Potential of chondrogenesis of bone marrow stromal cells co-cultured with chondrocytes on biodegradable scaffold: in vivo experiment with pigs and mice

Abstract

To explore the feasibility of in vivo chondrogenesis of bone marrow stromal cells (BMSCs) co-cultured with chondrocytes on biodegradable scaffold. Porcine BMSCs were isolated, expanded and labeled with enhanced green fluorescent protein (EGFP), and then were mixed with articular chondrocytes isolated from porcine knee joint at the ratio of 1:1. The mixed cells were seeded onto polyglycolic acid (PGA) scaffold at the ultimate concentration of 5.0 x 10(7)/ml (co-culture group). Pure chondrocytes and BMSCs of the same ultimate concentration were seeded respectively onto the scaffold as positive control group and negative control group. After two weeks' culture in vitro, they were planted subcutaneously into nude mice respectively. These specimens were collected after in vivo implantation for 8 weeks to undergo microscopy. Laser confocal microscopy was used to observe the distribution of EGFP-labeled cells in the tissue. RT-PCR was used to examine the expression of collagen type II and aggrecan. Immunohistochemistry was used to observe the protein expression of collagen type II. The cell-scaffold constructs of the co-culture group and positive control group, could maintain the original size and shape no matter in vitro or in vivo. After 8 weeks' in vivo implantation, the constructs in both co-culture group and positive control group formed cartilage-like tissue with typical histological structure and extracellular matrix staining similar to those of the normal cartilage. The GAG content and compressive modulus of the co-culture group reached over 80% of those of the positive control group. Confocal microscopy revealed the presence of EGFP-labeled cells in the engineered cartilage lacuna. Histological examination showed that the constructs of the negative control group shrunk gradually after in vivo implantation with no typical cartilage-like tissue formation. In vitro co-cultured BMSC-chondrocyte-PGA constructs have the potential to form mature cartilage-like tissue in subcutaneous non-chondrogenesis environment, indicating that chondrocytes still provide enough signals for BMSC chondrogenic differentiation.