CD44 Modulates Cell Migration and Invasion in Ewing Sarcoma Cells.

The chimeric EWSR1::FLI1 transcription factor is the main oncogenic event in Ewing sarcoma. Recently, it has been proposed that EWSR1::FLI1 levels can fluctuate in Ewing sarcoma cells, giving rise to two cell populations. EWSR1::FLI1low cells present a migratory and invasive phenotype, while EWSR1::FLI1high cells are more proliferative. In this work, we described how the CD44 standard isoform (CD44s), a transmembrane protein involved in cell adhesion and migration, is overexpressed in the EWSR1::FLI1low phenotype. The functional characterization of CD44s (proliferation, clonogenicity, migration, and invasion ability) was performed in three doxycycline-inducible Ewing sarcoma cell models (A673, MHH-ES1, and CADO-ES1). As a result, CD44s expression reduced cell proliferation in all the cell lines tested without affecting clonogenicity. Additionally, CD44s increased cell migration in A673 and MHH-ES1, without effects in CADO-ES1. As hyaluronan is the main ligand of CD44s, its effect on migration ability was also assessed, showing that high molecular weight hyaluronic acid (HMW-HA) blocked cell migration while low molecular weight hyaluronic acid (LMW-HA) increased it. Invasion ability was correlated with CD44 expression in A673 and MHH-ES1 cell lines. CD44s, upregulated upon EWSR1::FLI1 knockdown, regulates cell migration and invasion in Ewing sarcoma cells.

Effects of Chia Seeds on Growth Performance, Carcass Traits and Fatty Acid Profile of Lamb Meat.

The aim of this work was to supplement a diet with chia seeds (Salvia hispanica L.) based on the requirements of finishing lambs for meat, and to analyze biometric parameters and fatty acid profiles in meat. Eighteen male Rambouillet lambs with a bodyweight of 25 kg were used. Animals were kept in individual pens with water and feed provided ad libitum. Three finishing diets were designed with the inclusion of 0, 50 and 100 g dry matter chia seeds and divided among the animals (n = 6). The experimental period lasted 60 days. The weights of the individual lambs were recorded every 14 days. At the end of the experiment, the animals were slaughtered and the weights of the hot carcasses and non-meat components were registered. In addition, an analysis of the fatty acid composition was carried out in the muscles (Longissimus thoracis). The total weight gain and average daily gain displayed significant differences (p < 0.05). Initial and final bodyweights, such as the dry matter intake, did not display differences. The fatty acid profile of the meat tended to decrease the SFA (stearic acid) and increase MUFA (oleic acid) (p < 0.0001) when chia seeds were added to the lamb diets. In conclusion, chia seed supplementation did not increase meat production or other biometric parameters; however, it modified the fatty acid profile in L. thoracis.

A Systematic Review of the Adherence to Home-Practice Meditation Exercises in Patients with Chronic Pain.

Mindfulness-, compassion-, and acceptance-based (i.e., "third wave") psychotherapies are effective for treating chronic pain conditions. Many of these programs require that patients engage in the systematic home practice of meditation experiences so they can develop meditation skills. This systematic review aimed at evaluating the frequency, duration, and effects of home practice in patients with chronic pain undergoing a "third wave" psychotherapy. A comprehensive database search for quantitative studies was conducted in PubMed, Embase, and Web of Sciences Core Collection; 31 studies fulfilled the inclusion criteria. The reviewed studies tended to indicate a pattern of moderately frequent practice (around four days/week), with very high variability in terms of time invested; most studies observed significant associations between the amount of practice and positive health outcomes. Mindfulness-Based Stress Reduction and Mindfulness-Based Cognitive Therapy were the most common interventions and presented low levels of adherence to home practice (39.6% of the recommended time). Some studies were conducted on samples of adolescents, who practiced very few minutes, and a few tested eHealth interventions with heterogeneous adherence levels. In conclusion, some adaptations may be required so that patients with chronic pain can engage more easily and, thus, effectively in home meditation practices.

Effectiveness of four deconstructive meditative practices on well-being and self-deconstruction: study protocol for an exploratory randomized controlled trial.

