Pathogenic implications of dysregulated miRNAs in propionic acidemia related cardiomyopathy

Cardiac alterations (hypertrophic/dilated cardiomyopathy, and rhythm alterations) are one of the major causes of mortality and morbidity in propionic acidemia (PA), caused by the deficiency of the mitochondrial enzyme propionyl-CoA carboxylase (PCC), involved in the catabolism of branched-chain amino acids, cholesterol, and odd-chain fatty acids. Impaired mitochondrial oxidative phosphorylation has been documented in heart biopsies of PA patients, as well as in the hypomorphic Pcca−/−(A138T) mouse model, in the latter correlating with increased oxidative damage and elevated expression of cardiac dysfunction biomarkers atrial and brain natriuretic peptides (ANP and BNP) and beta-myosin heavy chain (β-MHC). Here we characterize the cardiac phenotype in the PA mouse model by histological and echocardiography studies and identify a series of upregulated cardiac-enriched microRNAs (miRNAs) in the PA mouse heart, some of them also altered as circulating miRNAs in PA patients’ plasma samples. In PA mice hearts, we show alterations in signaling pathways regulated by the identified miRNAs, which could be contributing to cardiac remodeling and dysfunction; notably, an activation of the mammalian target of rapamycin (mTOR) pathway and a decrease in autophagy, which are reverted by rapamycin treatment. In vitro studies in HL-1 cardiomyocytes indicate that propionate, the major toxic metabolite accumulating in the disease, triggers the increase in expression levels of miRNAs, BNP, and β-MHC, concomitant with an increase in reactive oxygen species. Our results highlight miRNAs and signaling alterations in the PCC-deficient heart which may contribute to the development of PA-associated cardiomyopathy and provide a basis to identify new targets for therapeutic interventionThis work was supported by Spanish Ministry of Economy and Competitiveness and European Regional Development Fund (grant number SAF2016-76004-R) and by Fundación Isabel Gemio and Fundación La Caixa (LCF/PR/PR16/ 11110018). AFG is funded by the FPI-UAM program, EAB and ARB by the Spanish Ministry of Science, Innovation and Universities (predoctoral fellowships FPU15/02923 and BES-2014-069420, respectively

Comparison of covered distances by u14 players in competition

[ES] Introducción En fútbol, el patrón de actividad de jugadores adultos, ha sido ampliamente estudiado, pero son escasos los que han centrado su atención en el análisis de los desplazamientos de los jugadores de categorías infantiles (Barbero et al., 2007). El objetivo es cuantificar la distancia total y tipos de intensidades que recorren los jugadores de un equipo infantil en competición. Método La muestra la forman 16 jugadores de fútbol infantil con una edad media de 12.69 años (± .479). Para la obtención de los datos, se utilizó un acelerómetro de tres ejes más giroscopio de tres ejes, extrayendo los resultados con la aplicación Zepp Play Soccer en concreto: distancia total recorrida, andando, en carrera y en sprint. Todos los datos fueron recogidos en partidos oficiales de la categoría infantil, en los cuatro primeros partidos de la liga. Resultados Los principales resultados medios de distancia total recorrida son de 4288.44 m. (± 1410.31), andando realizaron 2463 m. (± 904.31), en carrera 1520.94 m. (± 761.71) y en sprint 304.61 m. (± 266.02). Por puestos, el jugador que menos distancia total realiza es el portero 2862 m. (± 172.26) y el que más son los medios centros con 4662.58 m. (± 1513.57) y con datos similares defensas y delanteros. Sin embargo, en cuanto a la distancia en sprint son los delanteros y extremos los que más acciones a máxima intensidad realizan con una media de 545.44 M. (± 273.84), seguido por los medios centros y el que menor distancia recorre es el portero. Por último, existen diferencias significativas (p< .05) en la distancia corriendo entre el portero y el resto de puestos, y entre defensa y medios siendo mayor en los medios centros. También en el sprint, el portero con medios centros y delanteros, y entre delanteros con el resto, al realizar más metros en sprint los delanteros que el resto. Discusión y Conclusiones Los resultados del estudio muestran que en edades infantiles ya habría que comenzar con entrenamientos por puestos y adaptar las tareas de entrenamiento a las exigencias de los jugadores en sus respectivos puestos de juego, del mismo modo a lo que indica Echeazarra (2014).[EN] Introduction In football, the activity pattern of adult players has been widely studied, but there are few who have focused their attention on the analysis of the movements of the trainees (Barbero et al., 2007). The aim is to quantify the total distance and types of intensities that U14 players covered in competition. Method The sample is formed by 16 U14 football players with an average age of 12.69 years (± .479). The data were obtained with a three-axis accelerometer plus three-axis gyroscope, extracting the results with the Zepp Play Soccer application, in particular: total distance covered, walking, running and sprinting. All data was collected in official matches of U14 category, in the first four games of the league Results The main average results of total covered distance are 4288.44 m. (± 1410.31), walking perform 2463 m. (± 904.31), running 1520.94 m. (± 761.71) and in sprint 304.61 m. (± 266.02). By positions, the player with the least total distance is the goalkeeper 2862 m. (± 172.26) and the most midfielders with 4662.58 m. (± 1513.57) and with similar data defenders and forwards. However, in terms of sprinting distance, forwards, right midfielder and left midfielder are the players who cover most actions at maximum intensity with an average of 545.44 M. (± 273.84), followed by the midfielders and the shortest distance runs through the goalkeeper. Finally, there are significant differences (p< .05) in the running distance between the goalkeeper and the other posts, and between defenders and midfielders. Also in the sprint, there are significant differences, between the goalkeeper with midfielders and forwards, and forwards with the rest of the players, to make more meters in sprint. Discussion and Conclusions The results of the study show that in U14 players would be necessary to begin with training for positions and adapt the training tasks to the players´ demands in their respective game positions, in the same way as Echeazarra (2014).S

