Calcified cartilage tissue engineering: a comparison of hydroxyapatite and β-tricalcium phosphate bioinks

Abstract

654 million people worldwide suffer from knee osteoarthritis, resulting in a reduced quality of life for patients and a burden on the healthcare system. Tissue engineering strategies represent a promising alternative to current surgical interventions in the repair of osteochondral tissue. Tissue engineering strategies have neglected the critical role of the calcified cartilage zone in providing a mechanically competent interphase between soft viscoelastic cartilage and stiff bone. This results in bi-phasic osteochondral scaffolds failing due to phase delamination and dislodgement. The aim of the present study is to propose a new bioink for engineering calcified cartilage tissue. Our study presents a comparison between a hydroxyapatite (HA)-based biomaterial ink and a β-tricalcium phosphate (β-TCP)-based biomaterial ink. While solvent-based extrusion printing has been used to fabricate HA- PCL (hydroxyapatite- polycaprolactone) scaffolds, to our knowledge, this technique has not been used for the fabrication of β-TCP-PCL scaffolds. Solvent-based extrusion printing can be performed at room temperature avoids the disadvantages associated with printing at high temperatures. The hypothesis is that β-TCP-PCL will induce chondrocyte hypertrophy and faster deposition of a mineralized calcified cartilage layer due to the higher solubility of β-TCP compared to HA. Our study is the first to show that the novel bioink composed of β-TCP-PCL can be processed with solvent-based extrusion printing in the same way as HA-PCL. The biomaterial inks with the highest ceramic concentrations (70%HA- 30% PCL and 70% β-TCP- 30% PCL) have the highest yield stress and consequently best shape fidelity, while printing resolution is similar with a minimum interfibre space of 0.5-1 mm for all concentrations of both groups. Solvent based extrusion printing can be used to print centimeter sized porous scaffolds with both materials (70%HA- 30% PCL and 70% β-TCP -30% PCL). Macropores were in the range of 300-900 um, which facilitates to bone ingrowth. We found that PCL does not mask the ceramic phase and there is a higher release of calcium and phosphorous ions from the β-TCP material. The in vitro experiments showed that cells remained viable and metabolically active over 28 days. In terms of the production of matrix components, small amounts of glycosaminoglycans were produced. Collagen type II was not produced by any of the groups, while Collagen type I was produced by all groups. It is possible that the chondrocytes de-differentiated and consequently produce fibrocartilage. However, to investigate whether the ceramic ions induce chondrocyte hypertrophy and the deposition of a mineralized matrix, a Collagen type X staining and ALP assay should be performed. Therefore, further investigation is required to truly conclude whether the higher soluble β-TCP results in the faster production of calcified cartilage.