Evaluation of two human dental pulp stem cell cryopreservation methods
Cargando...
Archivos
Fecha
2015
Título de la revista
Publicado en
Acta Odontológica Latinoamericana, 1852-4834, Vol. 28, Nro. 2, 2015, p. 114-121
Publicado por
Sociedad Argentina de Investigación Odontológica
URL de la fuente
ISSN de la revista
Título del volumen
Resumen
La pulpa dental es una fuente promisoria de celulas madre mesenquimales para ser utilizadas en terapia celular y medicina regenerativa. El desarrollo de metodos que permitan almacenar las celulas madre con minimo compromiso de la viabilidad celular, capacidad de diferenciacion y funcion es necesario para aplicaciones clinicas e investigacion. El objetivo de este estudio fue evaluar si las celulas troncales de pulpa dental humana (hDPSCs) aisladas y criopreservadas durante 1, 7 y 30 dias conservan la viabilidad y expresion de marcadores especificos de celulas troncales pos crio-preservacion. Para esto, las hDPSCs se aislaron de 23 pacientes sanos entre 18 y 31 anos. La pulpa dental se disocio enzimaticamente, y las celulas CD105+ se separaron mediante el sistema Miltenyi™. Posteriormente, las hDPSCs se criopreservaron utilizando el metodo de Kamath y de Papaccio, se evaluo la viabilidad pos crio-preservacion por citometria de flujo (7AAD) y la expresion de marcadores CD105+/ CD73+, CD34-/CD45-. El metodo de Papaccio, mostro mayor viabilidad celular a los 30 dias (59,5%) comparado con el metodo de Kamath, a 1 dia (65,5%) y 7 dias (56%) respectivamente. Se observo mayor expresion de los marcadores CD105+/CD34- a 1 y 7 dias pos-criopreservacion con el metodo Kamath y CD105+/CD45- a los 3 tiempos de criopreservacion. CD73+ presento mayor expresion en las hDPSCs a las 24 horas y 7 dias con el metodo de Kamath, y al mes con el metodo Papaccio. Estos resultados sugieren que las hDPSCs expresan marca - dores de celulas troncales mesenquimales postcriopreservacion. Sin embargo el tiempo de criopreservacion podria modificar la expresion de los marcadores probablemente por alterar
Descripción
Abstract
Dental pulp is a promising source of mesenchymal stem cells for use in cell therapy and regenerative medicine. Methods for storing stem cells with minimum compromise of cell viability, differentiation capacity and function should be developed for clinical and research applications. The aim of this study was to evaluate whether human dental pulp stem cells (hDPSCs) isolated and cryopreserved for 1, 7 and 30 days maintain viability and expression of specific stem cell markers. Human dental pulp stem cells were isolated from 23 healthy patients aged 18 to 31 years. Dental pulp was enzymatically dissociated, and CD105+ cells were separated using the Miltenyi™ system. The hDPSCs were cryopreserved using the Kamath and Papaccio methods. Post-cryopreservation viability was measured by flow cytometry (7AAD) and by the expression of the phenotype markers CD105+/ CD73+, CD34-/CD45-. The Papaccio method showed greater cell viability for cells that had been frozen for 30 days (59.5%) than the Kamath method (56.2%), while the Kamath method provided better results for 1 day (65.5%) and 7 days (56%). Post-cryopreservation expression of the markers CD105+/CD34- was greater after 1 and 7 days with the Kamath method and CD105+/CD45- were expressed after all 3 cryopreservation times. There was greater expression of CD73+ in the hDPSCs after 1 and 7 days with the Kamath method, and after 30 days with the Papaccio method. These results suggest that hDPSCs express mesenchymal stem cell markers after cryopreservation. However, cryopreservation time may affect marker expression, probably by altering the spatialconfiguration of cell membrane proteins or by compromising cells at a certain level of differentiation.
Palabras clave
Criopreservación, Células troncales mesenquimales, Fenotipo, Viabilidad celular, Pulpa dental, Medicina regenerativa
Keywords
Cryopreservation, Mesenchymal stem cells, Phenotype, Cell viabilit, Dental pulp, Regenerative medicine