membrane permeabilization and increased oxidative stress. These events initiate the apoptosis program. Meanwhile, WSSV begins to express its genes, including two anti-apoptosis proteins: AAP-1, which is a direct caspase inhibitor, and WSV222, which is an E3 ubiquitin ligase that blocks apoptosis through the ubiquitin-mediated degradation of shrimp TSL protein (an apoptosis inducer). WSSV also induces the expression of a shrimp anti-apoptosis protein, Pm-fortilin, which can act on Bax to inhibit mitochondria-triggered apoptosis. This is a life and death struggle because the virus needs to prevent apoptosis in order to replicate. If WSSV succeeds in replicating in sufficient numbers, this will result in the death of the infected penaeid shrimp host.
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Apoptosis/inmunología , Infecciones por Virus ADN/inmunología , Penaeidae/inmunología , Virus del Síndrome de la Mancha Blanca 1/inmunología , Animales , Infecciones por Virus ADN/patología , Infecciones por Virus ADN/virología , Penaeidae/virologíaRESUMEN
White spot syndrome virus (WSSV) is a serious shrimp pathogen that has spread globally to all major shrimp farming areas, causing enormous economic losses. Here we investigate the role of hermit crabs in transmitting WSSV to Penaeus monodon brooders used in hatcheries in Vietnam. WSSV-free brooders became PCR-positive for WSSV within 2 to 14 d, and the source of infection was traced to hermit crabs being used as live feed. Challenging hermit crabs with WSSV confirmed their susceptibility to infection, but they remained tolerant to disease even at virus loads equivalent to those causing acute disease in shrimp. As PCR screening also suggests that WSSV infection occurs commonly in hermit crab populations in both Vietnam and Taiwan, their use as live feed for shrimp brooders is not recommended.
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Alimentación Animal , Anomuros , Dieta , Penaeidae/virología , Virus del Síndrome de la Mancha Blanca 1/fisiología , Animales , Acuicultura , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Factores de RiesgoRESUMEN
PURPOSE: To investigate how mitomycin C (MMC) modulates corneal fibroblast migration and its molecular mechanisms in the wound healing process. METHODS: After treatment with 0 and 0.2 mg · mL(-1) MMC for 5 minutes, effect of MMC on cell migration of human corneal fibroblasts (HCFs) was examined with a cell migration assay. Both focal adhesion kinase (FAK) and paxillin (PXN) expressions in HCFs were analyzed by semiquantitative real-time PCR, immunoblotting, and immunofluorescence confocal microscopy. Using gene silencing or gene overexpression with lentiviral-based pseudovirion infection, the phosphorylation level of FAK, PXN, and mutated PXNs at tyrosine sites 31 (Y31F-EGFP) and 118 (Y118F-EGFP) were verified in HCFs. RESULTS: MMC retarded HCF migration at 1 and 2 days posttreatment (dpt). MMC reduced levels of FAK transcript and FAK protein, but increased both transcript and protein expression of PXN at 1 and 2 dpt. Furthermore, MMC upregulated FAK-pY397, which subsequently enhanced PXN-pY31 in a dose-dependent manner at 1 dpt. Concurrently, MMC downregulated PXN-pY118 at 1 dpt. However, MMC treatment resulted in dephosphorylation of FAK-pY397, PXN-pY31, and PXN-pY118 at 2 dpt. The FAK/PXN complex in MMC-treated HCFs was detected at focal adhesion sites more than at the leading edge at 1 and 2 dpt, contributing to retardation of HCF migration. Y118F-EGFP-expressing HCFs exhibited lower mobility than that of PXN-EGFP- or Y31F-EGFP-expressing HCFs. CONCLUSIONS: The sustained PXN-pY118 dephosphorylation resulted in steadfastness of an incompletely active FAK/PXN complex at focal adhesion sites and played a pivotal role in MMC-retarded HCF migration.
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Movimiento Celular/efectos de los fármacos , Fibroblastos/efectos de los fármacos , Mitomicina/farmacología , Inhibidores de la Síntesis del Ácido Nucleico/farmacología , Paxillin/metabolismo , Movimiento Celular/fisiología , Epitelio Corneal/citología , Epitelio Corneal/efectos de los fármacos , Epitelio Corneal/metabolismo , Fibroblastos/metabolismo , Proteína-Tirosina Quinasas de Adhesión Focal/metabolismo , Regulación de la Expresión Génica , Humanos , Immunoblotting , Fosforilación/efectos de los fármacos , Reacción en Cadena en Tiempo Real de la Polimerasa , Tirosina/metabolismo , Cicatrización de Heridas/efectos de los fármacosRESUMEN
The Warburg effect is an abnormal glycolysis response that is associated with cancer cells. Here we present evidence that metabolic changes resembling the Warburg effect are induced by a nonmammalian virus. When shrimp were infected with white spot syndrome virus (WSSV), changes were induced in several metabolic pathways related to the mitochondria. At the viral genome replication stage (12 h postinfection [hpi]), glucose consumption and plasma lactate concentration were both increased in WSSV-infected shrimp, and the key enzyme of the pentose phosphate pathway, glucose-6-phosphate dehydrogenase (G6PDH), showed increased activity. We also found that at 12 hpi there was no alteration in the ADP/ATP ratio and that oxidative stress was lower than that in uninfected controls. All of these results are characteristic of the Warburg effect as it is present in mammals. There was also a significant decrease in triglyceride concentration starting at 12 hpi. At the late stage of the infection cycle (24 hpi), hemocytes of WSSV-infected shrimp showed several changes associated with cell death. These included the induction of mitochondrial membrane permeabilization (MMP), increased oxidative stress, decreased glucose consumption, and disrupted energy production. A previous study showed that WSSV infection led to upregulation of the voltage-dependent anion channel (VDAC), which is known to be involved in both the Warburg effect and MMP. Here we show that double-stranded RNA (dsRNA) silencing of the VDAC reduces WSSV-induced mortality and virion copy number. For these results, we hypothesize a model depicting the metabolic changes in host cells at the early and late stages of WSSV infection.