Experiencias en evaluación en España: El caso de la Universidad De Granada

La siguiente presentación resume las experiencias españolas en función de la evaluación integral de las bibliotecas. El principal objetivo, lograr la Certificación ISO 9001:2000, mediante la aplicación del Sistema de Gestión de la Calidad (SGC) en la Biblioteca Universitaria, dado por la necesidad de demostrar la capacidad de la Organización para satisfacer de forma consistente y continua productos y/o servicios que satisfagan las necesidades, requisitos y expectativas de los usuarios.Facultad de Humanidades y Ciencias de la Educació

Guidelines for the creation of institutional repositories at universities and higher education institutions = Directrices para la creación de repositorios institucionales en universidades y organizaciones de educación superior = Diretrizes para criação dos repositórios institucionais nas universidades e organizações de educação superior

The ALFA programme of the European Commission (Latin America Academic Training) promotes and supports cooperation activities between universities of both continents1. The universities who are members of the ALFA Network Babel Library2 assume, as part of their mission, the search for excellence and educational quality. In the initial work proposal, it was determined, as one of the expected results, to write a document, as Guidelines, about the development of services based on the use of new information and communication technology. The Institutional Repository (IR) is understood as an information system that collects, preserves, disseminates and provides access to the intellectual and academic output of the university community. Nowadays,the IR is a key tool of the scientific and academic policy of the university. On the other hand, access to the full text of the digital learning objects makes the repository become a fundamental support tool for teaching and research, whilst at the same time multiplying the institution’s visibility in the international community. Within this scenario, it is the university libraries that must lead the implementation of the IRs to enhance the university’s educational competitiveness, because of their experience in information management in all its forms and contact with knowledge

Guidelines for the creation of institutional repositories at universities and higher education institutions = Directrices para la creación de repositorios institucionales en universidades y organizaciones de educación superior = Diretrizes para criação dos repositórios institucionais nas universidades e organizações de educação superior

The ALFA programme of the European Commission (Latin America Academic Training) promotes and supports cooperation activities between universities of both continents1. The universities who are members of the ALFA Network Babel Library2 assume, as part of their mission, the search for excellence and educational quality. In the initial work proposal, it was determined, as one of the expected results, to write a document, as Guidelines, about the development of services based on the use of new information and communication technology. The Institutional Repository (IR) is understood as an information system that collects, preserves, disseminates and provides access to the intellectual and academic output of the university community. Nowadays,the IR is a key tool of the scientific and academic policy of the university. On the other hand, access to the full text of the digital learning objects makes the repository become a fundamental support tool for teaching and research, whilst at the same time multiplying the institution’s visibility in the international community. Within this scenario, it is the university libraries that must lead the implementation of the IRs to enhance the university’s educational competitiveness, because of their experience in information management in all its forms and contact with knowledge

Proceso para la producción en continuo de lovastatina

Número de publicación: ES2319029 A1 (01.05.2009) También publicado como: ES2319029 B1 (10.02.2010) Número de Solicitud: Consulta de Expedientes OEPM (C.E.O.)P200700519 (09.02.2007)La presente invención consiste en la producción de lovastatina, a partir de cultivos de Aspergillus terreus en forma de pellets, mediante un proceso de fermentación en continuo en un biorreactor de tipo columna de burbujeo operando de forma análoga a un biorreactor de lecho fluidizado (BLF) con un cultivo en tres fases, gas, líquido y sólido. La producción de lovastatina está regulada mediante un mecanismo de inhibición por producto, dando lugar al cese de la síntesis cuando la concentración alcanza un determinado valor. El proceso permite la obtención en continuo de un caldo de fermentación libre de biomasa conteniendo éste la lovastatina, lo cual facilita las posteriores etapas de extracción y purificación. La operación en continuo da lugar a que parte de la lovastatina generada abandone el biorreactor de lecho fluidizado, disminuyendo de esta forma los efectos de la inhibición por producto.Universidad de Almerí

Communication as a Strategy to Promote Sports and Health Activities Designed for Adolescents