Effect of small sided games on motor coordination and coda of u-12 football players

[ES] El fútbol de alto nivel es muy estudiado, no siendo así las etapas de formación (González-Villoria, García-López & Contreras-Jordán, 2015). En este ámbito los cambios de dirección (CODA) y la coordinación son elementos importantes para el rendimiento (Pardeiro & Yanci, 2017). Los Small Side Games (SSGs) son juegos reducidos que simulan las características propias del fútbol (Rampinini et al., 2007). Método Para la obtención de los datos se les aplicó el test de coordinación motriz SportComp validado por Ruiz-Pérez, et al., (2017). Para la medida de los CODA se utiliza el MAT T (Modified Agility Test), sin y con balón. Se realizan ocho sesiones de SSGs en el grupo experimental (GE) y con metodología tradicional en el grupo control (GC). El GE lo forman 15 jugadores con edad media de 10.73 años y el GC lo forman 12 jugadores con edad media de 10.42 años. Resultados El GE mejoran en todas las variables, existiendo diferencias significativas (p <0.05), a excepción de “saltos a la pata coja” en la coordinación. En los CODA mejoran la media en el restest tanto con balón como sin balón, con diferencias significativas en el caso de los CODA con balón (regate y conducción). El GC sólo mejora en el desplazamiento sobre soportes, con diferencias significativas negativas en tres de las cinco variables de coordinación, y en los CODA no presentan resultados superiores en el retest ni diferencias significativas. Discusión y conclusiones La aplicación de un programa de entrenamiento basado en SSGs mejora la coordinación motriz y los CODA (con y sin balón) de jugadores Sub-12, con mejoras significativas tras la intervención. Los SSGs mejoran los factores que afectan al rendimiento de los jugadores de fútbol.[EN] High-level football is well studied, but the stages of formation are not (Gonzalez-Villoria, García-López & Contreras-Jordán, 2015). In this area, changes of direction (CODA) and the coordination are important elements for sport performance (Pardeiro & Yanci, 2017). Small Side Games (SSGs) are small games that simulate the characteristics of football (Rampinini et al, 2007). Method In order to obtain the data, the motor coordination test SportComp validated by Ruiz-Pérez, et al., (2017) was applied. For the measurement of CODA the MAT T (Modified Agility Test) is used, without and with a ball. There are eight sessions of SSGs in the experimental group (EG) and with traditional methodology in the control group (CG). The GE is made up of 15 players with an average age of 10.73 years and the CG is made up of 12 players with an average age of 10.42 years. Results The EG improve in all the variables, with significant differences (p <0.05), with the exception of "jumps to the lame leg", in the motor coordination. In the CODA, they improve the average in the restest both, with and without ball, with differences significant in the case of the CODA with a ball (dribbling and driving). CG only improves in the “displacement on supports”, with significant negative differences in three of the five variables in motor coordination, and in the CODA they do not present superior results in the retest nor differences significant. Discusión and Conclusions The application of a training program based on SSGs improve motor coordination and the CODA (with and without ball) of U-12 players, with improvements after the intervention. SSGs improve the factors that affect the performance of the football players.S

