Figure 3: Chondrocytes introduced to the hydrogel region of the composite recellularize microparticles and maintain chondrogenic expression.

(A, B) Chondrocytes are extracted from bovine knee joints, stained with a fluorescent proliferation dye (CFSE), and encapsulated in the hydrogel portion of tissue clay. The number of cells migrating into particles was measured by quantification of CFSE stained cells present within image-thresholded microparticles at 0,2,4,7, and 14 days post seeding. (C) Chondrocyte migration into the microparticles occurs within the first 2 days of culture. (D) Cells were observed migrating into the microparticles within the first 24 hours. (E) Image montaging supports that the migration was observed broadly throughout the tissue clay construct (Figure S2 illustrates cells embedded in particles with a z-stack image). (F) Quantitative RT-PCR of primary chondrocytes seeded in empty constructs, constructs at percolation, and constructs past percolation shows increased chondrogenic expression in tissue clay (HPRT1 used as housekeeping gene, fold change from TCP plated cells). Scale bar = 100 μm (B, D), 1 mm (E). (*p<0.05, RT-qPCR data compared to hydrogel control and ⁺p<0.05, compared to native tissue).