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INTRODUCTION: Critical COVID-19 survivors have a high risk of respiratory sequelae. Therefore, we aimed to identify key factors associated with altered lung function and CT scan abnormalities at a follow-up visit in a cohort of critical COVID-19 survivors. METHODS: Multicenter ambispective observational study in 52 Spanish intensive care units. Up to 1327 PCR-confirmed critical COVID-19 patients had sociodemographic, anthropometric, comorbidity and lifestyle characteristics collected at hospital admission; clinical and biological parameters throughout hospital stay; and, lung function and CT scan at a follow-up visit. RESULTS: The median [p25-p75] time from discharge to follow-up was 3.57 [2.77-4.92] months. Median age was 60 [53-67] years, 27.8% women. The mean (SD) percentage of predicted diffusing lung capacity for carbon monoxide (DLCO) at follow-up was 72.02 (18.33)% predicted, with 66% of patients having DLCO<80% and 24% having DLCO<60%. CT scan showed persistent pulmonary infiltrates, fibrotic lesions, and emphysema in 33%, 25% and 6% of patients, respectively. Key variables associated with DLCO<60% were chronic lung disease (CLD) (OR: 1.86 (1.18-2.92)), duration of invasive mechanical ventilation (IMV) (OR: 1.56 (1.37-1.77)), age (OR [per-1-SD] (95%CI): 1.39 (1.18-1.63)), urea (OR: 1.16 (0.97-1.39)) and estimated glomerular filtration rate at ICU admission (OR: 0.88 (0.73-1.06)). Bacterial pneumonia (1.62 (1.11-2.35)) and duration of ventilation (NIMV (1.23 (1.06-1.42), IMV (1.21 (1.01-1.45)) and prone positioning (1.17 (0.98-1.39)) were associated with fibrotic lesions. CONCLUSION: Age and CLD, reflecting patients' baseline vulnerability, and markers of COVID-19 severity, such as duration of IMV and renal failure, were key factors associated with impaired DLCO and CT abnormalities.
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COVID-19 , Enfisema Pulmonar , Humanos , Femenino , Persona de Mediana Edad , Masculino , Enfermedad Crítica , Estudios de Seguimiento , COVID-19/complicaciones , Progresión de la Enfermedad , Pulmón/diagnóstico por imagenRESUMEN
BACKGROUND: The primary aim of our study was to investigate the association between intubation timing and hospital mortality in critically ill patients with coronavirus disease 2019 (COVID-19)-associated respiratory failure. We also analysed both the impact of such timing throughout the first four pandemic waves and the influence of prior noninvasive respiratory support on outcomes. METHODS: This is a secondary analysis of a multicentre, observational and prospective cohort study that included all consecutive patients undergoing invasive mechanical ventilation due to COVID-19 from across 58 Spanish intensive care units (ICUs) participating in the CIBERESUCICOVID project. The study period was between 29 February 2020 and 31 August 2021. Early intubation was defined as that occurring within the first 24â h of ICU admission. Propensity score matching was used to achieve a balance across baseline variables between the early intubation cohort and those patients who were intubated after the first 24â h of ICU admission. Differences in outcomes between early and delayed intubation were also assessed. We performed sensitivity analyses to consider a different time-point (48â h from ICU admission) for early and delayed intubation. RESULTS: Of the 2725 patients who received invasive mechanical ventilation, a total of 614 matched patients were included in the analysis (307 for each group). In the unmatched population, there were no differences in mortality between the early and delayed groups. After propensity score matching, patients with delayed intubation presented higher hospital mortality (27.3% versus 37.1%; p=0.01), ICU mortality (25.7% versus 36.1%; p=0.007) and 90-day mortality (30.9% versus 40.2%; p=0.02) compared with the early intubation group. Very similar findings were observed when we used a 48-h time-point for early or delayed intubation. The use of early intubation decreased after the first wave of the pandemic (72%, 49%, 46% and 45% in the first, second, third and fourth waves, respectively; first versus second, third and fourth waves p<0.001). In both the main and sensitivity analyses, hospital mortality was lower in patients receiving high-flow nasal cannula (HFNC) (n=294) who were intubated earlier. The subgroup of patients undergoing noninvasive ventilation (n=214) before intubation showed higher mortality when delayed intubation was set as that occurring after 48â h from ICU admission, but not when after 24â h. CONCLUSIONS: In patients with COVID-19 requiring invasive mechanical ventilation, delayed intubation was associated with a higher risk of hospital mortality. The use of early intubation significantly decreased throughout the course of the pandemic. Benefits of such an approach occurred more notably in patients who had received HFNC.