Comparison of the degree of current and future commitment of u14 football players. Difference by sex

[ES]El fútbol es el deporte más practicado y con mayor repercusión en España, especialmente en hombres, no siendo así en las mujeres (Pérez, et al., 2018). Cada día hay más mujeres y niñas que juegan al fútbol, de ahí la necesidad de estudiar el fútbol formativo por sexos. Así, Peñaloza et al., (2013) definen este concepto como el reflejo del deseo y la resolución de persistir en el esfuerzo deportivo durante cierto periodo de tiempo. Por ello conocer el grado de compromiso deportivo actual y futuro en los jóvenes jugadores, puede conllevar a tomar diferentes directrices para evitar la inactividad, mejorar la promoción deportiva y el burnout (estar quemado) en el deporte, en función del sexo de los jugadores Método Para la obtención de los datos se les aplicó la Escala de Grado de Compromiso Deportivo, en su versión castellana (Belando et al., 2012). Mide dos factores, el compromiso actual y futuro de los jugadores de fútbol. La muestra la componen 36 jugadores sub-14, siendo 18 hombres y 18 mujeres, con edad media de 13.5 años. El cuestionario se aplicó 15 minutos antes del comienzo del entrenamiento respetando la Declaración Ética de Helsinki. Resultados El factor más valorado por los deportistas es el compromiso futuro (M: 4.42), con escasa diferencia con el compromiso actual (M: 4.28). Diferenciando por sexos, las mujeres valoran más el compromiso futuro (M: 4.78) y actual (M: 4.33), los hombres los valoran en menor medida, el compromiso futuro (M: 4.06) y el compromiso actual (M: 4.22). Existen diferencias significativas (p<.05) por sexos en el compromiso futuro (p< .000), es decir, que las mujeres valoran más el compromiso en el futuro frente a los hombres, mientras que en el factor compromiso actual no existen diferencias entre sexos. En el resto de factores como posición o puesto, tanto general como por sexos no existen diferencias significativas. Discusión y Conclusiones Los datos en la categoría infantil indican que son las mujeres las que tienen más compromiso futuro frente a los hombres, pese a las dificultades del fútbol femenino. Se sienten más comprometidas con este deporte, frente a los hombres que no muestran tanto compromiso futuro. Estos datos corroboran los resultados obtenidos por Pérez et al., (2018) en el que las jugadoras tienen un mayor nivel de motivación intrínseca y otros estudios en los que se considera que mayores niveles de compromiso implican una tasa menor de abandono deportivo (Langarica, Gómez & Pelayo, 2018; Torregrosa, 2007).[EN] Football is the most practiced sport with the greatest repercussion in Spain, especially in men, but not in women (Pérez et al., 2018). Every day, there are more women and girls who play football, hence the importance to study formative football by sex. Thus, Peñaloza et al., (2013) define this concept as the reflection of desire and the resolution to resist in sports effort during a certain period of time. Therefore, knowing the degree of current and future sports commitment en young players, can lead to take different guidelines to avoid the inactivity, improve sports promotion and burnout in sport, depending on the sex of the players. Method In order to obtain the data, the Scale of Sports Commitment Degree was applied, in its Spanish version (Belando et al., 2012). It measures two factors, the current and future commitment of football players. The sample is composed of 36 U14 players, 18 men and 18 women with an average age of 13.5 years. The questionnaire was applied 15 minutes before the beginning of the training, respecting the Helsinki Ethical Declaration. Results The most valued factor by football players ir the future commitment (M: 4.42), with little difference with current commitment (M: 4.28). Differentiating by sex, women value future (M: 4.78) and current (M: 4.33) more than men, the future commitment (M: 4.06) and the current commitment (M: 4.22). There are significant differences (p<.05) in the future commitment by sex (p< .000), that is, women value more the commitment in the future in front of men, while in the current commitment factor, there are no significant differences between sex. In the other factors, such as the position of the players, there are no significant differences. Discussion and Conclusions The results in U14 show that women have more future commitment to men, depite the difficultis of women´s football. They feel more committed to this sport, compared to men who do not show so much future commitment. These data corroborate the results obtained by Pérez et al., (2018) in which women players have a higher level of intrinsic motivation and other studies in which it is considered that higher levels of commitment imply a lower rate of sports dropout (Langarica, Gómez & Pelayo, 2018; Torregrosa, 2007).S