INTRODUCTION: The efficacy of interventions based on mindfulness and compassion has been demonstrated in both clinical and general population, and in different social contexts. These interventions include so-called attentional and constructive meditation practices, respectively. However, there is a third group, known as deconstructive meditation practices, which has not been scientifically studied. Deconstructive practices aim to undo maladaptive cognitive patterns and generate knowledge about internal models of oneself, others and the world. Although there are theoretical and philosophical studies on the origin of addiction to the self or on the mechanisms of action associated with the deconstruction of the self, there are no randomized controlled trials evaluating these techniques in either a healthy population or clinical samples. This study aims to evaluate the effect of three deconstructive techniques by comparing them to mindfulness in the general population. METHODS AND ANALYSIS: A randomized controlled clinical trial will be conducted with about 240 participants allocated to four groups: (a) mindful breathing, (b) prostrations, according to Tibetan Buddhist tradition; (c) the Koan Mu, according to Zen Buddhist tradition; and (d) the mirror exercise, according to Toltec tradition. The primary outcome will be the qualities of the non-dual experience and spiritual awakening, measured by the Nondual Embodiment Thematic Inventory, assessed at pre- and post-treatment and at 3- and 6-month follow-ups. Other outcomes will be mindfulness, happiness, compassion, affectivity and altered state of consciousness. Quantitative data will be compared using mixed-effects linear regression models, and qualitative data will be analysed through thematic analysis and using the constant comparative method from grounded theory. ETHICS AND DISSEMINATION: Approval was obtained from the Research Ethics Committee of Aragon, Spain. The results will be submitted to peer-reviewed specialized journals, and brief reports will be sent to participants on request. TRIAL REGISTRATION: ClinicalTrials.gov NCT05317754. Registered on August 2,2022.

Effects of eHealth interventions on stress reduction and mental health promotion in healthcare professionals: A systematic review.

AIMS: To evaluate the effectiveness of eHealth interventions to reduce stress and promote mental health in healthcare professionals, and to compare the efficacy of different types of programs (guided vs. self-guided; 'third-wave' psychotherapies vs. other types). BACKGROUND: Healthcare workers present high levels of stress, which constitutes a risk factor for developing mental health problems such as depression and anxiety. eHealth interventions have been designed to reduce these professional's stress considering that the characteristics of this delivery method make it a cost-effective and very appealing alternative because of its fast and easy access. DESIGN: A systematic review of quantitative studies. METHODS: A comprehensive database search for quantitative studies was conducted in PubMed, EMBASE and Cochrane (until 1 April 2022). The systematic review was conducted in accordance with the PRISMA and SWiM reporting guidelines. The quality of the studies was assessed using the National Heart, Lung and Blood Institute tools. RESULTS: The abstracts of 6349 articles were assessed and 60 underwent in-depth review, with 27 fulfilling the inclusion criteria. The interventions were classified according to their format (self-guided vs. guided) and contents ('third-wave' psychotherapies vs. others). Twenty-two interventions emerged, 13 of which produced significant posttreatment reductions in stress levels of health professionals (9 self-guided, 8 'third wave' psychotherapies). Significant effects in improving depressive symptomatology, anxiety, burnout, resilience and mindfulness, amongst others, were also found. CONCLUSION: The evidence gathered in this review highlights the heterogeneity of the eHealth interventions that have been studied; self-guided and 'third-wave' psychotherapy programs are the most common, often with promising results, although the methodological shortcomings of most studies hinder the extraction of sound conclusions. PROTOCOL REGISTRATION: PROSPERO CRD42022310199. No Patient or Public Contribution.

Effectiveness and implementation of an online intervention (MINDxYOU) for reducing stress and promote mental health among healthcare workers in Spain: a study protocol for a stepped-wedge cluster randomized trial.