Physical activity reduces the risk of developing noncommunicable diseases and improves quality of life, providing health benefits for present and future generations. This is especially relevant for adolescents. Educational institutions are promoters of healthy habits through the organisation of different activities such as extracurricular sports programmes. These activities increase the rates of sports practice among adolescents. The literature shows that the perceived quality of sports and health services is an antecedent of users' behavioural intentions. The aim of this paper is to find out whether communication from educational/sports organisations influences adolescents' intentions to continue engaging in physical activity. A total of 1080 students participated, with a mean age of 13.76 +/- 1.39 years, 34.1% of whom were girls. Tests were conducted to verify the validity and reliability of the model that relates communication with value, satisfaction, and future intentions. Tests were conducted to verify the validity (average variance extracted was between 0.754 and 0.583) and reliability (composite reliability was between 0.925 and 0.813) of the model that relates communication with value, satisfaction, and future intentions. Confirmatory analyses and factor invariance tests were performed. The results revealed that communication is an antecedent of value, satisfaction, and future intentions. In conclusion, communication is a good strategy to consolidate sporting habits in both male and female adolescents

Embryogenic competence of microspores is associated to their ability to form a callosic, osmoprotective subintinal layer

[EN] Microspore embryogenesis is an experimental morphogenic pathway with important applications in basic research and applied plant breeding, but its genetic, cellular, and molecular bases are poorly understood. We applied a multi-disciplinary approach using confocal and electron microscopy, detection of Ca2+, callose, and cellulose, treatments with caffeine, digitonin, and endosidin7, morphometry, qPCR, osmometry, and viability assays in order to study the dynamics of cell wall formation during embryogenesis induction in a high-response rapeseed (Brassica napus) line and two recalcitrant rapeseed and eggplant (Solanum melongena) lines. Formation of a callose-rich subintinal layer (SL) was common to microspore embryogenesis in the different genotypes. However, this process was directly related to embryogenic response, being greater in high-response genotypes. A link could be established between Ca2+ influx, abnormal callose/cellulose deposition, and the genotype-specific embryogenic competence. Callose deposition in inner walls and SLs are independent processes, regulated by different callose synthases. Viability and control of internal osmolality are also related to SL formation. In summary, we identified one of the causes of recalcitrance to embryogenesis induction: a reduced or absent protective SL. In responding genotypes, SLs are markers for changes in cell fate and serve as osmoprotective barriers to increase viability in imbalanced in vitro environments. Genotype-specific differences relate to different responses against abiotic (heat/osmotic) stresses.Thanks are due to the Electron Microscopy Service of Universitat Politecnica de Valencia, Marisol Gascon (IBMCP Microscopy Service), Dr Kim Boutilier (WUR, Wageningen) for hosting ARS at her lab, and Dr Samantha Vernhettes (INRA Versailles) for kindly providing us with S4B. This work supported by grants AGL2014-55177-R and AGL2017-88135-R to JMSS from MINECO jointly funded by FEDER.Rivas-Sendra, A.; Corral Martínez, P.; Porcel, R.; Camacho-Fernández, C.; Calabuig-Serna, A.; Seguí-Simarro, JM. (2019). Embryogenic competence of microspores is associated to their ability to form a callosic, osmoprotective subintinal layer. Journal of Experimental Botany. 70(4):1267-1281. https://doi.org/10.1093/jxb/ery458S12671281704Abramova, L. I. (2003). Russian Journal of Plant Physiology, 50(3), 324-329. doi:10.1023/a:1023866019102Adkar-Purushothama, C. R., Brosseau, C., Giguère, T., Sano, T., Moffett, P., & Perreault, J.-P. (2015). Small RNA Derived from the Virulence Modulating Region of the Potato spindle tuber viroid Silences callose synthase Genes of Tomato Plants. The Plant Cell, 27(8), 2178-2194. doi:10.1105/tpc.15.00523Cordewener, J., Bergervoet, J., & Liu, C.