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COVID-19 , Ventilación no Invasiva , Insuficiencia Respiratoria , Humanos , Estudios Prospectivos , Pandemias , Intubación Intratraqueal/efectos adversos , Respiración Artificial/efectos adversos , Insuficiencia Respiratoria/terapia , Insuficiencia Respiratoria/etiología , Unidades de Cuidados IntensivosRESUMEN
Previous studies have shown that intermittent exposure to a 50 Hz, 100 µT sinusoidal magnetic field (MF) promotes proliferation of human neuroblastoma cells, NB69. This effect is mediated by activation of the epidermal growth factor receptor through a free radical-dependent activation of the p38 pathway. The present study investigated the possibility that the oxidative stress-sensitive protein p53 is a potential target of the MF, and that field exposure can affect the protein expression. To that end, NB69 cells were exposed to short intervals of 30 to 120 min to the aforementioned MF parameters. Two specific anti-p53 antibodies that allow discrimination between the wild and unfolded forms of p53 were used to study the expression and cellular distribution of both isoforms of the protein. The expression of the antiapoptotic protein Bcl-2, whose regulation is mediated by p53, was also analyzed. The obtained results revealed that MF exposure induced increases in p53 gene expression and in protein expression of the wild-type form of p53. Field exposure also caused overexpression of the unfolded form of p53, together with changes in the nuclear/cytoplasmic distribution of both forms of the protein. The expression of protein Bcl-2 was also significantly increased in response to the MF. As a whole, these results indicated that the MF is capable of interacting with the function, distribution and conformation of protein p53. Such interactions could be involved in previously reported MF effects on NB69 proliferation promotion.
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Background: The clinical heterogeneity of COVID-19 suggests the existence of different phenotypes with prognostic implications. We aimed to analyze comorbidity patterns in critically ill COVID-19 patients and assess their impact on in-hospital outcomes, response to treatment and sequelae. Methods: Multicenter prospective/retrospective observational study in intensive care units of 55 Spanish hospitals. 5866 PCR-confirmed COVID-19 patients had comorbidities recorded at hospital admission; clinical and biological parameters, in-hospital procedures and complications throughout the stay; and, clinical complications, persistent symptoms and sequelae at 3 and 6 months. Findings: Latent class analysis identified 3 phenotypes using training and test subcohorts: low-morbidity (n=3385; 58%), younger and with few comorbidities; high-morbidity (n=2074; 35%), with high comorbid burden; and renal-morbidity (n=407; 7%), with chronic kidney disease (CKD), high comorbidity burden and the worst oxygenation profile. Renal-morbidity and high-morbidity had more in-hospital complications and higher mortality risk than low-morbidity (adjusted HR (95% CI): 1.57 (1.34-1.84) and 1.16 (1.05-1.28), respectively). Corticosteroids, but not tocilizumab, were associated with lower mortality risk (HR (95% CI) 0.76 (0.63-0.93)), especially in renal-morbidity and high-morbidity. Renal-morbidity and high-morbidity showed the worst lung function throughout the follow-up, with renal-morbidity having the highest risk of infectious complications (6%), emergency visits (29%) or hospital readmissions (14%) at 6 months (p<0.01). Interpretation: Comorbidity-based phenotypes were identified and associated with different expression of in-hospital complications, mortality, treatment response, and sequelae, with CKD playing a major role. This could help clinicians in day-to-day decision making including the management of post-discharge COVID-19 sequelae. Funding: ISCIII, UNESPA, CIBERES, FEDER, ESF.
