Advanced Bioprinting of Hydrogels with Controlled Mineral Gradients for Regenerative Engineering of the Osteochondral Interface

Abstract

The osteochondral (OC) interface exhibits a mineral gradient in the subchondral bone and articular cartilage interface, varying in thickness by several hundred micrometers across different species. Disruptions to this interface can cause severe damage to OC tissues, leading to osteoarthritis (OA), a debilitating and irreversible condition. Regenerative engineering approaches hold promise for addressing this issue by replicating the natural architecture and composition of native OC interface within a biomaterial scaffold. This study introduces a novel one-step bioprinting process using a twin-screw extruder that facilitates the fabrication of a unitary synthetic graft (USG), which mimics the native OC interfaces mineral concentration gradient. The newly developed USG is composed of an agarose-based cartilage layer and a bone layer, which consists of agarose enriched with 20% hydroxyapatite (w/vol). The USG features a gradient interface with the mineral concentration seamlessly transitioning from 0 to 20wt% from the cartilage to the bone layer. The mineral gradients in the USG and the native tissue were documented using thermogravimetric analysis (TGA), micro-CT, and energy dispersive x-ray (EDX). TGA revealed that the gradient transition length in the graft (647{+/-}21m) compared well to that of native OC tissue (633{+/-}124m) harvested from bovine knee. The strain sweep and frequency sweep tests in oscillatory shear evaluated the linear viscoelastic properties of the grafts, indicating a dominant storage modulus over loss modulus similar to that of native OC tissues. Additionally, the compressive and stress relaxation behaviors of the USGs were quantified using multi-extensional tests, highlighting the grafts ability to maintain structural integrity under mechanical stress. Furthermore, viability assays performed after bioprinting showed that chondrocytes and human fetal osteoblast cells successfully integrated and survived within their designated regions of the graft. The USGs engineered in this study exhibit properties that make them promising candidates for regenerating OC defects and restoring knee joint functionality.