-M. (2000). Changes in Protein Synthesis and Phosphorylation during Microspore Embryogenesis in Brassica napus. Journal of Plant Physiology, 156(2), 156-163. doi:10.1016/s0176-1617(00)80300-4Corral-Martínez, P., García-Fortea, E., Bernard, S., Driouich, A., & Seguí-Simarro, J. M. (2016). Ultrastructural Immunolocalization of Arabinogalactan Protein, Pectin and Hemicellulose Epitopes Through Anther Development inBrassica napus. Plant and Cell Physiology, 57(10), 2161-2174. doi:10.1093/pcp/pcw133Fortes, A. M., Testillano, P. S., Del Carmen Risueño, M., & Pais, M. S. (2002). Studies on callose and cutin during the expression of competence and determination for organogenic nodule formation from internodes of Humulus lupulus var. Nugget. Physiologia Plantarum, 116(1), 113-120. doi:10.1034/j.1399-3054.2002.1160114.xFurch, A. C. U., Hafke, J. B., Schulz, A., & van Bel, A. J. E. (2007). Ca2+-mediated remote control of reversible sieve tube occlusion in Vicia faba. Journal of Experimental Botany, 58(11), 2827-2838. doi:10.1093/jxb/erm143Grewal, R. K., Lulsdorf, M., Croser, J., Ochatt, S., Vandenberg, A., & Warkentin, T. D. (2009). Doubled-haploid production in chickpea (Cicer arietinum L.): role of stress treatments. Plant Cell Reports, 28(8), 1289-1299. doi:10.1007/s00299-009-0731-1Hoekstra, S., van Bergen, S., van Brouwershaven, I. ., Schilperoort, R. ., & Wang, M. (1997). Androgenesis in Hordeum vulgare L.: Effects of mannitol, calcium and abscisic acid on anther pretreatment. Plant Science, 126(2), 211-218. doi:10.1016/s0168-9452(97)00096-4Hong, Z., Delauney, A. J., & Verma, D. P. S. (2001). A Cell Plate–Specific Callose Synthase and Its Interaction with Phragmoplastin. The Plant Cell, 13(4), 755-768. doi:10.1105/tpc.13.4.755Jacobs, A. K., Lipka, V., Burton, R. A., Panstruga, R., Strizhov, N., Schulze-Lefert, P., & Fincher, G. B. (2003). An Arabidopsis Callose Synthase, GSL5, Is Required for Wound and Papillary Callose Formation. The Plant Cell, 15(11), 2503-2513. doi:10.1105/tpc.016097Jacquard, C., Mazeyrat-Gourbeyre, F., Devaux, P., Boutilier, K., Baillieul, F., & Clément, C. (2008). Microspore embryogenesis in barley: anther pre-treatment stimulates plant defence gene expression. Planta, 229(2), 393-402. doi:10.1007/s00425-008-0838-6Jensen, W. A. (1968). Cotton embryogenesis: The zygote. Planta, 79(4), 346-366. doi:10.1007/bf00386917Joosen, R., Cordewener, J., Supena, E. D. J., Vorst, O., Lammers, M., Maliepaard, C., … Boutilier, K. (2007). Combined Transcriptome and Proteome Analysis Identifies Pathways and Markers Associated with the Establishment of Rapeseed Microspore-Derived Embryo Development. Plant Physiology, 144(1), 155-172. doi:10.1104/pp.107.098723KAY, R., CHAN, A., DALY, M., & MCPHERSON, J. (1987). Duplication of CaMV 35S Promoter Sequences Creates a Strong Enhancer for Plant Genes. Science, 236(4806), 1299-1302. doi:10.1126/science.236.4806.1299Ochatt, S., Pech, C., Grewal, R., Conreux, C., Lulsdorf, M., & Jacas, L. (2009). Abiotic stress enhances androgenesis from isolated microspores of some legume species (Fabaceae). Journal of Plant Physiology, 166(12), 1314-1328. doi:10.1016/j.jplph.2009.01.011Park, E., Díaz-Moreno, S. M., Davis, D. J., Wilkop, T. E., Bulone, V., & Drakakaki, G. (2014). Endosidin 7 Specifically Arrests Late Cytokinesis and Inhibits Callose Biosynthesis, Revealing Distinct Trafficking Events during Cell Plate Maturation. Plant Physiology, 165(3), 1019-1034. doi:10.1104/pp.114.241497Parra-Vega, V., Corral-Martínez, P., Rivas-Sendra, A., & Seguí-Simarro, J. M. (2015). Induction of Embryogenesis in Brassica Napus Microspores Produces a Callosic Subintinal Layer and Abnormal Cell Walls with Altered Levels of Callose and Cellulose. Frontiers in Plant Science, 6. doi:10.3389/fpls.2015.01018Paul, D. C., & Goff, C. W. (1973). Comparative effects of caffeine, its analogues and calcium deficiency on cytokinesis. Experimental Cell Research, 78(2), 399-413. doi:10.1016/0014-4827(73)90085-2Pauls, K. P., Chan, J., Woronuk, G., Schulze, D., & Brazolot, J. (2006). When microspores decide to become embryos — cellular and molecular changesThis review is one of a selection of papers published in the Special Issue on Plant Cell Biology. Canadian Journal of Botany, 84(4), 668-678. doi:10.1139/b06-064Reynolds, T. L. (1990). Interactions between calcium and auxin during pollen androgenesis in anther cultures of Solanum carolinense L. Plant Science, 72(1), 109-114. doi:10.1016/0168-9452(90)90192-qReynolds, T. L. (2000). Effects of calcium on embryogenic induction and the accumulation of abscisic acid, and an early cysteine-labeled metallothionein gene in androgenic microspores of Triticum aestivum. Plant Science, 150(2), 201-207. doi:10.1016/s0168-9452(99)00187-9Rivas-Sendra, A., Calabuig-Serna, A., & Seguí-Simarro, J. M. (2017). Dynamics of Calcium during In vitro Microspore Embryogenesis and In vivo