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Figure 4. Culturing and differentiation of fetal neural stem cells (NSCs). Phase contrast pictures of live cells in culture and confocal images of fixed and immunostained cells. A and B NSCs were grown in floating conditions where they formed clusters of cells (A) and secondary cultures could be formed after dissociation and re-plating (B). C NSCs expressed stem cell markers Nestin and SOX2. D-G Expression of GFAP and βIII-tubulin was low in NSCs. H-O After differentiation with 5% FCS for 7 or 14 days, strong immunoreactivity for βIII-tubulin and GFAP was visible. Scale bar in B: 100 μm (A-B); scale bar in C, N, O: 50 μm (C-O).
ferentiation in NSCs was investigated using staining for βIII-tubulin and GFAP. The βIII- tubulin/total cell count ratio was increased after transfection with miR146a (p=0.0043) and miR147b mimics (p=0.0022, Fig 5 A-M). In order to elucidate possible mecha- nisms, mRNA expression levels of several potential targets were examined (Fig 5 N-O). Transfection with miR146a mimic led to decreased expression of Notch homolog 1, trans- location-associated protein (NOTCH1, p=0.0260), miR147b mimic transfection decreased the expression of both NOTCH1 (p=0.0152) and Janus kinase 2 (JAK2, p=0.0173). We also