Dr. Alan J. Nixon
This proposal broadly aims to enhance the ability of stem cells to repair damaged joints. Despite popular perception, cultured stem cells or marrow derived cell mixtures seem incapable of forming durable cartilage. Moreover, in adult horses, focal cartilage injury has little to no potential for spontaneous repair. We hypothesize that this is due, at least in part, to the absence of a recruitable local stem cell niche that would normally contribute and coordinate repair. These experiments examine this hypothesis and ways to overcome the functional decline of progenitor action with aging and disease. In a series of aims, this research will characterize a putative cartilage progenitor stem cell “niche”, and then develop methods to supplement this meager cell pool with gene-programmed stem cells that can seed and re-populate a progenitor nidus in extensive cartilage injuries. The long-term goal of our studies is to reliably regenerate cartilage and avert secondary osteoarthritis (OA) which plays a major role in abbreviating a horse’s athletic career. The work will characterize developing, immature, adult, and diseased cartilage to define progenitor cell populations by immunocharacterization and gene expression profiling. Having identified target regions for supplemental regenerative stem cell pools, this proposal will then develop a reliable mechanism to effect a chromosomal based shift in stem cell phenotypic programming that may drive in vivo formation of a dedicated stem cell niche in cartilage. Grafted stem cells have been heralded as a break-through in repair of numerous connective tissue diseases. However, applying undifferentiated or poorly differentiated MSCs to a joint surface and expecting durable repair has so far been ineffective. This studies proposed here will strengthen this field by clearly defining the target layers and developing a cartilage progenitor stem cell niche of autologous MSCs stably transduced with chondroinductive genes. The fundamental goal is to use stem-cell based cartilage repair to lessen arthritis and hasten return to performance. Presently, joint disease is still one of the leading causes of retirement in racehorses. Treatment costs have spiraled without emerging curative techniques. While it is well established that pluripotent cells can respond to the bodies call for repair of musculoskeletal tissues, the extent of local differentiation to target cells declines with maturity. Driving stem cell chondrogenesis may overcome this aging effect and promote durable cartilage to prevent debilitating arthritis. Previous studies in the investigator’s laboratory indicate that implanting simple stem cell cultures has only transitory effect in improving cartilage repair. We hypothesize this is due to inadequate chondrogenic differentiation of MSCs down the target lineage. This grant utilizes Sox transcription factors and TGF-β family members to recapitulate fetal cartilage development. These equine specific genes have been recombined into Sleeping Beauty non-viral vectors that integrate to the chromosomal DNA of target cells. There we expect they will exert cellular and tissue specific pressure to drive cartilage differentiation and resist matrix degradation in stem cell pools. We will use a novel 3D culture system to characterize these cells and their progenitor to lineage pathway, and later evaluate repair in long-term in vivo studies. This grant will be the first to use sequential gene delivery to deliberately “pre-program” a stem cell line toward the cartilage lineage. The long-range goal is to provide a ready source of autologous chondrocyte-like stem cells for transfer to acutely damaged joints.