Tissue engineering via 3D bioprinting offers a novel solution to treating large bone voids. The optimum bioink for bone tissue engineering is unclear. The purpose of this study was to investigate the suitability of an extrusion-based 3D bioink composed of gelatin methacryloyl (GelMA), gelatin, hydroxyapatite (HA), demineralized bone matrix (DBM) and osteoblasts for bone tissue engineering.
A mouse calvarial osteoblast-laden GelMA-gelatin-HA bioink consisting of various concentrations of DBM was 3D-bioprinted into porous hydrogel constructs. The 3D-fabricated constructs were cross-linked via photopolymerization and cultured in osteogenic medium. After 1, 14, and 28 days, a cohort of constructs were analyzed. The water weight percent differences of the hydrogels were characterized following fabrication along with the degradation behavior in standard culture medium for 28 days. Cell survivability and proliferation was determined.
The addition of DBM to the bioink significantly decreased water content (%) from baseline GelMA-Gelatin-HA hydrogels with a significant trend of decreased swelling with increasing DBM content. The addition of 40, 80, and 120 mg/ml of DBM significantly reduced hydrogel swelling ratios (p ≤ 0.0001). DBM decreased hydrogel breakdown in a concentration dependent manner. Alamar Blue assay demonstrated significantly increased cell proliferation across groups. There was a slight trend of increasing metabolic activity with increasing DBM content. Live/dead staining at 1, 14, and 28 days after printing showed the majority of the osteoblasts were alive at each time-point.
Addition of DBM significantly decreased hydrogel swelling and suggests DBM to be essential in construct stability. DBM may also be altered to modulate the rate of degradation in vitro. Additionally, Alamar Blue results indicate that DBM addition to GelMA-Gelatin-HA bioink did not adversely affect cell viability and cells were able to survive the bioprinting process. These findings support GelMA-gelatin-HA-DBM as a viable bioink for bone tissue engineering.