Re-inventing Multimode Interference Couplers Using Subwavelength Gratings

We use the concept of subwavelength grating (SWG) refractive-index-engineering to propose and experimentally demonstrate a reduced size, slotted 2x2 MMI coupler. We also present an ultra-broadband 2x2 MMI coupler which is based on SWG dispersion engineering.This work was supported in part by the Spanish Ministerio de Ciencia (project TEC2009-10152), a Formación del Profesorado Universitario scholarship (AP-2006-03355), the European Mirthe project (FP7-2010-257980) and “Universidad de Málaga. Campus de Excelencia Internacional Andalucía Tech

Newly impaired glucose metabolism and prognosis after percutaneous revascularization

Background: Clinical practice guidelines recommend ad hoc screening of diabetes in patients admitted for macrovascular disease; however, these recommendations are rarely followed in real practice. This study was undertaken to assess whether impaired glucose metabolism, newly diagnosed after percutaneous coronary intervention (PCI) or known diabetes, provides prognostic information. Methods: We studied 374 patients who underwent PCI. An oral glucose tolerance test was carried out in the known non-diabetic patients with fasting glucose &lt; 7 mmol/L. Results: Eighty-one percent of the patients presented impaired glucose metabolism, from which 35.3% were previously diagnosed with diabetics, 21.4% were newly detected diabetics, and 24.3% were pre-diabetics. After a mean follow-up of 35.8 ± 13.4 months, only a known history of diabetes was an independent predictor of revascularization (OR = 2.03, p = 0.025), non-fatal acute myocardial infarction (OR = 2.70, p = 0.029) and readmission due to heart failure during the follow-up (OR = 3.82, p = 0.022). Conclusions: Screening for impaired glucose metabolism after PCI permits the detection of a high proportion of patients with abnormal glucose regulations. However, previously known diabetes remains the only independent predictor of cardiovascular events in the follow-up.

Association of breast and gut microbiota dysbiosis and the risk of breast cancer: a case-control clinical study

We would like to thank M Luisa Puertas-Martin and Isabel Manzano-Jimenez, nurses at the Unit of Mammary Pathology, General Surgery Service, San Cecilio University Hospital (Granada), without whose enthusiasm the enrolment of participants in Granada would still be stalled. We are indebted to all the women taking part in the study.The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.Trial registration ClinicalTrials.gov NCT03885648, 03/25/2019. Retrospectively registered.Background Breast cancer ranks first in women, and is the second cause of death in this gender. In addition to genetics, the environment contributes to the development of the disease, although the factors involved are not well known. Among the latter is the influence of microorganisms and, therefore, attention is recently being paid to the mammary microbiota. We hypothesize that the risk of breast cancer could be associated with the composition and functionality of the mammary/gut microbiota, and that exposure to environmental contaminants (endocrine disruptors, EDCs) might contribute to alter these microbiota. Methods We describe a case-control clinical study that will be performed in women between 25 and 70 years of age. Cases will be women diagnosed and surgically intervened of breast cancer (stages I and II). Women with antecedents of cancer or advanced tumor stage (metastasis), or who have received antibiotic treatment within a period of 3 months prior to recruitment, or any neoadjuvant therapy, will be excluded. Controls will be women surgically intervened of breast augmentation or reduction. Women with oncological, gynecological or endocrine history, and those who have received antibiotic treatment within a period of 3 months prior to recruitment will also be excluded. Blood, urine, breast tissue and stool samples will be collected. Data regarding anthropometric, sociodemographic, reproductive history, tumor features and dietary habits will be gathered. Metabolomic studies will be carried out in stool and breast tissue samples. Metagenomic studies will also be performed in stool and breast tissue samples to ascertain the viral, fungal, bacterial and archaea populations of the microbiota. Quantitation of estrogens, estrogen metabolites and EDCs in samples of serum, urine and breast tissue will also be performed. Discussion: This is the first time that the contribution of bacteria, archaea, viruses and fungi together with their alteration by environmental contaminants to the risk of breast cancer will be evaluated in the same study. Results obtained could contribute to elucidate risk factors, improve the prognosis, as well as to propose novel intervention studies in this disease.This work is funded by grants PI-0538-2017 (Junta de Andalucía, Spain, to LF) and Biomedical Research Networking Center-CIBER de Epidemiología y Salud Pública (CIBERESP) of the Institute of Health Carlos III -supported by European Regional Development Fund/FEDER (FIS-PI16/01812) (to MFF)

