Figure 1.
A) Modified 3DKUBE™ design where the original imaging interface is bored out to extend the depth of the chamber from 8.8 mm to 10.05 mm deep and replaced with No. 1.5 cover glass. This brings the tissue sample closer to the objective in order to accommodate a shorter working distance required for higher magnification levels. B) Photograph of the modified 3DKUBE™.
Figure 2.
Experimental timeline describing cell seeding, induction and imaging time points.
The total number of cells seeded on each scaffold was scaled up by scaffold volume to maintain a consistent density with previous work [15].
Figure 3.
False colored redox maps of adipose tissues in vitro after 0, 1, and 2 wks within the perfusion environment.
The grayscale structures in the images are the silk biomaterial 3D scaffolds, while the sparsely distributed colored objects are cells.
Figure 4.
Changes in cell redox ratio and density with time.
A) Average cellular redox ratio at 0, 1, and 2 wks in perfusion compared to previously published static culture experiments [9]. In both static and perfusion culture, no change in redox was observed between 0 and 1 wks, but the redox ratio decreased (p<0.001) from 1 to 2 wks. Although a significant decrease was observed in both cultures at wk 2, the redox ratio was signficantly higher in the perfusion cultures. B) Cell population measurements per image stack (238×238×40 µm volume) at 0, 1, and 2 wks in perfusion flow demonstrate an increase over time, but substantial sample-to-sample variability.
Figure 5.
H&E stained images taken of the edges and middle of sacrificed adipose samples after 0, 1, and 2 wks in perfusion.
Images were acquired at 10× and 40×. The dark purple structures are part of the silk scaffold, the lighter purple is ECM, and the dark spots within elongated structures are nuclei within cells. The blue arrows and dashed line specify the location of the edges of the tissues. The red arrows point out ECM formation in at the 2 wk time point. 10× scale bars are 500 µm. 40× scale bars are 200 µm.