IDG-SW3 Osteocyte Differentiation and Bone Extracellular Matrix Deposition Are Enhanced in a 3D Matrix Metalloproteinase-Sensitive Hydrogel

Osteocytes reside within a heavily mineralized matrix, making them difficult to study in vivo and to extract for studies in vitro. IDG-SW3 cells are capable of producing a mineralized collagen matrix and transitioning from osteoblasts to mature osteocytes, thus offering an alternative to study osteoblast to late osteocyte differentiation in vitro . The goal for this work was to develop a 3D degradable hydrogel to support IDG-SW3 differentiation and deposition of bone extracellular matrix. In 2D, the genes Mmp2 and Mmp13 increased during IDG-SW3 differentiation and were used as targets to create a matrix metalloproteinase (MMP)-sensitive poly(ethylene glycol) hydrogel containing the peptide cross-link GCGPLG-LWARCG and RGD to promote cell attachment. IDG-SW3 differentiation in the MMP-sensitive hydrogels improved over nondegradable hydrogels and standard 2D culture. Alkaline phosphatase activity at day 14 was higher, Dmp1 and Phex were 8.1-fold and 3.8-fold higher, respectively, and DMP1 protein expression was more pronounced in the MMP-sensitive hydrogels compared to nondegradable hydrogels. Cell-encapsulation density (cells/mL of precursor) influenced the formation of dendrite-like cellular processes and mineral and collagen deposition, with 80 × 10 6 cells/mL of precursor performing better than 2 × 10 6 or 20 × 10 6 cells/mL of precursor, while connexin 43 was not affected by cell density. The cell density effects were more pronounced in the MMP-sensitive hydrogels over nondegradable hydrogels. This study identified that high cell encapsulation density and hydrogels susceptible to cell-mediated degradation enhanced the mineralized collagen matrix and osteocyte differentiation. Overall, a promising hydrogel is presented that supports IDG-SW3 cell maturation from osteoblasts to osteocytes in 3D.