3D-PRINTED OSTEOCHONDRAL GRAFTS AND THEIR CHARACTERIZATION

Abstract

The osteochondral (OC) interface is a complex tissue with a hierarchical structure found at the ends of the bones of the knee joint consisting of a layer of soft tissue (cartilage) overlaying hard tissue in the subchondral bone. It exhibits a gradient of its constituents, especially in terms of mineral concentration, cell phenotype, collagens, and glycosaminoglycans, with a thickness of around 0.5 mm. The tidemark, a critical yet often overlooked component of OC interface tissue, plays a pivotal role in maintaining tissue function by acting as a barrier against vascular invasion of the cartilage. Fabricating scaffolds that mimic the complex physiology and functionalities of the OC tissue within the physiological thickness remains a challenge. This study aimed at fabricating a unitary composite scaffold that is similar of the OC interface in terms of distribution of its mineral content. It was hypothesized that the interface formed between the layers of the multilayer graft will possess a thickness of hydroxyapatite (HAP) gradient similar to that seen at the native rabbit OC tissue. To test the hypothesis, a multilayer composite OC graft was fabricated using gelatin and oxidized alginate (OXA) compositions with and without HAP for the bone and cartilage regions, respectively, and a gradient of HAP was formed in between. The two layers were formed using a 3D bioprinting method, while a porous electrospun mesh of polycaprolactone was placed in the graded region between cartilage and bone to represent the tidemark. The change in mineral content across the rabbit OC interface tissue and the OC graft interface was investigated using energy dispersive X-ray (EDX) and micro computed tomography (CT) characterization. The printability of the bioinks was verified by a strain sweep test, and volumetric expansion of both inks, with and without HAP, was examined using a swelling test. Findings revealed that both bioinks exhibited a shear thinning behavior. In addition, swelling test showed that both inks possessed similar volumetric expansion when immersed in water, demonstrating its feasibility to be used as a defect filler. EDX scan for calcium (Ca) and phosphorus (P) verified the gradient of mineral in both OC grafts and native rabbit OC tissue. The CT characterization verified a HAP gradient created in the OC graft within 168m thickness similar to the mineral gradient thickness determined for rabbit OC interface. Furthermore, the electrospun membrane was found to have pore diameters less than 1m that is sufficient to prevent vascular invasion of the articular cartilage tissue. Overall, the OC graft fabricated using combined bioprinting and electrospinning techniques demonstrated a potential to serve as a biomimetic hydrogel filler for regenerating OC defects to restore the function of the knee joint. It is expected that the proposed OC graft will be effectively used to address a significant clinical problem that affects millions of people, with significant societal and economic impacts.