Microspore Development in Brassica napus and Solanum melongena. Frontiers in Plant Science, 8. doi:10.3389/fpls.2017.01177Rivas-Sendra, A., Campos-Vega, M., Calabuig-Serna, A., & Seguí-Simarro, J. M. (2017). Development and characterization of an eggplant (Solanum melongena) doubled haploid population and a doubled haploid line with high androgenic response. Euphytica, 213(4). doi:10.1007/s10681-017-1879-3Rivas-Sendra, A., Corral-Martínez, P., Camacho-Fernández, C., & Seguí-Simarro, J. M. (2015). Improved regeneration of eggplant doubled haploids from microspore-derived calli through organogenesis. Plant Cell, Tissue and Organ Culture (PCTOC), 122(3), 759-765. doi:10.1007/s11240-015-0791-6Saidi, Y., Finka, A., Muriset, M., Bromberg, Z., Weiss, Y. G., Maathuis, F. J. M., & Goloubinoff, P. (2009). The Heat Shock Response in Moss Plants Is Regulated by Specific Calcium-Permeable Channels in the Plasma Membrane. The Plant Cell, 21(9), 2829-2843. doi:10.1105/tpc.108.065318Samuels, A. L., & Staehelin, L. A. (1996). Caffeine inhibits cell plate formation by disrupting membrane reorganization just after the vesicle fusion step. Protoplasma, 195(1-4), 144-155. doi:10.1007/bf01279193Schindelin, J., Arganda-Carreras, I., Frise, E., Kaynig, V., Longair, M., Pietzsch, T., … Cardona, A. (2012). Fiji: an open-source platform for biological-image analysis. Nature Methods, 9(7), 676-682. doi:10.1038/nmeth.2019Schl�pmann, H., Bacic, A., & Read, S. (1993). A novel callose synthase from pollen tubes of Nicotiana. Planta, 191(4). doi:10.1007/bf00195748Shi, X., Sun, X., Zhang, Z., Feng, D., Zhang, Q., Han, L., … Lu, T. (2014). GLUCAN SYNTHASE-LIKE 5 (GSL5) Plays an Essential Role in Male Fertility by Regulating Callose Metabolism During Microsporogenesis in Rice. Plant and Cell Physiology, 56(3), 497-509. doi:10.1093/pcp/pcu193Slewinski, T. L., Baker, R. F., Stubert, A., & Braun, D. M. (2012). Tie-dyed2 Encodes a Callose Synthase That Functions in Vein Development and Affects Symplastic Trafficking within the Phloem of Maize Leaves. Plant Physiology, 160(3), 1540-1550. doi:10.1104/pp.112.202473Sun, F., Fan, G., Hu, Q., Zhou, Y., Guan, M., Tong, C., … Wang, H. (2017). The high-quality genome ofBrassica napuscultivar ‘ZS11’ reveals the introgression history in semi-winter morphotype. The Plant Journal, 92(3), 452-468. doi:10.1111/tpj.13669Tan, H., Yang, X., Zhang, F., Zheng, X., Qu, C., Mu, J., … Zuo, J. (2011). Enhanced Seed Oil Production in Canola by Conditional Expression of Brassica napus LEAFY COTYLEDON1 and LEC1-LIKE in Developing Seeds. Plant Physiology, 156(3), 1577-1588. doi:10.1104/pp.111.175000Töller, A., Brownfield, L., Neu, C., Twell, D., & Schulze-Lefert, P. (2008). Dual function of Arabidopsis glucan synthase-like genes GSL8 and GSL10 in male gametophyte development and plant growth. The Plant Journal, 54(5), 911-923. doi:10.1111/j.1365-313x.2008.03462.xVerma, D. P. S. (2001). CYTOKINESIS ANDBUILDING OF THECELLPLATE INPLANTS. Annual Review of Plant Physiology and Plant Molecular Biology, 52(1), 751-784. doi:10.1146/annurev.arplant.52.1.751Verma, D. P. S., & Hong, Z. (2001). Plant Molecular Biology, 47(6), 693-701. doi:10.1023/a:1013679111111Vithanage, H. I. M. V., Gleeson, P. A., & Clarke, A. E. (1980). The nature of callose produced during self-pollination inSecale cereale. Planta, 148(5), 498-509. doi:10.1007/bf00552666Waldmann, T., Jeblick, W., & Kauss, H. (1988). Induced net Ca2+ uptake and callose biosynthesis in suspension-cultured plant cells. Planta, 173(1), 88-95. doi:10.1007/bf00394492WHITE, P. J. (2003). Calcium in Plants. Annals of Botany, 92(4), 487-511. doi:10.1093/aob/mcg164Xie, B., Deng, Y., Kanaoka, M. M., Okada, K., & Hong, Z. (2012). Expression of Arabidopsis callose synthase 5 results in callose accumulation and cell wall permeability alteration. Plant Science, 183, 1-8. doi:10.1016/j.plantsci.2011.10.015Ling You, X., Seon Yi, J., & Eui Choi, Y. (2006). Cellular change and callose accumulation in zygotic embryos of Eleutherococcus senticosus caused by plasmolyzing pretreatment result in high frequency of single-cell-derived somatic embryogenesis. Protoplasma, 227(2-4), 105-112. doi:10.1007/s00709-006-0149-3Yu, Y., Jiao, L., Fu, S., Yin, L., Zhang, Y., & Lu, J. (2016). Callose Synthase Family Genes Involved in the Grapevine Defense Response to Downy Mildew Disease. Phytopathology®, 106(1), 56-64. doi:10.1094/phyto-07-15-0166-rZhang, C., Guinel, F. C., & Moffatt, B. A. (2002). A comparative ultrastructural study of pollen development in Arabidopsis thaliana ecotype Columbia and male-sterile mutant apt1-3. Protoplasma, 219(1-2), 59-71. doi:10.1007/s00709020000

