Chondrogenesis of adipose stem cells in a porous PLGA scaffold impregnated with plasmid DNA containing SOX trio (SOX-5,-6 and -9) genes.

摘要

We developed a chondrogenic scaffold system in which plasmid DNA (pDNA) containing SOX trio (SOX-5, -6, and -9) genes was incorporated into a PLGA scaffold and slowly released to transfect adipose stem cells (ASCs) seeded in the scaffold. The purpose of this study was to test the in vitro and in vivo efficacy of the system to induce chondrogenic differentiation of ASCs. The pDNA/PEI-PEG complex-incorporated PLGA/Pluronic F127 porous scaffolds were fabricated by a precipitation/particulate leaching method. The following five kinds of pDNA were incorporated into the scaffolds: 1) pECFP-C1 vector without an interposed gene (control group); 2) SOX-5 plasmids; 3) SOX-6 plasmids; 4) SOX-9 plasmids; and 5) one-third doses of each plasmid (SOX-5, -6, and -9). ASCs were seeded on pDNA-incorporated PLGA scaffolds and cultured in chondrogenic media for 21 days. ASCs were also isolated from rabbits, seeded in pDNA-incorporated PLGA scaffolds, and then implanted in the osteochondral defect created on the patellar groove. The rabbits were sacrificed and analyzed grossly and microscopically 8 weeks after implantation. The percentage of transfected cells was highest on day 14, around 70%. After 21 days, PLGA scaffolds incorporated with each gene showed markedly increased expression of the corresponding gene and protein. Glycosaminoglycan (GAG) assay and Safranin-O staining showed an increased proteoglycan production in SOX trio pDNA-incorporated scaffolds. The COL2A1 gene and protein were notably increased in SOX trio pDNA-incorporated scaffolds than in the control, while COL10A1 protein expression decreased. Gross and histological findings from the in vivo study showed enhanced cartilage regeneration in ASCs/SOX trio pDNA-incorporated PLGA scaffolds.