BACKGROUND: The World Health Organization has formally recognized that healthcare professionals are at risk of developing mental health problems; finding ways to reduce their stress is mandatory to improve both their quality of life and, indirectly, their job performance. In recent years, particularly since the COVID-19 pandemic outbreak, there has been a proliferation of online interventions with promising results. The purpose of the present study is twofold: to test the effectiveness of an online, self-guided intervention, MINDxYOU, to reduce the stress levels of healthcare workers; and to conduct an implementation study of this intervention. Additionally, an economic evaluation of the intervention will be conducted. METHODS: The current study has a hybrid effectiveness-implementation type 2 design. A stepped wedge cluster randomized trial design will be used, with a cohort of 180 healthcare workers recruited in two Spanish provinces (Malaga and Zaragoza). The recruitment stage will commence in October 2022. Frontline health workers who provide direct care to people in a hospital, primary care center, or nursing home setting in both regions will participate. The effectiveness of the intervention will be studied, with perceived stress as the main outcome (Perceived Stress Scale), while other psychopathological symptoms and process variables (e.g., mindfulness, compassion, resilience, and psychological flexibility) will be also assessed as secondary outcomes. The implementation study will include analysis of feasibility, acceptability, adoption, appropriateness, fidelity, penetration, and sustainability. The incremental costs and benefits, in terms of quality-adjusted life years, will be examined by means of cost-utility and cost-effectiveness analyses. DISCUSSION: MINDxYOU is designed to reduce healthcare workers' stress levels through the practice of mindfulness, acceptance, and compassion, with a special focus on how to apply these skills to healthy habits and considering the particular stressors that these professionals face on a daily basis. The present study will show how implementation studies are useful for establishing the framework in which to address barriers to and promote facilitators for acceptability, appropriateness, adoption, feasibility, fidelity, penetration, and sustainability of online interventions. The ultimate goal is to reduce the research-to-practice gap. TRIAL REGISTRATION: This study was registered in ClinicalTrials.gov on 29/06/2022; registration number: NCT05436717.

Examining the Relation Between Practicing Meditation and Having Peak Experiences and Lucid Dreams. A Cross-Sectional Study.

The aim of this study was to compare meditators and non-meditators in terms of their tendency to have peak experiences and their dream lucidity, while examining the associations between these outcomes and some related variables such as non-dual awareness, mindfulness facets and absorption. In this cross-sectional study, 237 participants from general Spanish population completed an online survey that included ad hoc questions related to the study aim, along with the Five Facet Mindfulness Questionnaire (FFMQ), the Non-dual Embodiment Thematic Inventory (NETI), the Tellegen Absorption Scale (TAS) and the Lucidity and Consciousness in Dreams Scale (LUCID). Of the total, 110 participants were identified as meditators and 127 as non-meditators. More than half of the sample (58.2%) reported having experienced at least one peak experience in their life; these showed no differences in the number, intensity, or self-inducing ability of these experiences between both groups but were significantly more common among meditators (71.8% vs. 46.8%; p < 0.001), who also presented higher scores in most of the questionnaires, except for some LUCID subscales. Regression models demonstrated that being a meditator was a significant predictor of having had a peak experience, but not of LUCID scores. These results, which need to be interpreted considering the study limitations, support the potential of meditation to facilitate having peak experiences, while its impact on lucid dreams remains unclear.

Relationship between initial symptoms and the prognosis, sex, and demographic area of patients with COVID-19.

Background: A method of determining the initial symptoms and main prognostic identifiers for COVID-19 can be a key tool for physicians, especially primary care physicians. Therefore, the objective of this study was to examine the prognosis of patients with COVID-19 from two different demographic regions according to baseline and main symptoms, age, and sex. Methods: All individuals selected from both urban and rural health centers were over 18 years of age, had COVID-19 before 2 March 2021, and were followed up with a primary care physician. All patients included in this study were recruited in terms of sex, age at the time of infection, type of contact, baseline symptoms, primary and secondary symptomatology, emergency assistance, hospitalization, intensive care unit (ICU) admission, and death. Results: A total of 219 and 214 subjects were recruited from rural and urban health centers, respectively. Subjects with COVID-19 from rural areas were significantly older in age, with a higher proportion of men, and had significantly lower baseline and main symptoms than those from urban areas. In addition, the presence of both fever and dyspnea as the initial or main symptom is significantly associated with emergency assistance, hospitalization, and death, regardless of sex, age, and demographic area. This type of illness was reported to be significantly less frequent in the rural population than in the urban population. Conclusion: The presence of both fever and dyspnea as both initial and main symptoms is a poor prognostic factor for COVID-19, regardless of age, sex, and demographic areas. In addition, women reported lower levels of fever and dyspnea, requiring minimal emergency assistance and fewer hospitalization, and a lower rate of mortality than men. During a COVID-19 infection follow-up, subjects in rural areas seem to have less access to medical care than those in urban areas.

