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International Journal of Bioprinting Curved cell-guided structures printed by FDM

Figure 2. Cell proliferation on the patterns with varying curvature. (A) Curved structures for cell culture and proliferation test (scale bar = 10 mm).
(B) The proliferation of M-22 cells (CCK-8 assay) after 1, 2, and 4 days of culture on the curved structures. The significant difference between the groups
was indicated (*P < 0.05, **P < 0.01).
3. Results ratio corresponds to the more rounded shape of the cells.
The curved channels used to observe the morphology of
3.1. Cell proliferation on curved structures cells has the same length (500π μm), width (100 μm), and
The CCK-8 test assessed cell proliferation. The structures depth (150 μm) (Figure 3B). The straight channel had the
for cell culture and proliferation test were the entire circle same width and depth as the curved groups (Figure 3C).
spreading over the entire glass sheet with the same perimeter The effect of pattern curvature on the cell morphology
but varying radii (Figure 2A). The cells proliferated well and orientation was evaluated after 2 days of cell seeding
in all groups, indicating that the printed pattern materials by analyzing the microscopic fluorescence images
and glass substrate were not toxic to cells (Figure 2B). The (Figure 3D). The aspect ratio of cells first declined and
cell proliferation first increased and then decreased as the then increased as the guided radius increased, and reached
channel radius increased and was significantly faster in the minimum in group R2. The cells were elongated in the
group R2 than that in the other groups on day 2 (P < 0.01) groups R1.5, R2.5, and R3 but shortened in the groups SL
and day 4 (P < 0.05). The proliferation of groups R1.5, R2.5, and R2 compared to the control (Figure 3E). Specifically,
and R3 showed no significant differences on day 2 and day the elongation of cells in R3 was significantly larger than
4. The proliferation in straight lines was approximately that in SL (P < 0.01), R2 (P < 0.01), and control (P < 0.05).
equal to the proliferation in control on day 2 and day 4. The elongation of cells in R1.5 was significantly greater
Notably, the cells proliferated faster in all curved groups than in SL (P < 0.01). Interestingly, the aspect ratio at R2
than in the straight lines and control on day 4, where was closest to the control.
groups R2 and R3 were both significantly faster than the
straight lines and control (P < 0.05), and group R2.5 was The orientation angle was employed to characterize cell
considerably faster than the control (P < 0.05). orientation, which was defined as the acute angle formed
by the major axis of the cell fitting ellipse and the tangent
3.2. Cell morphology and orientation in curved line on the pendant foot of the nucleus to the channel
channels (Figure 3F). Cells were more oriented when the orientation
Cell morphology was characterized by aspect ratio, which angle was smaller. The distribution of cell orientation angle
was defined as the ratio of the major axis to the minor axis was uniform in the control, concentrated within 60° in the
of the cell fitting ellipse (Figure 3A). The smaller aspect straight channels, and dispensed within 30° in the curved

Volume 9 Issue 3 (2023) 42 https://doi.org/10.18063/ijb.681