Discrimination of primitive endoderm in embryoid bodies by Raman microspectroscopy
- 283 Downloads
Embryoid bodies (EBs), derived from aggregated embryonic stem (ES) cells, are capable of differentiating into all three germ layers, including the endoderm, mesoderm, and ectoderm. The initial stage of EB differentiation is the formation of a primitive endoderm (PE) layer located at the periphery of the aggregate. Raman microspectroscopy was employed to segregate PE cells from undifferentiated ES cells. The Raman spectra of the PE cells of the periphery of EBs, formed upon the withdrawal of leukemia inhibitory factor (LIF), were compared with those of the undifferentiated ES cells of the core of cell aggregates, formed in the presence of LIF. It was noticed that the PE cells have high contents of proteins and low contents of nucleic acids, lipids, and carbohydrates compared with ES cells. Also, we established the presence of another population of PE cells located in the core of the EBs. In addition, we identified some specific Raman markers to distinguish PE cells from ES cells (e.g., I 1003/I 937). This is the first study to investigate the PE cells of live EBs and define some Raman markers to distinguish them from undifferentiated ES cells.
KeywordsEmbryoid bodies Differentiation Primitive endoderm Raman microspectroscopy Glycogen
This work was supported by Special Coordination Funds for Promoting Science and Technology/Special Grants for Projects Promoting Science and Technology through Regional Industry–Academia–Government Cooperation and by Special Coordination Funds for Promoting Science and Technology, Ministry of Education, Culture, Sports, Science and Technology, Japan.
- 2.Beddington RS, Robertson EJ (1989) Development 105:733–737Google Scholar
- 6.Doetschman TC, Eistetter H, Katz M, Schmidt W, Kemler R (1985) J Embryol Exp Morphol 87:27–45Google Scholar
- 8.Keller G, Kennedy M, Papayannopoulou T, Wiles MV (1993) Mol Cell Biol 13:473–486Google Scholar
- 10.Hopfl G, Gassmann M, Desbaillets I (2004) Methods Mol Biol 254:79–98Google Scholar
- 18.Notingher I, Verrier S, Romanska H, Bishop AE, Polak JM, Hench LL (2002) Spectrosc-Int J 16:43–51Google Scholar