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Due to their alleged analgesic, anti-inflammatory and tissue regenerative effects, capacitive-resistive electrothermal therapy (CRET), which is based on non-invasive exposure to radiofrequency (RF) currents, is often applied to chemotherapeutically treated patients with cancer. Our previous studies have demonstrated that subthermal CRET currents can elicit a number of cell responses, including anti-proliferative effects, in the human liver cancer cell line HepG2. Such effects involve significant changes in the regulation of proteins involved in MAPK signaling pathways, which are also implicated in the cancer cell response to standard anticancer drugs such as sorafenib. This overlap in response pathways may lead to competitive, neutralizing or blocking interactions between the electrical and chemical treatments, thus raising questions on the advisability of CRET treatment for their analgesic, anti-inflammatory or other purposes in patients undergoing chemotherapy. The present study analyzed the effects of simultaneous treatment with sorafenib and 448-kHz, subthermal CRET current on the proliferation and viability of HepG2 cell cultures. Cell viability was assessed through Trypan blue or XTT assays, while flow cytometry was applied for cell cycle and apoptosis analysis. The expression of proteins involved in cell proliferation were assessed by immunoblotting and immunofluorescence. The results revealed no evidence to suggest that the electrical treatment counteracted or neutralized the cellular response to sorafenib at the different conditions evaluated. Furthermore, at the standard pharmacological sorafenib concentration, 5 µM, the combined treatment elicited an anti-proliferative response significantly stronger than that induced by each of the treatments when applied separately in HepG2 cells. These data do not support the hypothesis that CRET exposure may inhibit or diminish the effects of a chemotherapeutic drug used in cancer treatment, and highlights the requirement for further investigation into the cell response to the combined action of electrical and chemical treatments.
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Capacitive-resistive electric transfer (CRET) therapies have been proposed as strategies for regeneration of cutaneous tissue lesions. Previous studies by our group have shown that intermittent stimulation with 448 kHz CRET currents at subthermal densities promotes in vitro proliferation of human stem cells involved in tissue regeneration. The present study investigates the effects of the in vitro exposure to these radiofrequency (RF) currents on the proliferation and migration of keratinocytes and fibroblasts, the main cell types involved in skin regeneration. The effects of the electric stimulation on cell proliferation and migration were studied through XTT and wound closure assays, respectively. The CRET effects on the expression and location of proteins involved in proliferation and migration were assessed by immunoblot and immunofluorescence. The obtained results reveal that electrostimulation promotes proliferation and/or migration in keratinocytes and fibroblasts. These effects would be mediated by changes observed in the expression and location of intercellular adhesion proteins such as ß-catenin and E-cadherin, of proteins involved in cell-to-substrate adhesion such as vinculin, p-FAK and the metalloproteinase MMP-9, and of other proteins that control both processes: MAP kinases p-p38, p-JUNK and p-ERK1/2. These responses could represent a mechanism underlying the promotion of normotrophic wound regeneration induced by CRET. Indeed, electric stimulation would favor completion of granulation tissue formation prior to the closure of the outer tissue layers, thus preventing abnormal wound cicatrization or chronification.
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Fibroblastos , Queratinocitos , Proliferación Celular , Humanos , Ondas de Radio , PielRESUMEN
Capacitive-Resistive Electric Transfer (CRET) thermotherapies aim at tissue repair and regeneration through non-invasive application of RF currents. We have reported that the cellular response to subthermal CRET currents is non-linearly dependent on the signal frequency, and that in vitro exposure to a 448-kHz CRET signal promotes ADSC proliferation, as well as collagen and glycosaminoglycan synthesis in prechondrocytic cells. The present work investigates the response of neonatal fibroblasts to subthermal exposure (100 µA/mm2) to two CRET signals: a 448-kHz, non-modulated sinusoidal wave vs. a 20-kHz amplitude-modulation of the 448-kHz carrier. To that end, cell proliferation and expression of the biomarkers Hsp47, Hsp27 and decorin were assessed by cell count, PCNA and Western blotting. The results revealed that while both signals significantly and equivalently increased early (4 h) expression of Hsp47, the modulated signal was more efficient in inducing Hsp27 and decorin overexpression and promoting cell proliferation. These data indicate that the cellular response is dependent on the RF signal modulation and suggest that the therapeutic effects of CRET could be mediated by promotion of fibroblastic proliferation and overexpression of biomarkers that are essential in skin regeneration.
