RESUMEN
In culture, human pluripotent stem cells (PSCs) are phenotypically (for instance, the SSEA3 expression level) and functionally (capacity to survive after single-cell dissociation) heterogeneous. We report here that the side scatter (SSC) signal measured by flow cytometry, a variable correlated with membrane irregularity and cell granularity, is very high in PSCs, even higher than in blood polymorphonuclear cells, and markedly heterogeneous. Moreover, SSC intensity rapidly and strongly decreases upon PSC differentiation into any of the three germ layers. PSCs with high SSC (HSSC cells) or low SSC (LSSC cells) values both express pluripotency markers, but HSSC cells are characterized by more frequent simultaneous expression of the membrane pluripotency factors SSEA3, SSEA4, TRA-1-81, TRA-1-60, and CD24 and by a higher mitochondrial content. Functionally, HSSC cells are more likely to generate colonies upon single-cell passage than LSSC cells. SSC monitoring might provide a simple, but robust and rapid method to estimate pluripotency variations in culture and unveils a new phenotypic and functional heterogeneity in PSCs.
Asunto(s)
Células Madre Embrionarias/citología , Heterogeneidad Genética , Estratos Germinativos/citología , Células Madre Pluripotentes/citología , Animales , Antígenos de Superficie/genética , Antígenos de Superficie/metabolismo , Biomarcadores/metabolismo , Antígeno CD24/genética , Antígeno CD24/metabolismo , Diferenciación Celular , Línea Celular , Células Clonales , Células Madre Embrionarias/metabolismo , Citometría de Flujo , Expresión Génica , Estratos Germinativos/metabolismo , Humanos , Ratones , Ratones SCID , Células Madre Pluripotentes/metabolismo , Proteoglicanos/genética , Proteoglicanos/metabolismo , Antígenos Embrionarios Específico de Estadio/genética , Antígenos Embrionarios Específico de Estadio/metabolismo , Teratoma/metabolismo , Teratoma/patologíaRESUMEN
Human pluripotent stem cells (PSCs), encompassing embryonic stem cells and induced pluripotent stem cells, proliferate extensively and differentiate into virtually any desired cell type. PSCs endow regenerative medicine with an unlimited source of replacement cells suitable for human therapy. Several hurdles must be carefully addressed in PSC research before these theoretical possibilities are translated into clinical applications. These obstacles are: (1) cell proliferation; (2) cell differentiation; (3) genetic integrity; (4) allogenicity; and (5) ethical issues. We discuss these issues and underline the fact that the answers to these questions lie in a better understanding of the biology of PSCs. To contribute to this aim, we have developed a free online expression atlas, Amazonia!, that displays for each human gene a virtual northern blot for PSC samples and adult tissues (http://www.amazonia.transcriptome.eu).