Pilot study of cutaneous tolerability of fibrin-agarose substitutes in healthy volunteers

Objetivos: En el presente estudio se persigue comprobar posibles reacciones adversas, derivadas del uso tópico de láminas de fibrina-agarosa en el antebrazo de voluntarios sanos. Metodología: Se llevó a cabo un estudio experimental en siete voluntarios sanos, cinco varones y dos mujeres, que no presentaban ningún tipo de lesión cutánea visible. En el antebrazo de cada voluntario se colocaron dos láminas de fibrina-agarosa de 4 cm2 . Cada lámina se cubrió con un apósito impregnado y sobre una de las láminas se aplicó pomada antibiótica con mupirocina. Ambas láminas se cubrieron finalmente con un apósito protector y se mantuvieron en contacto directo sobre la piel durante 48 horas. Resultados: Los resultados determinaron que no se detectaron reacciones adversas después de 48 horas de evolución ni en los siguientes 7 días en ningún voluntario. Se observaron diferencias entre las dos láminas implantadas en cada voluntario, ya que al retirar el apósito cubierto con pomada antibiótica, la lámina presentaba un aspecto más hidratado que la que no llevaba pomada antibiótica. Conclusiones: El uso tópico de las láminas de fibrina-agarosa en voluntarios sanos no presenta reacciones adversas del tipo irritación o alergia al aplicarse directamente por vía tópica. Aunque el tamaño muestral del estudio es limitado, sugiere que la combinación de fibrina-agarosa se presenta como el biomaterial idóneo para el desarrollo de un modelo de piel artificial humana.Purpose: This study aims to analyse possible adverse reactions resulting from the topical use of fibrin-agarose substitutes in the forearm of healthy volunteers. Methods: An experimental study was carried out in seven healthy volunteers, five males and two females, who did not have any cutaneous lesion. Two fibrin-agarose substitutes of 4 cm2 were placed in the forearm of each volunteer. Each substitute was covered with an impregnated dressing and one of the substitutes was covered with antibiotic ointment (mupirocin). Both substitutes were finally covered with a protective dressing. The substitutes were maintained for 48 hours. Results: The results determined that no adverse reactions were detected in any volunteer after 48 hours and a week of evolution. Differences were observed between the two substitutes implanted in each volunteer, since when removing the covered dressing with antibiotic ointment, the substitute presented a more hydrated appearance than the one without antibiotic cream. Conclusions: The implant of fibrin-agarose substitutes in healthy volunteers does not present irritation or allergic type adverse reactions when they applied directly topically on the skin. Although the sample size is low, the fibrin-agarose combination is presented as the biomaterial suitable for the development of an artificial human skin model