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Fibroblastos/citología , Fibroblastos/efectos de la radiación , Ondas de Radio , Relación Dosis-Respuesta en la Radiación , Humanos , Recién NacidoRESUMEN
Our previous studies have shown that intermittent exposure to a 50-Hz, 100-µT sine wave magnetic field (MF) promotes human NB69 cell proliferation, mediated by activation of the epidermal growth factor receptor (EGFR) and pathways MAPK-ERK1/2 and p38; being the effects on proliferation and p38 activation blocked by the chelator N-acetylcysteine. The present work investigates the MF effects on free radical (FR) production, and the potential involvement of NADPH oxidase, the main source of reactive oxygen species (ROS), in the MF-induced activation of MAPK pathways. To this end, the field effects on MAPK-ERK1/2, -p38 and -JNK activation in the presence or absence of the NADPH oxidase inhibitor, diphenyleneiodonium chloride (DPI), as well as the expression of the p67phox subunit, were analyzed. The results revealed that field exposure increases FR production and induces early, transient expression of the cytosolic component of the NADPH oxidase, p67phox. Also, the MF-induced activation of the MAPK-JNK pathway, but not that of -ERK1/2 or -p38 pathways, was prevented in the presence of the DPI, which has been shown to significantly reduce p67phox expression. These data, together with those from previous studies, identify various, FR-dependent or -independent mechanisms, involved in the MF-induced proliferative response mediated by MAPK signaling activation.
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Sistema de Señalización de MAP Quinasas , Campos Magnéticos , NADPH Oxidasas/metabolismo , Neuroblastoma/patología , Línea Celular Tumoral , HumanosRESUMEN
Unlike general anaesthesia, neuraxial anaesthesia (NA) reduces the burden and risk of respiratory adverse events in the post-operative period. However, both patients affected by chronic obstructive pulmonary disease (COPD) and chest wall disorders and/or neuromuscular diseases may experience the development or the worsening of respiratory failure, even during surgery performed under NA; this latter negatively affects the function of accessory respiratory muscles, resulting in a blunted central response to hypercapnia and possibly in an exacerbation of cardiac dysfunction (NA-induced relative hypovolemia). According to European Respiratory Society (ERS) and American Thoracic Society (ATS) guidelines, non-invasive ventilation (NIV) is effective in the post-operative period for the treatment of both impaired pulmonary gas exchange and ventilation, while the intra-operative use of NIV in association with NA is just anecdotally reported in the literature. Whilst NIV does not assure a protected patent airway and requires the patient's cooperation, it is a handy tool during surgery under NA: NIV is reported to be successful for treatment of acute respiratory failure; it may be delivered through the patient's home ventilator, may reverse hypoventilation induced by sedatives or inadvertent spread of anaesthetic up to cervical dermatomes, and allow the avoidance of intubation in patients affected by chronic respiratory failure, prolonging the time of non-invasiveness of respiratory support (i.e., neuromuscular patients needing surgery). All these advantages could make NIV preferable to oxygen in carefully selected patients.
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Anestesia Epidural/métodos , Anestesia Raquidea/métodos , Ventilación no Invasiva/métodos , Humanos , Enfermedades Neuromusculares/complicaciones , Enfermedades Neuromusculares/fisiopatología , Selección de Paciente , Enfermedad Pulmonar Obstructiva Crónica/complicaciones , Enfermedad Pulmonar Obstructiva Crónica/fisiopatología , Insuficiencia Respiratoria/etiología , Insuficiencia Respiratoria/prevención & control , Enfermedades Torácicas/complicaciones , Enfermedades Torácicas/fisiopatologíaRESUMEN
BACKGROUND: Capacitive-resistive electric transfer (CRET) is a non-invasive therapeutic strategy that applies radiofrequency electric currents within the 400-600 kHz range to tissue repair and regeneration. Previous studies by our group have shown that 48 h of intermittent exposure to a 570 kHz CRET signal at a subthermal density of 50 µA/mm2 causes significant changes in the expression and activation of cell cycle control proteins, leading to cycle arrest in human cancer cell cultures. The present study investigates the relevance of the signal frequency in the response of the human neuroblastoma cell line NB69 to subthermal electric treatment with four different signal frequency currents within the 350-650 kHz range. METHODS: Trypan blue assay, flow cytometry, immunofluorescence and immunoblot were used to study the effects of subthermal CRET currents on cell viability, cell cycle progression and the expression of several marker proteins involved in NB69 cell death and proliferation. RESULTS: The results reveal that among the frequencies tested, only a 448 kHz signal elicited both proapoptotic and antiproliferative, statistically significant responses. The apoptotic effect would be due, at least in part, to significant changes induced by the 448 kHz signal in the expression of p53, Bax and caspase-3. The cytostatic response was preceded by alterations in the kinetics of the cell cycle and in the expression of proteins p-ERK1/2, cyclin D1 and p27, which is consistent with a potential involvement of the EGF receptor in electrically induced changes in the ERK1/2 pathway. This receives additional support from results indicating that the proapototic and antiproliferative responses to CRET can be transiently blocked when the electric stimulus is applied in the presence of PD98059, a chemical inhibitor of the ERK1/2 pathway. CONCLUSION: The understanding of the mechanisms underlying the ability of slowing down cancer cell growth through electrically-induced changes in the expression of proteins involved in the control of cell proliferation and apoptosis might afford new insights in the field of oncology.