Abatacept in patients with rheumatoid arthritis and interstitial lung disease: A national multicenter study of 63 patients

OBJECTIVE: Interstitial lung disease (ILD) is one of the most serious complications of rheumatoid arthritis (RA). In the present study, we aimed to assess the efficacy of abatacept (ABA) in patients with ILD associated to RA. METHODS: National multicenter, non-controlled, open-label registry study of RA patients with ILD treated with ABA. RESULTS: 63 patients (36 women) with RA-associated ILD undergoing ABA therapy were studied. The mean ± standard deviation age at the time of the study was 63.2 ± 9.8 years. The median duration of RA and ILD from diagnosis were 6.8 and 1 year, respectively. RA was seropositive in 55 patients (87.3%). In 15 (23.8%) of 63 patients the development of ILD was closely related to the administration of synthetic or biologic disease modifying anti-rheumatic drugs. After a follow-up of 9.4 ± 3.2 months, two-thirds of patients remained stable whereas one-quarter experienced improvement in the Modified Medical Research Council scale. At that time forced vital capacity remained stable in almost two-thirds of patents and improved in one out of five patients assessed. Also, diffusing capacity of the lung for carbon monoxide remained stable in almost two-thirds and showed improvement in a quarter of the patients assessed. At 12 months, 50% of the 22 patients in whom chest HRCT scan was performed due persistence of respiratory symptoms showed stabilization, 8 (36.4%) improvement and 3 worsening of the HRCT scan pattern. Eleven of 63 patients had to discontinue ABA, mainly due to adverse events. CONCLUSION: ABA appears to be an effective in RA-associated ILD.Funding: This work was partially supported by RETICS Programs, RD08/0075 (RIER) and RD12/0009/0013 from ‘‘Instituto de Salud Carlos III’’ (ISCIII), Spain

Intermediate Molecular Phenotypes to Identify Genetic Markers of Anthracycline-Induced Cardiotoxicity Risk.

Cardiotoxicity due to anthracyclines (CDA) affects cancer patients, but we cannot predict who may suffer from this complication. CDA is a complex trait with a polygenic component that is mainly unidentified. We propose that levels of intermediate molecular phenotypes (IMPs) in the myocardium associated with histopathological damage could explain CDA susceptibility, so variants of genes encoding these IMPs could identify patients susceptible to this complication. Thus, a genetically heterogeneous cohort of mice (n = 165) generated by backcrossing were treated with doxorubicin and docetaxel. We quantified heart fibrosis using an Ariol slide scanner and intramyocardial levels of IMPs using multiplex bead arrays and QPCR. We identified quantitative trait loci linked to IMPs (ipQTLs) and cdaQTLs via linkage analysis. In three cancer patient cohorts, CDA was quantified using echocardiography or Cardiac Magnetic Resonance. CDA behaves as a complex trait in the mouse cohort. IMP levels in the myocardium were associated with CDA. ipQTLs integrated into genetic models with cdaQTLs account for more CDA phenotypic variation than that explained by cda-QTLs alone. Allelic forms of genes encoding IMPs associated with CDA in mice, including AKT1, MAPK14, MAPK8, STAT3, CAS3, and TP53, are genetic determinants of CDA in patients. Two genetic risk scores for pediatric patients (n = 71) and women with breast cancer (n = 420) were generated using machine-learning Least Absolute Shrinkage and Selection Operator (LASSO) regression. Thus, IMPs associated with heart damage identify genetic markers of CDA risk, thereby allowing more personalized patient management.J.P.L.’s lab is sponsored by Grant PID2020-118527RB-I00 funded by MCIN/AEI/10.13039/ 501100011039; Grant PDC2021-121735-I00 funded by MCIN/AEI/10.13039/501100011039 and by the “European Union Next Generation EU/PRTR”, the Regional Government of Castile and León (CSI144P20). J.P.L. and P.L.S. are supported by the Carlos III Health Institute (PIE14/00066). AGN laboratory and human patients’ studies are supported by an ISCIII project grant (PI18/01242). The Human Genotyping unit is a member of CeGen, PRB3, and is supported by grant PT17/0019 of the PE I + D + i 2013–2016, funded by ISCIII and ERDF. SCLl is supported by MINECO/FEDER research grants (RTI2018-094130-B-100). CH was supported by the Department of Defense (DoD) BCRP, No. BC190820; and the National Cancer Institute (NCI) at the National Institutes of Health (NIH), No. R01CA184476. Lawrence Berkeley National Laboratory (LBNL) is a multi-program national laboratory operated by the University of California for the DOE under contract DE AC02-05CH11231. The Proteomics Unit belongs to ProteoRed, PRB3-ISCIII, supported by grant PT17/0019/0023 of the PE I + D +i, 2017–2020, funded by ISCIII and FEDER. RCC is funded by fellowships from the Spanish Regional Government of Castile and León. NGS is a recipient of an FPU fellowship (MINECO/FEDER). hiPSC-CM studies were funded in part by the “la Caixa” Banking Foundation under the project code HR18-00304 and a Severo Ochoa CNIC Intramural Project (Exp. 12-2016 IGP) to J.J.S

