Microfluidic Perfusion for Regulating Diffusible Signaling in Stem Cells
Figure 3
FGF signaling in perfusion culture.
A. Phase images and close-ups of representative mESC colonies growing under differentiation and self-renewal conditions. Representative images of time-course changes in mESC morphology under two differentiation conditions in perfusion culture (N2B27+FGF4 and N2B27+CM) (left) or under self-renewal conditions in static culture (serum+LIF) (right). B. FGF4 supplementation in perfused mESC differentiation. Growth curves for cells differentiated in N2B27+CM vs. N2B27+FGF4 (5, 20 ng/mL) in perfusion for 5 days. Data shown are average ± s.d. of 2 independent experiments for each FGF4 concentration (* Indicates P<0.05). (C–D) Inhibition of FGF signaling in perfused mESC differentiation cultures. C. Transmission images of cells cultured in perfusion for 6 days in N2B27+CM and N2B27+CM+FGFRi (300 ng/mL) (left). Growth analysis for cells cultured in the presence of FGFRi at 300 ng/mL. Fold increase in cell area after 5 days of perfusion culture for two different conditions, N2B27+CM and N2B27+CM+FGFRi (right). D. Fluorescence images of cells cultured in perfusion for 6 days in N2B27+CM and N2B27+CM+FGFRi (300 ng/mL) (left). Sox1-GFP+ cell frequency assessed by flow cytometry for cells differentiated in N2B27+CM and N2B27+CM+FGFRi condition on Day 6 of perfusion culture (right). Data are average ± s.d. of 3 independent experiments, (*** Indicates P<0.001).