RESUMEN
We employed Fourier transform infrared (FTIR) microspectroscopy to investigate the effects of different tissue culture environments on the FTIR spectra of undifferentiated human embryonic stem cells (hESCs) and their differentiated progeny. First we tested whether there were any possible spectral artifacts resulting from the use of transflectance measurements by comparing them with transmission measurements and found no evidence of these concluding that the lack of any differences resulted from the homogeneity of the dried cytospun cellular monolayers. We found that hESCs that were enzymatically passaged onto mouse embryonic fibroblasts (MEFs) in KOSR based hESC medium, hESCs enzymatically passaged onto Matrigel in mTESR medium and hESCs mechanically passaged onto MEFs in KOSR-based hESC medium, possessed unique FTIR spectroscopic signatures that reflect differences in their macromolecular chemistry. Further, these spectroscopic differences persisted even upon differentiation towards mesendodermal lineages. Our results suggest that FTIR microspectroscopy is a powerful, objective, measurement modality that complements existing methods for studying the phenotype of hESCs and their progeny, particularly changes induced by the cellular environment.
Asunto(s)
Diferenciación Celular , Medios de Cultivo Condicionados/farmacología , Células Madre Embrionarias/citología , Fibroblastos/citología , Espectroscopía Infrarroja por Transformada de Fourier/métodos , Animales , Linaje de la Célula , Células Cultivadas , Colágeno/metabolismo , Análisis Discriminante , Combinación de Medicamentos , Células Madre Embrionarias/efectos de los fármacos , Fibroblastos/efectos de los fármacos , Humanos , Laminina/metabolismo , Análisis de los Mínimos Cuadrados , Ratones , Microscopía de Fuerza Atómica , Fenotipo , Proteoglicanos/metabolismoRESUMEN
Fourier transform infrared (FTIR) microspectroscopy shows potential as a benign, objective and rapid tool to screen pluripotent and multipotent stem cells for clinical use. It offers a new experimental approach that provides a holistic measurement of macromolecular composition such that a signature representing the internal cellular phenotype is obtained. The use of this technique therefore contributes information that is complementary to that acquired by conventional genetic and immunohistochemical methods.
Asunto(s)
Células Madre Multipotentes/química , Células Madre Pluripotentes/química , Diferenciación Celular , Análisis por Conglomerados , Análisis Discriminante , Humanos , Análisis de los Mínimos Cuadrados , Células Madre Multipotentes/citología , Células Madre Pluripotentes/citología , Análisis de Componente Principal , Espectroscopía Infrarroja por Transformada de FourierRESUMEN
Fourier transform infrared (FTIR) microspectroscopy was employed to elucidate the macromolecular phenotype of human embryonic stem cells (hESCs) and human induced pluripotent stem cells (hiPSCs) and their differentiated progeny. Undifferentiated hESCs and hiPSC lines were found to be not clearly distinguishable from each other. However, although both hESC and hiPSC variants appeared to undergo similar changes during differentiation in terms of cell surface antigens, the derived cell types from all cell lines could be discriminated using FTIR spectroscopy. We foresee a possible future role for FTIR microspectroscopy as a powerful and objective investigative and quality control tool in regenerative medicine.