Investigation of cell proliferation in adult human NSCs during expansion. The portion of NSCs with potential to self renewal expanded in culture medium supplemented with EGF and FGF-2 or CSF without additional growth factors was determined 24 and 72 hours after medium change. A, Percentage of Ki67+ cells after 24 h, number of BrdU+ cells per cm2 after 72 h. Values represent the mean ± S.E.M from at least three independent experiments. *indicates P < 0.05 and ** P < 0.01 compared to each corresponding value in CSF. B, Representative microphotographs demonstrating the influence of CSF on proliferation markers in NSCs. For immunostainings cells were stained with anti-BrdU (upper panel) and anti-Ki67 (lower panel) and anti-Nestin (upper and lower panel). Nuclei were counterstained with DAPI (lower panel). Altered morphology of neurospheres with CSF treatment (right panel) is most likely due to the beginning differentiation process. Scale bar = 50 μm.