Figure 1
Illustration of the agarose-scaffolded alginate bead culture for the 3D static compression model and timeline for experiments. (a) Procedure for the agarose-scaffolded bead culture for compression. (A) Agarose molding. Twenty-five milliliter of the low-melting agarose containing DMEM/F12, 10% FBS, and 1% PS was poured into a 100 mm tissue culture plate. To embed the CAF-alginate beads, the agarose gel was punched using a sterile 15 ml tube. (B) Alginate bead embedding. A thin layer of agarose was made on the bottom of a punched hole. The cell-alginate beads pre-cultured for ECM deposition a day before embedding were laid on the agarose layer. The rest part around the cells-alginate beads was filled with agarose gel. (C) Loading static compression. Compression was loaded using the cube filled with iron ball bearings. An empty cube was loaded for control. (b) Timeline for the experiments. The samples for experiments were generally obtained in the same procedure except for histological analysis and function validation. For histological analysis, the alginate beads containing cells were fixed with 4% paraformaldehyde, embedded in paraffin, sectioned into 5-μm-thick sections, and stained with hematoxylin and eosin. For function validation, inhibitor treatment was started 1 days before cell harvest. For the sample preparation of decompressed cells, the cells were incubated for 1 day without compression
