Fig 1.
Micro and SEM images of the printed and template casted scaffolds at low (a, b) and high magnification (c, d) and at day 5 (e-h). Day 5 images show visual alignment of SCs on the printed scaffold. Arrows indicate SCs on scaffolds. Scale bar on a, b = 2mm; on c, d = 500μm; on e, g = 100μm; on f, h = 25μm.
Fig 2.
Live/ Dead cytotoxicity viability assay at days 3 (a, b, c), 7 (d, e, f), and 14 (g, h, i).
Staining also suggested directed growth of the cells on the 3D-printed scaffold in parallel with the struts. Staining highlights pattern of clustered growth on the scaffolds (e, f, h, i). Scale bar = 100μm.
Fig 3.
Immunostaining for p75LNGFR(green) and S100-β (red) for days 7 (a-c) and 14 (d-e).
Panels are divided horizontally by group and vertically by stain. The final column is a co-stain of DAPI, S100-β, and p75LNGFR. Majority of cells stain positive for both SC markers suggesting the scaffolds do not significantly influence cell character. Scale bar = 100μm.
Fig 4.
Results show the percentage of cells in a given random viewing field within a certain degree of parallel with the major scaffold struts. Data is from DAPI images taken at 7 (a) and 14 (b) days from 4 different fields per time point. Fields analyzed were different than those in Fig 3. Schematic (c-d) shows the method of measuring the angle between the cell axis in relation to strut. X marks the location of the cell being measured. A line was draw through the major axis of the cell’s nucleus and the angle this line formed with the indicated strut was recorded. The shaded region denoted by Z indicates the top facing regions of the strut from which nuclei were measured.
Fig 5.
Graph shows cell number at each time point. Starred bars designate significant differences between groups within a single time point (n = 3).
Fig 6.
Four genes were analyzed: ngf-β (a), nt–3 (b), vegf-a (c), and pdgf-bb (d) with GAPDH used as a house-keeping gene. Data is shown as expression relative to GAPDH at 7 and 14 days. Starred bars specify significant differences between groups.
Fig 7.
ELISA Assays for β-NGF (a) and VEGF-A (b).
Protein concentration per cell in the growth media at days 3, 7, and 14, was normalized using the cell numbers from the dsDNA (Fig 5) results. Data collected from three different samples at each time point. Starred bars indicate significant difference between groups at each time point. Significant differences between time points not shown.