Page 360 - IJB-9-2

International Journal of Bioprinting Laser bioprinting of hiPSC-derived neural stem cells and neurons

Figure 2. Cell viability, colony morphology, development of dendrites. (A and B) Fluorescent microscopic images of live/dead-staining 24 hours after
printing for NSCs, d5, d10, and d20 neurons with calcein AM (green, viable cells) and ethidium homodimer-1 (red, dead cells). Live/dead staining revealed
agglomeration of printed cells, increasing with duration of pre-differentiation. Dead cells could be seen within these agglomerations in high-magnification
images (B). (C) Left: Viability 24 hours post-printing of printed, donor, and control cells (NSCs, d5, d10, and d20 neurons), given in percent. Control cells
were not in contact to bioink or substrate, but stored in a vial, while printed and donor cells were suspended in bioink for printing. Obviously, viability
decreases with duration of pre-differentiation period and viability of printed cells is mostly higher than that of donor and control cells. Right: Statistical
analysis, P values for unpaired two-sample t-test. Statistical significance (highlighted in red) was determined for higher viability of printed cells when com-
pared to control and donor cells (five out of eight cases) and for decreased viability with increasing pre-differentiation period (15 out of 18 cases). (D and
E) Formation of neuronal dendrites in smaller (D) and larger (E) droplets of bioink with lower and higher number of NSCs printed on Matrigel substrate
TM
is depicted in phase contrast imaging 1 day post-printing and in fluorescence imaging with marker β3-tubulin and Hoechst 33342 at day 7 of post-printing
differentiation. The number of neuronal dendrites relative to the number of neuronal cells is lower in smaller droplets and higher in larger droplets. Scale
bars are 1 mm (D and E, except for right column), 500 µm (A) and 100 µm (all others).

Volume 9 Issue 2 (2023) 352 https://doi.org/10.18063/ijb.v9i2.672