Validation of Bact/Alert automathic system in the microbiological control of cell medicinal products of Advanced Therapies

Objetivo. El Control de calidad para demostrar que un producto está libre de agentes microbianos adventicios es un aspecto clave de control de procesos y evaluación de la calidad de todas las preparaciones medicinales celulares y en la ingeniería tisular. El objetivo de este estudio es validar el sistema de detección por hemocultivo BacT / ALERT para el control microbiológico de las células mesenquimales para terapia celular, según la Farmacopea Europea (EU.PH), 2.6.27. “Control microbiológico de productos celulares” (1). Método. Para el cálculo del límite de detección las botellas de hemocultivo fueron inoculadas e incubadas con 4 réplicas de 30 UFC, 5 réplicas de 15 UFC y 5 réplicas de 6 UFC de los microorganismos en ausencia de producto celular. Se llevaron a cabo también experimentos en presencia de producto con 400.000 células mesenquimales. Este método se ha comparado con el método de referencia de Esterilidad de la EU.PH (2). La especificidad se ensayó inoculando 5 réplicas con 400.000 células mesenquimales sin microorganismos. Resultados. Todas las botellas inoculadas con células mesenquimales sin microorganismos permanecieron negativas después de 7 días de incubación. Todas las botellas inoculadas con cepas bacterianas aerobias y anaerobias fueron detectadas como positivas por el sistema, en el caso del límite inferior (6 UFC) en menos de 36 horas. Se detectaron como positivas las botellas inoculadas con Candida albicans (6 UFC) en menos de 48 horas y con Aspergillus niger (6 UFC) en menos de 72 horas. No hubo diferencias notables en el tiempo de detección entre botellas inoculadas con y sin la presencia de células mesenquimales. Conclusión: El sistema de detección de hemocultivos Bact/Alert es un método fiable para la detección de la contaminación microbiana de medicamentos a base de células mesenquimales y cumple los requisitos de la UE PH, 2.6.27, para el control microbiológico de productos celulares.Objective. Quality control to demonstrate that a product is free from adventitious microbial agents is a key aspect of process control and quality evaluation of all cell medicinal preparations and in tisular engineering. Evaluate the validation of the BacT/ALERT Blood Culture System for the microbial control of mesenchymal cells for cell therapy according European Pharmacopoeia (EU.PH), 2.6.27. “Microbiological control of cellular products” (1). Method. Blood culture bottles were challenged with 4 replica of 30 cfu, 5 replica of 15 cfu and 5 replica of 6 cfu of the test microorganisms. Test were also carried out in the presence in each contaminated culture bottle of 400.000 mesenchymal cells. This method has been compared with the reference method for Sterility of the EU.PH (2). Specificity was tested inoculating 5 replicas of broth culture media with 400.000 cells without microorganisms. Results. All bottles challenged with mesenchymal cells without microorganisms remained negative after 7 days of incubation. All inoculated bottles with aerobic and anaerobic bacterial strains were flagged as positive for the system, in case of low inoculum (6 cfu) in less than 36 hours. Candida inoculated bottles (6 cfu) were detected in less than 48 hours and Aspergillus (6 cfu) in less than 72 hours. There were no significant differences in the detection time between bottles inoculated with and without the presence of mesenchymal cells. Conclusion: The BacT/ALERT blood culture detection system and is a reliable method for detection of microbial contamination of mesenchymal cells medicinal products that fulfils the requirements of the EU PH, 2.6.27, for the microbiological control of cellular products