miR-320c Regulates SERPINA1 Expression and Is Induced in Patients With Pulmonary Disease / El miR-320c regula la expresión del gen SERPINA1 y se induce en pacientes con enfermedad pulmonar

Introduction: Alpha-1 antitrypsin deficiency (AATD) is a genetic condition resulting in lung and liver disease with a great clinical variability. MicroRNAs have been identified as disease modifiers; therefore miRNA deregulation could play an important role in disease heterogeneity. Members of miR-320 family are involved in regulating of multiple processes including inflammation, and have potential specific binding sites in the 3′UTR region of SERPINA1 gene. In this study we explore the involvement of miR-320c, a member of this family, in this disease. Methods: Firstly in vitro studies were carried out to demonstrate regulation of SERPINA1 gene by miR-320. Furthermore, the expression of miR-320c was analyzed in the blood of 98 individuals with different AAT serum levels by using quantitative PCR and expression was correlated to clinical parameters of the patients. Finally, HL60 cells were used to analyze induction of miR-320c in inflammatory conditions. Results: Overexpression of miR-320 members in human HepG2 cells led to inhibition of SERPINA1 expression. Analysis of miR-320c expression in patient's samples revealed significantly increased expression of miR-320c in individuals with pulmonary disease. Additionally, HL60 cells treated with the pro-inflammatory factor lipopolysaccharide (LPS) showed increase in miR-320c expression, suggesting that miR-320c responds to inflammation. Conclusion: Our findings demonstrate that miR-320c inhibits SERPINA1 expression in a hepatic cell line and its levels in blood are associated with lung disease in a cohort of patients with different AAT serum levels. These results suggest that miR-320c can play a role in AAT regulation and could be a biomarker of inflammatory processes in pulmonary diseases. (AU)

Introducción: La deficiencia de alfa-1 antitripsina (DAAT) es una condición genética que produce enfermedad pulmonar y hepática con una gran variabilidad clínica. Los microARN se han identificado como modificadores de la gravedad de algunas enfermedades y su desregulación podría desempeñar un papel en la heterogeneidad de esta enfermedad. Los miembros de la familia miR-320 regulan múltiples procesos, incluyendo la inflamación, y tienen lugares de unión en la región 3’UTR del gen SERPINA1. En este estudio exploramos la implicación del miR-320c, un miembro de esta familia, en la DAAT. Métodos: Primero se realizaron estudios in vitro para demostrar la regulación del gen SERPINA1 por parte del miR-320. Además, se analizó la expresión de miR-320c en la sangre de 98 individuos con diferentes niveles de AAT mediante PCR cuantitativa y se correlacionó con los parámetros clínicos. Por último, se utilizaron células HL60 para analizar la inducción de miR-320c en condiciones inflamatorias. Resultados: La sobreexpresión del miR-320 en células HepG2 inhibía la expresión del gen SERPINA1. El análisis de expresión de miR-320c en los pacientes reveló una expresión significativamente aumentada en los casos con enfermedad pulmonar. Por otro lado, las células HL60 tratadas con LPS como factor proinflamatorio mostraron un aumento de expresión de miR-320c, lo que sugiere que este miARN responde a procesos inflamatorios. Conclusión: Nuestros resultados demuestran que el miR-320c inhibe la expresión de SERPINA1 en células hepáticas y que sus niveles en sangre están asociados con la presencia de enfermedad pulmonar en pacientes con diferentes niveles de AAT. Esto sugiere que el miR-320c desempeña un papel en la regulación de los niveles de AAT y podría ser un biomarcador de inflamación en enfermedades pulmonares. (AU)

miR-320c Regulates SERPINA1 Expression and Is Induced in Patients With Pulmonary Disease.