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Cresta Neural/efectos de la radiación , Neuroblastoma/metabolismo , Neuroblastoma/patología , Tratamiento de Radiofrecuencia Pulsada/métodos , Apoptosis/efectos de la radiación , Caspasa 3/metabolismo , Puntos de Control del Ciclo Celular/efectos de la radiación , Línea Celular Tumoral , Proliferación Celular/efectos de la radiación , Supervivencia Celular/efectos de la radiación , Ciclina D1/metabolismo , Inhibidor p27 de las Quinasas Dependientes de la Ciclina/metabolismo , Receptores ErbB/metabolismo , Flavonoides/farmacología , Humanos , Sistema de Señalización de MAP Quinasas/efectos de los fármacos , Sistema de Señalización de MAP Quinasas/efectos de la radiación , Neuroblastoma/radioterapia , Transducción de Señal/efectos de la radiación , Proteína p53 Supresora de Tumor/metabolismo , Proteína X Asociada a bcl-2/metabolismoRESUMEN
BACKGROUND/AIMS: Previous studies have shown that a 63-hour, intermittent exposure to a 50 Hz, 100 µT magnetic field (MF) induces in the NB69 line of human neuroblastoma a proliferative response that is mediated by activation of the MAPK pathways ERK1/2 and p38. The present study aims to investigate the potential involvement of the epidermal growth factor receptor (EGFR) in the field-induced cell proliferation and activation of MAPK pathways. METHODS: NB69 cultures were MF- or sham-exposed for 5 to 30 minute intervals and 63 hours. Cell proliferation and activation of MAPK-ERK1/2, -p38 and -JNK was analyzed in the presence or absence of erlotinib, an effective inhibitor of EGFR tyrosine kinase. The expression of p-EGFR and MMP-9 in the presence or absence of MF was also studied. Between 3 and 7 replicates of each experiment were performed, using between 3 and 4 samples per experimental condition and replicate. At the end of each replicate, the samples were analyzed at short times (5-30 min) through immunofluorescence and Western blotting, and the growth response was assessed (63 hours interval) through dye exclusion with Trypan blue. RESULTS: The results confirmed that field exposure induces cell proliferation and activation of ERK1/2, p38 and JNK, and revealed that these effects were blocked with erlotinib. The data also showed that, compared to shamexposed controls, the MF exposure induces early and transient increases in the expression of p-EGFR and MMP-9 at 15 and 5 min from the exposure onset, respectively. CONCLUSION: The obtained results reveal that the activation of the MAPK-ERK1/2 and -p38 pathways by the MF is mediated by the EGF receptor. Taken together with our previously published results, this dataset suggests that the proliferative response induced in NB69 by a 63-hour exposure to a weak, power frequency MF, is mediated by early transient activation of EGFR in which MMP-9 would be involved.
