Research Article

Comparative Analysis of Spontaneous and Stimulus-Evoked Calcium Transients in Proliferating and Differentiating Human Midbrain-Derived Stem Cells

Figure 5

Immunocytochemical characterization of proliferating human midbrain-derived stem cells. Immunostaining of cells during, (1) standard propagation (upper panel) in a medium with epidermal growth factor (EGF) and basic fibroblast growth factor (bFGF) and (2) after removal of EGF/bFGF and one-day exposure to fibroblast growth factor 8 (FGF8) (lower panel), which represents the initial step of the induced neuronal differentiation (sequential addition of various factors, see Materials and Methods for details). To visualize all cells, cultures were immunostained using an antibody against human nuclei (HN). Almost all cells in both experimental groups expressed Ki67, a marker of dividing cells. Moreover, a very large proportion of the cells expressed the intermediate filament protein nestin, a marker of neural progenitor cells. Only very few cells had spontaneously differentiated into β-tubulin III- (β-tub III-) positive or doublecortin- (DCX-) positive immature neurons in the EGF/bFGF group, whereas some β-tub III-positive and DCX-positive neuronal cells were seen after short-term FGF8 treatment. Scale bar = 20 μm.