Assessment of a bovine co-culture, scaffold-free method for growing meniscus-shaped constructs

Using a self-assembly (SA), scaffoldless method, five high-density co-cultures with varied ratios of meniscal fibrochondrocytes (MFCs) and articular chondrocytes (ACs) were seeded into novel meniscus-specific, ring-shaped agarose wells. The following ratios of MFCs to ACs were used: 0% MFC, 25% MFC, 50 % MFC, 75 % MFC, and 100 % MFC. Over 4 weeks, all ratios of cells self-assembled into three-dimensional constructs with varying mechanobiological and morphological properties. All groups stained for collagen II (Col II), and all groups except the 0% MFC group stained for collagen I (Col I). It was found that the tensile modulus was proportional to the percentage of MFCs employed. The 100% MFC group yielded the greatest mechanical stiffness with 432.2 +/- 47 kPa tensile modulus and an ultimate tensile strength of 23.7 +/- 2.4 kPa. On gross inspection, the 50 % MFC constructs were the most similar to our idealized meniscus shape, our primary criterion. A second experiment was performed to examine the anisotropy of constructs as well as to directly compare the scaffoldless, SA method with a poly-glycolic acid (PGA) scaffold-based construct. When compared to PGA constructs, the SA groups were 2-4 times stiffer and stronger in tension. Further, at 8 weeks, SA groups exhibited circumferential fiber bundles similar to native tissue. When pulled in the circumferential direction, the SA group had significantly higher tensile modulus (226 76 kPa) than when pulled in the radial direction (67 +/- 32 kPa). The PGA constructs had neither a directional collagen fiber orientation nor differences in mechanical properties in the radial or circumferential direction. It is suggested that the geometric constraint imposed by the ring-shaped, non-adhesive mold guides collagen fibril directionality and, thus, (a)lters mechanical properties. Co-culturing ACs and MFCs in this manner appears to be a promising new method for tissue engineering fibrocartilaginous tissues exhibiting a spectrum of mechanical and biomechanical properties.