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Proliferación Celular , Sistema de Señalización de MAP Quinasas , Campos Magnéticos , Proteínas de Neoplasias/metabolismo , Neuroblastoma/metabolismo , Línea Celular Tumoral , Receptores ErbB/genética , Receptores ErbB/metabolismo , Humanos , Proteínas de Neoplasias/genética , Neuroblastoma/genética , Neuroblastoma/patologíaRESUMEN
Occupational exposure to static and radiofrequency fields emitted by nuclear magnetic resonance spectrometers was assessed through systematic field metering during operation of 19 devices in nine research centers. Whereas no measurable levels of radiofrequency radiation were registered outside the spectrometers, significant exposure to static field was detected, with maximum values recorded at the user's hand (B = 683.00 mT) and head-thorax (B = 135.70 mT) during spectrometer manipulation. All values were well below the exposure limits set by the European standard for workers protection against the effects of acute field exposure only. As for potential effects of chronic exposure, waiting for more complete knowledge, adoption of technical and operational strategies for exposure minimizing is advisable. In this respect, the data revealed that compared with standard magnetic shielding, ultrashield technology allows a 20-65-fold reduction of the field strength received by the operator.
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Campos Electromagnéticos , Espectroscopía de Resonancia Magnética , Exposición Profesional/análisis , Monitoreo de Radiación/métodos , Protección Radiológica/métodos , Ondas de Radio , Europa (Continente) , Humanos , Factores de RiesgoAsunto(s)
Delirio , Respiración Artificial , Humanos , Factores de Tiempo , Desconexión del VentiladorRESUMEN
BACKGROUND: To assess the performance of Candida albicans germ tube antibody (CAGTA), (1 â 3)-ß-D-glucan (BDG), mannan antigen (mannan-Ag), anti-mannan antibodies (mannan-Ab), and Candida DNA for diagnosing invasive candidiasis (IC) in ICU patients with severe abdominal conditions (SAC). METHODS: A prospective study of 233 non-neutropenic patients with SAC on ICU admission and expected stay ≥ 7 days. CAGTA (cutoff positivity ≥ 1/160), BDG (≥80, 100 and 200 pg/mL), mannan-Ag (≥60 pg/mL), mannan-Ab (≥10 UA/mL) were measured twice a week, and Candida DNA only in patients treated with systemic antifungals. IC diagnosis required positivities of two biomarkers in a single sample or positivities of any biomarker in two consecutive samples. Patients were classified as neither colonized nor infected (n = 48), Candida spp. colonization (n = 154) (low-grade, n = 130; high-grade, n = 24), and IC (n = 31) (intra-abdominal candidiasis, n = 20; candidemia, n = 11). RESULTS: The combination of CAGTA and BDG positivities in a single sample or at least one of the two biomarkers positive in two consecutive samples showed 90.3 % (95 % CI 74.2-98.0) sensitivity, 42.1 % (95 % CI 35.2-98.8) specificity, and 96.6 % (95 % CI 90.5-98.8) negative predictive value. BDG positivities in two consecutive samples had 76.7 % (95 % CI 57.7-90.1) sensitivity and 57.2 % (95 % CI 49.9-64.3) specificity. Mannan-Ag, mannan-Ab, and Candida DNA individually or combined showed a low discriminating capacity. CONCLUSIONS: Positive Candida albicans germ tube antibody and (1 â 3)-ß-D-glucan in a single blood sample or (1 â 3)-ß-D-glucan positivity in two consecutive blood samples allowed discriminating invasive candidiasis from Candida spp. colonization in critically ill patients with severe abdominal conditions. These findings may be helpful to tailor empirical antifungal therapy in this patient population.