Epithelial in vitro differentiation of human mesenchymal stem cells (hMSCs) from adipose tissue (AT) and bone marrow (BM): cellular characterization and study of HLA I and II expression

AGRADECIMIENTOS Laboratorio de Citogenética del servicio de Análisis Clínicos del Hospital Universitario Virgen de las Nieves. Servicio de Análisis Clínicos (Sección de Citometría/Biopatología tumoral) del Hos- pital Universitario Virgen de las Nieves.Introducción: Las células troncales mesenquimales derivadas de tejido adiposo o médula ósea constituyen uno de los tratamientos de terapia celular más utilizados en los ensayos clínicos actuales por su capacidad inmunomoduladora. Además, por su potencial de diferenciación a células epiteliales pueden ser utilizadas en ingeniería tisular incorporadas a tejidos artificiales como la piel o córnea, sustituyendo a las células epiteliales autólogas de estos tejidos. Es necesario realizar una correcta caracterización de estas células diferenciadas y estudiar el efecto de la diferenciación en la expresión del HLA de clase I y II. Objetivos: Caracterizar y realizar los controles de calidad GMP en dos líneas de células mesenquimales troncales humanas de distintos orígenes (tejido adiposo y médula ósea) tras diferenciarlas a células epiteliales in vitro, y analizar si se modifica la expresión de los marcadores HLA I y II antes y después del proceso diferenciador. Metodología: Se ha realizado el aislamiento y expansión de las dos líneas celulares de células mesenquimales troncales a partir del tejido fuente y se ha procedido a su diferenciación in vitro a células epiteliales mediante medios de cultivos suplementados con factores de crecimiento específico. Se han realizado controles de calidad siguiendo los requerimientos de las normas de correcta fabricación y se ha estudiado por citometría de flujo la expresión de HLA tipo I y II antes y después del proceso diferenciador. Finalmente se ha comprobado mediante estudio histológico e inmunohistoquímico las características de las células diferenciadas. Resultados: Se han aislado dos líneas de células mesenquimales troncales de tejido adiposo y médula ósea que cumplen los controles de calidad propuestos. Tras el proceso diferenciador in vitro, las células mesenquimales troncales humanas no expresan marcadores HLA (I y II) importantes en la respuesta inmune, pero sí expresan débilmente proteínas relacionadas con los principales estratos epiteliales (CK5, CK6 y CK14). Conclusión: La ausencia de expresión de marcadores de HLA I y II por citometría de flujo en las células diferenciadas favorecería su uso con carácter alogénico en la construcción de piel y córneas humanas por ingeniería de tejidos, sin embargo, son necesarios más estudios que confirmen estos resultados preliminares y protocolos que optimicen el proceso diferenciador in vitro de las células mesenquimales troncales.Background: Human mesenchymal stem cells derived from adipose tissue and bone marrow are one of the most common cell therapy procedures used in recent clinical trials due to their immunomodulation capacity. Furthermore, for their epithelial differentiation potential can be used in tissue engineering, incorporated in artificial tissues such as skin and cornea, replacing autologous epithelial cells. It is necessary to make a correct cellular characterization of differentiated cells and to study the effect in HLA I and II expression. Objetives: Characterization and quality controls under GMP conditions of in vitro differentiated human mesenchymal stem cells from different sources (adipose tissue and bone marrow) to epithelial lineage, and study of HLA I and II expression before and after differentiation. Methods: Isolation and expansion of two human mesenchymal stem cells lines from their tissues of origin and in vitro differentiation to epithelial cells using culture mediums supplemented with specific growth factors. Quality controls according Good Manufacturing Practices have been made and HLA I and II expression before and after differentiation have been studied. Finally, characteristics of differentiated cells have been demonstrated by histological and immunohistochemical analysis. Results: Two human mesenchymal stem cells lines from adipose tissue and bone marrow have been isolated complying with the proposed quality controls. After in vitro differentiation, human mesenchymal stem cells do not express HLA (I and II) markers, which are important in immune response, but weakly express proteins related to main epithelial layers of human skin (CK5, CK6 and CK14). Conclusion: The absence of expression of HLA I and II by flow cytometry in differentiated cells would promote the use of them with allogenic character to construct human skin and cornea by tissue engineering, however, more studies and protocols are required to confirm these preliminary results and to optimize in vitro differentiation of human mesenchymal stem cells.FIS ISC-III and FEDER PI13/0257