INTRODUCTION: Alpha-1 antitrypsin deficiency (AATD) is a genetic condition resulting in lung and liver disease with a great clinical variability. MicroRNAs have been identified as disease modifiers; therefore miRNA deregulation could play an important role in disease heterogeneity. Members of miR-320 family are involved in regulating of multiple processes including inflammation, and have potential specific binding sites in the 3'UTR region of SERPINA1 gene. In this study we explore the involvement of miR-320c, a member of this family, in this disease. METHODS: Firstly in vitro studies were carried out to demonstrate regulation of SERPINA1 gene by miR-320. Furthermore, the expression of miR-320c was analyzed in the blood of 98 individuals with different AAT serum levels by using quantitative PCR and expression was correlated to clinical parameters of the patients. Finally, HL60 cells were used to analyze induction of miR-320c in inflammatory conditions. RESULTS: Overexpression of miR-320 members in human HepG2 cells led to inhibition of SERPINA1 expression. Analysis of miR-320c expression in patient's samples revealed significantly increased expression of miR-320c in individuals with pulmonary disease. Additionally, HL60 cells treated with the pro-inflammatory factor lipopolysaccharide (LPS) showed increase in miR-320c expression, suggesting that miR-320c responds to inflammation. CONCLUSION: Our findings demonstrate that miR-320c inhibits SERPINA1 expression in a hepatic cell line and its levels in blood are associated with lung disease in a cohort of patients with different AAT serum levels. These results suggest that miR-320c can play a role in AAT regulation and could be a biomarker of inflammatory processes in pulmonary diseases.

Development and validation of a new model for predicting hypocalcaemia after total thyroidectomy: the NuGra model.

BACKGROUND: Hypocalcaemia is the most frequent complication after total thyroidectomy. Finding a method for its early detection has become a priority. A single-center prospective cohort study was conducted to identify risk factors for postoperative hypocalcaemia, develop an early detection model, and test its validity in a different group of patients. METHODS: The sample was composed of patients who underwent a total thyroidectomy between May 2012 and September 2015. Demographic, clinical, laboratory, and surgical data were collected. The incidence of hypocalcaemia and permanent hypoparathyroidism was calculated. Bivariate and multivariate analysis identified several independent predictors of hypocalcaemia, which were used to design a predictive model. The validity of the model was subsequently tested in a different cohort. Area under the ROC curve (AUROC) was calculated to determine its predictive power. RESULTS: The study and validation groups included 352 and 118 patients, respectively. Seventy-three patients developed laboratory-confirmed hypocalcaemia (20.7%), and symptomatic in 43 (12.2%). Multivariate analysis confirmed as independent predictors of hypocalcaemia the higher number of parathyroid glands identified [OR 1.41(0.98, 2.02); p = 0.063] and pre-to-postoperative gradient of parathormone decline [OR 1.06(1.04, 1.08); p < 0.001]. Based on these variables, the NuGra (Number of parathyroid glands identified-Gradient of decline) model was developed for predicting laboratory-confirmed hypocalcaemia. Its predictive power was high (AUROC 0.902, CI 0.857-0.947) for the study and the validation group (AUROC 0.956, CI 0.919-0.993). CONCLUSIONS: A higher number of parathyroid glands identified and a higher gradient of parathormone decline are risk factors for post-thyroidectomy hypocalcaemia. The NuGra model is useful for early prediction of individual risk for hypocalcaemia.

miR-320c Regulates SERPINA1 Expression and Is Induced in Patients With Pulmonary Disease.

INTRODUCTION: Alpha-1 antitrypsin deficiency (AATD) is a genetic condition resulting in lung and liver disease with a great clinical variability. MicroRNAs have been identified as disease modifiers; therefore miRNA deregulation could play an important role in disease heterogeneity. Members of miR-320 family are involved in regulating of multiple processes including inflammation, and have potential specific binding sites in the 3'UTR region of SERPINA1 gene. In this study we explore the involvement of miR-320c, a member of this family, in this disease. METHODS: Firstly in vitro studies were carried out to demonstrate regulation of SERPINA1 gene by miR-320. Furthermore, the expression of miR-320c was analyzed in the blood of 98 individuals with different AAT serum levels by using quantitative PCR and expression was correlated to clinical parameters of the patients. Finally, HL60 cells were used to analyze induction of miR-320c in inflammatory conditions. RESULTS: Overexpression of miR-320 members in human HepG2 cells led to inhibition of SERPINA1 expression. Analysis of miR-320c expression in patient's samples revealed significantly increased expression of miR-320c in individuals with pulmonary disease. Additionally, HL60 cells treated with the pro-inflammatory factor lipopolysaccharide (LPS) showed increase in miR-320c expression, suggesting that miR-320c responds to inflammation. CONCLUSION: Our findings demonstrate that miR-320c inhibits SERPINA1 expression in a hepatic cell line and its levels in blood are associated with lung disease in a cohort of patients with different AAT serum levels. These results suggest that miR-320c can play a role in AAT regulation and could be a biomarker of inflammatory processes in pulmonary diseases.

