Cartilage repair in a rat model of osteoarthritis through intraarticular transplantation of muscle-derived stem cells expressing bone morphogenetic protein 4 and soluble Flt-1

Osteoarthritis (OA), a chronic degenerative joint disorder with worldwide impact, is characterized by articular cartilage destruction and osteophyte formation. OA affects _40 million individuals in the US alone and influences more lives than any other musculoskeletal condition (1). Since articular cartilage is a tissue type that is poorly supplied by blood vessels, nerves, and the lymphatic system, it has a very limited capacity for repair after injury. Although several therapies have been used for OA, no widely accepted treatments have been established, with the exception of arthroplasty. For this reason, tissue engineering techniques aimed at repairing articular cartilage have been extensively studied, and chondrocyte transplantation has already been performed (2–4). Currently, the most effective treatment for OA, besides arthroplasty, is autologous chondrocyte transplantation. However, this treatment has several limitations, including the need to use neighboring healthy donor cartilage, difficulty in treating large-scale defects, limited expansion capacity of primary chondrocytes, and the need for a periosteal patch to maintain engineered cartilage. In addition, in most cases only 30–40% of the defect regenerates articular cartilage, with the remaining defect being filled with fibrocartilage (5,6). In light of these limitations, it is important to find other sources of cells that are abundant and capable of chondrogenic differentiation. Muscle stem cells are more attractive than primary chondrocytes because of their superior capacity for self-renewal, proliferation, and survival following environmental stress (7–9). Recently, stem cell–based therapies have been used clinically for cartilage repair (10,11). The results of several previous studies, including those using muscle-derived stem cells (MDSCs), have indicated that stem cells can undergo chondrogenesis and repair articular cartilage in experimental cartilage injury models (12–15). We previously demonstrated that bone morphogenetic protein 4 (BMP-4)–transduced MDSCs improved cartilage formation in an in vitro pellet culture and regeneration in an in vivo cartilage defect model (13). Based on those results, the present study was designed to clarify the therapeutic efficacy of BMP-4–transduced MDSCs in OA. The control of angiogenesis during chondrogenic differentiation is one of the most important issues affecting the application of stem cells for cartilage repair. Among angiogenesis-modulating factors, including antiangiogenic factors such as troponin 1 (16) and chondromodulin 1 (17), vascular endothelial growth factor (VEGF) is an important mediator of angiogenesis (18). VEGF stimulates capillary formation in vivo and exerts direct mitogenic actions on various cells in vitro (19). In the growth plate, VEGF has been reported to play an essential role in cartilage vascularization and absorption of hypertrophic chondrocytes, which together lead to ossification (20,21). Similar to this endochondral ossification, osteophyte formation during OA development has been reported to involve VEGF signaling (22). Similarly, recent data reveal the expression of VEGF and its receptors (Flt-1 and Flk-1) in OA cartilage and reflect the ability of VEGF to enhance catabolic pathways in chondrocytes by stimulating matrix metalloproteinase (MMP) activity and reducing tissue inhibitors of MMPs (TIMPs) (23–25). These data suggest that, apart from the effect of VEGF on cartilage vascularization and proliferation of cells in the synovial membrane, chondrocyte-derived VEGF promotes catabolic pathways in the cartilage itself, thereby leading to a progressive breakdown of the extracellular matrix (ECM) of articular cartilage. In the current study, we used a gain- and loss-offunction approach based on tissue engineering techniques to assess the role of VEGF in MDSC-mediated cartilage repair. We demonstrated that genetically modified MDSCs expressing a VEGF antagonist and BMP-4 and transplanted intracapsularly in a rat model of OA enhanced chondrogenesis, repaired cartilage via the autocrine/paracrine effects of BMP-4, and contributed to an appropriate environment that prevented chondrocyte apoptosis by blocking both the intrinsic VEGF catabolic pathway and extrinsic VEGF-induced vascular invasion. This is the first report to describe the effects of VEGF on MDSC-mediated chondrogenesis and OA repair in vitro and in vivo.