Production of Hyaline-like Cartilage by Bone Marrow Mesenchymal Stem Cells in a Self-Assembly Model

Abstract

A scaffoldless or self-assembly approach to cartilage tissue engineering has been used to produce hyaline cartilage from bone marrow–derived mesenchymal stem cells (bMSCs), but the mechanical properties of such engineered cartilage and the effects the transforming growth factor (TGF) isoform have not been fully explored. This study employs a cell culture insert model to produce tissue-engineered cartilage using bMSCs. Neonatal pig bMSCs were isolated by plastic adherence and expanded in monolayer before being seeded into porous transwell inserts and cultured for 4 or 8 weeks in defined chondrogenic media containing either TGF-β1 or TGF-β3. Following biomechanical evaluation in confined compression, colorimetric dimethyl methylene blue and Sircol dye–binding assays were used to analyze glycosaminoglycan (GAG) and collagen contents, respectively. Histological sections were stained with toluidine blue for proteoglycans and with picrosirius red to reveal collagen orientation, and immunostained for detection of collagen types I and II. Neocartilage increased in thickness, collagen, and GAG content between 4 and 8 weeks. Proteoglycan concentration increased with depth from the top surface. The tissue contained much more collagen type II than type I, and there was a consistent pattern of collagen alignment. TGF-β1–treated and TGF-β3–treated constructs were similar at 4 weeks, but 8-week TGF-β1 constructs had a higher aggregate modulus and GAG content compared to TGF-β3. These results demonstrate that bMSCs can generate functional hyaline-like cartilage through a self-assembling process.