Liver organoids reproduce alpha-1 antitrypsin deficiency-related liver disease.

BACKGROUND AND AIMS: Alpha-1 antitrypsin (AAT) is a product of SERPINA1 gene mainly expressed by hepatocytes. Clinically relevant mutations in the SERPINA1 gene, such as Z (Glu342Lys), results in an expression of misfolded AAT protein having high propensity to polymerize, accumulate in hepatocytes and thus to enhance a risk for hepatocyte damage and subsequent liver disease. So far, the relationship between the Z-AAT accumulation and liver cell damage remains not completely understood. We present three-dimensional organoid culture systems, as a novel tool for modeling Z-AAT-related liver diseases. METHODS: We have established liver organoids from liver biopsies of patients with homozygous (ZZ) and heterozygous (MZ) deficiency and normal (MM) genotypes of AAT. The features of these organoid models were characterized by analyzing AAT protein secretion and intracellular aggregation in MZ and ZZ genotypes as well as SERPINA1 expression in differentiated cultures. RESULTS: Transcriptional analysis of differentiated organoid cultures by RNA-Seq showed hepatocyte-specific gene expression profile. Genes, such as ALB, APOB, CYP3A4 and SERPINA1, were validated and confirmed through quantitative-PCR analysis. The organoids from MZ and ZZ cases showed intracellular aggregation and lower secretion of AAT protein, and lower expression of ALB and APOB, as typically seen in hepatocytes from Z-AAT deficiency patients. Furthermore, organoids responded to external stimulus. Treatment with oncostatin M, a well-known inducer of SERPINA1, increased expression of the full-length transcripts (AAT-1C) as well as the short transcript of AAT (AAT-ST1C4). CONCLUSIONS: Liver organoid model recapitulates the key features of Z-AAT deficiency and provides a useful tool for disease modeling.

Characterization of Novel Missense Variants of SERPINA1 Gene Causing Alpha-1 Antitrypsin Deficiency.

The SERPINA1 gene is highly polymorphic, with more than 100 variants described in databases. SERPINA1 encodes the alpha-1 antitrypsin (AAT) protein, and severe deficiency of AAT is a major contributor to pulmonary emphysema and liver diseases. In Spanish patients with AAT deficiency, we identified seven new variants of the SERPINA1 gene involving amino acid substitutions in different exons: PiSDonosti (S+Ser14Phe), PiTijarafe (Ile50Asn), PiSevilla (Ala58Asp), PiCadiz (Glu151Lys), PiTarragona (Phe227Cys), PiPuerto Real (Thr249Ala), and PiValencia (Lys328Glu). We examined the characteristics of these variants and the putative association with the disease. Mutant proteins were overexpressed in HEK293T cells, and AAT expression, polymerization, degradation, and secretion, as well as antielastase activity, were analyzed by periodic acid-Schiff staining, Western blotting, pulse-chase, and elastase inhibition assays. When overexpressed, S+S14F, I50N, A58D, F227C, and T249A variants formed intracellular polymers and did not secrete AAT protein. Both the E151K and K328E variants secreted AAT protein and did not form polymers, although K328E showed intracellular retention and reduced antielastase activity. We conclude that deficient variants may be more frequent than previously thought and that their discovery is possible only by the complete sequencing of the gene and subsequent functional characterization. Better knowledge of SERPINA1 variants would improve diagnosis and management of individuals with AAT deficiency.