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Biomarcadores/sangre , Candidiasis Invasiva/diagnóstico , Anciano , Anciano de 80 o más Años , Anticuerpos Antifúngicos , Antifúngicos/uso terapéutico , Candida albicans/inmunología , Candida albicans/patogenicidad , Candidiasis Invasiva/mortalidad , Enfermedad Crítica/mortalidad , Enfermedad Crítica/enfermería , Femenino , Humanos , Unidades de Cuidados Intensivos , Masculino , Mananos/sangre , Persona de Mediana Edad , Estudios ProspectivosRESUMEN
The proliferative response of the neuroblastoma line NB69 to a 100 µT, 50 Hz magnetic field (MF) has been shown mediated by activation of the MAPK-ERK1/2 pathway. This work investigates the MF effect on the cell cycle of NB69, the participation of p38 and c-Jun N-terminal (JNK) kinases in the field-induced proliferative response and the potential involvement of reactive oxygen species (ROS) in the activation of the MAPK-ERK1/2 and -p38 signaling pathways. NB69 cultures were exposed to the 100 µT MF, either intermittently for 24, 42 or 63 h, or continuously for periods of 15 to 120 min, in the presence or absence of p38 or JNK inhibitors: SB203580 and SP600125, respectively. Antioxidant N-acetylcysteine (NAC) was used as ROS scavenger. Field exposure induced transient activation of p38, JNK and ERK1/2. The MF proliferative effect, which was mediated by changes in the cell cycle, was blocked by the p38 inhibitor, but not by the JNK inhibitor. NAC blocked the field effects on cell proliferation and p38 activation, but not those on ERK1/2 activation. The MF-induced proliferative effects are exerted through sequential upregulation of MAPK-p38 and -ERK1/2 activation, and they are likely mediated by a ROS-dependent activation of p38.
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Proliferación Celular , Sistema de Señalización de MAP Quinasas , Campos Magnéticos/efectos adversos , Neuroblastoma/etiología , Neuroblastoma/metabolismo , Especies Reactivas de Oxígeno/metabolismo , Ciclo Celular , Línea Celular Tumoral , Humanos , Neuroblastoma/patología , Proteínas Quinasas p38 Activadas por Mitógenos/metabolismoRESUMEN
The 448 kHz capacitiveresistive electric transfer (CRET) is an electrothermal therapy currently applied in anticellulite and antiobesity treatments. The aim of the present study was to determine whether exposure to the CRET electric signal at subthermal doses affected early adipogenic processes in adiposederived stem cells (ADSC) from human donors. ADSC were incubated for 2 or 9 days in the presence of adipogenic medium, and exposed or shamexposed to 5 min pulses of 448 kHz electric signal at 50 µA/mm2 during the last 48 h of the incubation. Colorimetric, immunofluorescence, western blotting and reverse transcriptionquantitative polymerase chain reaction assays were performed to assess adipogenic differentiation of the ADSC. Electric stimulation significantly decreased cytoplasmic lipid content, after both 2 and 9 days of differentiation. The antiadipogenic response in the 9 day samples was accompanied by activation of mitogenactivated protein kinase kinase 1/2, decreased expression and partial inactivation of peroxisome proliferatoractivated receptor (PPAR) γ, which was translocated from the nucleus to the cytoplasm, together with a significant decrease in the expression levels of the PPARG1 gene, perilipin, angiopoietinlike protein 4 and fatty acid synthase. These results demonstrated that subthermal stimulation with CRET interferes with the early adipogenic differentiation in ADSC, indicating that the electric stimulus itself can modulate processes controlling the synthesis and mobilization of fat, even in the absence of the concomitant thermal and mechanical components of the thermoelectric therapy CRET.
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Tejido Adiposo/metabolismo , Diferenciación Celular , Células Madre Mesenquimatosas/metabolismo , Tejido Adiposo/citología , Células Cultivadas , Estimulación Eléctrica , Humanos , Células Madre Mesenquimatosas/citología , Quinasas de Proteína Quinasa Activadas por Mitógenos/metabolismo , PPAR gamma/metabolismo , Perilipina-1/metabolismoRESUMEN
BACKGROUND/AIMS: Semicircular lipoatrophy (SL) is an idiopathic condition characterized by atrophy of subcutaneous fatty tissue. Although several studies have suggested a possible association between SL and occupational exposure to power frequency magnetic fields (MF), no mechanism has been proposed so far that explains an influence of these fields on adipogenesis. METHODS: The study investigates the effects of a 50 Hz, 100 µT MF on the adipogenesis of stem cells isolated from human adipose tissue (ADSC). Cells were plated in Petri dishes and either exposed intermittently to the field for 42 hours or sham-exposed. RESULTS: Field exposure significantly reduced lipid accumulation within the cells, revealed in Oil Red O stained samples by spectrophotometry and colorimetry. Early cell passages were particularly sensitive to the effect: 30.40 ± 5.77% and 47.96 ± 12.47% below controls in the spectrophotometric and colorimetric assays, respectively. Such antiadipogenic effect was accompanied by significant changes in the expression of key effectors/regulators of early adipogenesis: PPARx03B3;, p-ERK1/2 and Sox9, indicating that at least the ERK/PPARx03B3; signaling pathway could be involved in the effect. CONCLUSIONS: These results constitute an experimental support to the hypothesis that power frequency MF can be one of the factors involved in the etiology of SL.
