Comparison of neuropoietic activity of SB623 and MSCs in cocultures with rat embryonic neural cells. (A) Rat neural cells were grown in the presence or absence of MSCs or SB623 (rat/human cell ratio was 10:1) and immunostained for rat nestin (upper panel) and GFAP (middle panel) on day 5, or for CNP on day 12. Nuclei were stained with DAPI (B) An example of microplate neuropoiesis assay data: a comparison of rat neural differentiation marker induction in cocultures of rat cells with MSCs and SB623 from Donor A. Rat neural cells (5,000/well) were cocultured with 500, 250, and 125 cells/well of either MSCs or SB623; and expression of rat-specific nestin, GFAP, and CNP, and human-specific GAP was quantified by using qRT-PCR. Stimulation with 10% MSC-CM and no stimulation (“No add”) were used as positive control and background, respectively. Relative units correspond to standard samples used in qPCR run. Error bars represent SD of biologic duplicates. (C) Neuropoietic coefficients of MSCs and SB623 from Donor A were calculated based on data presented in (B) by first subtracting the background expression of a corresponding neural marker and then normalizing the expression of the neural marker to the human GAP for each number of human cells per well, followed by averaging normalized values. Error bars represent SD from three normalized values.