A PD-L1/EGFR bispecific antibody combines immune checkpoint blockade and direct anti-cancer action for an enhanced anti-tumor response

Immune checkpoint blockade (ICB) with antibodies has shown durable clinical responses in a wide range of cancer types, but the overall response rate is still limited. Other effective therapeutic modalities to increase the ICB response rates are urgently needed. New bispecific antibody (bsAb) formats combining the ICB effect and a direct action on cancer cells could improve the efficacy of current immunotherapies. Here, we report the development of a PD-L1/EGFR symmetric bsAb by fusing a dual-targeting tandem trimmer body with the human IgG1 hinge and Fc regions. The bsAb was characterized in vitro and the antitumor efficacy was evaluated in humanized mice bearing xenografts of aggressive triple-negative breast cancer and lung cancer. The IgG-like hexavalent bsAb, designated IgTT-1E, was able to simultaneously bind both EGFR and PD-L1 antigens, inhibit EGF-mediated proliferation, effectively block PD-1/PD-L1 interaction, and induce strong antigen-specific antibody-dependent cellular cytotoxicity activity in vitro. Potent therapeutic efficacies of IgTT-1E in two different humanized mouse models were observed, where tumor growth control was associated with a significantly increased proportion of CD8+ T cells. These results support the development of IgTT-1E for the treatment of EGFR+ cancers.L.A-V. was supported by grants from the MCIN/AEI/10.13039/ 501100011033 (PID2020-117323RB-100 and PDC2021-121711-100), the Instituto de Salud Carlos III (DTS20/00089), the CRIS Cancer Foundation (FCRIS-2021-0090), the Spanish Association Against Cancer (PROYE19084ALVA), the Fundación ‘‘La Caixa’’ (HR21-00761 project IL7R_LungCan) and the Fundación de Investigación Biomédica 12 de Octubre Programa Investiga (2022-0082). B.B and L.S. were supported by grants PI20/01030 and PI19/00132 from the Instituto de Salud Carlos III (PI20/01030). FJB and MF-G were supported by grants PID2020- 113225GB-I00 and PRE2018-085788 funded by MCIN/AEI/10.13039/ 501100011033. L.R-P. was supported by a predoctoral fellowship from the Immunology Chair, Universidad Francisco de Vitoria/Merck. C. D-A. was supported by a predoctoral fellowship from the MCIN/AEI/ 10.13039/501100011033 (PRE2018-083445). L.D-A. was supported by a Rio Hortega fellowship from the Instituto de Salud Carlos III (CM20/ 00004). O.H. was supported by an industrial PhD fellowship from the Comunidad de Madrid (IND2020/BMD-17668). AE-L was supported industrial PhD fellowship from the Instituto de Salud Carlos III (IFI18/ 00045)Peer reviewe