Optimization of human keratinocyte culture to develop an artificial human skin model: cell alternatives as feeder layer of Advanced Therapies

Agradecimientos: Servicio de Medicina Nuclear del Complejo Hospitalario Universitario de GranadaObjetivos: En el presente estudio se persigue optimizar el cultivo de queratinocitos para desarrollar un modelo de piel artificial humana. Para ello, se utilizan como capa alimentadora células de origen humano: fibroblastos dérmicos humanos y células mesenquimales troncales derivadas de tejido adiposo. Los resultados obtenidos se comparan con los fibroblastos 3T3, capa alimentadora de origen murino utilizada desde hace décadas. Metodología: Se llevó a cabo un estudio experimental, utilizando células de origen humano y células de origen murino subletalmente irradiadas, como capa alimentadora para el establecimiento del cultivo de queratinocitos. Se evaluó la tasa de expansión celular y la tasa de duplicación en el pase celular de queratinocitos y en la recuperación celular final que se llevó a cabo a las 3 semanas de cultivo; así como el rendimiento celular y la viabilidad celular, que también se evaluaron en el procesamiento inicial. Resultados: Los resultados determinan que los fibroblastos dérmicos humanos irradiados y las células mesenquimales troncales derivadas de tejido adiposo pueden actuar como capa alimentadora promoviendo la adhesión y la expansión celular de los queratinocitos. Los fibroblastos dérmicos humanos proporcionan resultados equiparables a los obtenidos con los fibroblastos 3T3 murinos. Conclusiones: Los fibroblastos dérmicos humanos irradiados proporcionan una capa alimentadora funcional que permite la expansión in vitro de manera eficaz de los queratinocitos que se van a utilizar con fines clínicos para el desarrollo de un modelo de piel artificial humana.Purpose: This study aims to optimize keratinocyte culture to develop an artificial human skin model. For this purpose, human cells are used as feeder layer: human dermal fibroblasts and adipose derived mesenchymal stem cells. The results obtained are compared with 3T3 fibroblasts, murine feeder layer used for decades. Methods: We conducted an experimental study using human and murine sub-lethally irradiated cells as feeder layer for the establishment of keratinocyte culture. Cell expansion rate and doubling rate were evaluated in the keratinocyte cell passage and in the final cell recovery (was carried out at 3 weeks). The yield and viability of keratinocytes were also evaluated in the initial processing. Results: The results determine that irradiated human dermal fibroblasts and irradiated adipose derived mesenchymal stem cells can act as feeder layer promoting adhesion and expansion of keratinocytes. Human dermal fibroblasts provide comparable results to those obtained with murine 3T3 fibroblasts. Conclusions: Irradiated human dermal fibroblasts provide a functional feeder layer which allows effectively in vitro expansion of keratinocytes to be used for clinical purposes for the development of an artificial human skin model