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Tejido Adiposo/citología , Diferenciación Celular , Magnetismo , Tejido Adiposo/metabolismo , Células Cultivadas , Técnica del Anticuerpo Fluorescente Indirecta , Humanos , Metabolismo de los Lípidos , Sistema de Señalización de MAP Quinasas , PPAR gamma/metabolismo , Factor de Transcripción SOX9/metabolismoRESUMEN
BACKGROUND/AIMS: Capacitive-resistive electric transfer (CRET) is a non invasive electrothermal therapy that applies electric currents within the 400 kHz - 450 kHz frequency range to the treatment of musculoskeletal lesions. Evidence exists that electric currents and electric or magnetic fields can influence proliferative and/or differentiating processes involved in tissue regeneration. This work investigates proliferative responses potentially underlying CRET effects on tissue repair. METHODS: XTT assay, flow cytometry, immunofluorescence and Western Blot analyses were conducted to asses viability, proliferation and differentiation of adipose-derived stem cells (ADSC) from healthy donors, after short, repeated (5 m On/4 h Off) in vitro stimulation with a 448-kHz electric signal currently used in CRET therapy, applied at a subthermal dose of 50 µA/mm(2) RESULTS: The treatment induced PCNA and ERK1/2 upregulation, together with significant increases in the fractions of ADSC undergoing cycle phases S, G2 and M, and enhanced cell proliferation rate. This proliferative effect did not compromise the multipotential ability of ADSC for subsequent adipogenic, chondrogenic or osteogenic differentiation. CONCLUSIONS: These data identify cellular and molecular phenomena potentially underlying the response to CRET and indicate that CRET-induced lesion repair could be mediated by stimulation of the proliferation of stem cells present in the injured tissues.
Asunto(s)
Proliferación Celular/fisiología , Células Madre Mesenquimatosas/fisiología , Tejido Adiposo/metabolismo , Tejido Adiposo/fisiología , Adulto , Anciano , Ciclo Celular/fisiología , Diferenciación Celular/fisiología , Células Cultivadas , Estimulación Eléctrica/métodos , Electricidad , Femenino , Humanos , Sistema de Señalización de MAP Quinasas/fisiología , Masculino , Células Madre Mesenquimatosas/metabolismo , Células Madre/metabolismo , Células Madre/fisiología , Regulación hacia Arriba/fisiologíaRESUMEN
Capacitive Resistive Electric Transfer (CRET) therapy applies currents of 0.4-0.6 MHz to treatment of inflammatory and musculoskeletal injuries. Previous studies have shown that intermittent exposure to CRET currents at subthermal doses exert cytotoxic or antiproliferative effects in human neuroblastoma or hepatocarcinoma cells, respectively. It has been proposed that such effects would be mediated by cell cycle arrest and by changes in the expression of cyclins and cyclin-dependent kinase inhibitors. The present work focuses on the study of the molecular mechanisms involved in CRET-induced cytostasis and investigates the possibility that the cellular response to the treatment extends to other phenomena, including induction of apoptosis and/or of changes in the differentiation stage of hepatocarcinoma cells. The obtained results show that the reported antiproliferative action of intermittent stimulation (5 m On/4 h Off) with 0.57 MHz, sine wave signal at a current density of 50 µA/mm(2), could be mediated by significant increase of the apoptotic rate as well as significant changes in the expression of proteins p53 and Bcl-2. The results also revealed a significantly decreased expression of alpha-fetoprotein in the treated samples, which, together with an increased concentration of albumin released into the medium by the stimulated cells, can be interpreted as evidence of a transient cytodifferentiating response elicited by the current. The fact that this type of electrical stimulation is capable of promoting both, differentiation and cell cycle arrest in human cancer cells, is of potential interest for a possible extension of the applications of CRET therapy towards the field of oncology.