| Connective tissue growth factor (CTGF/CCN2), a cysteine-rich module of the CCN family, plays multiple roles in development, cell proliferation and differentiation, and has been considered one of the key factors in tissue fibrosis. The overall objective of this thesis is to test the hypothesis that CTGF may act as a potent profibrogenic factor and promote fibrogenesis that is necessary for tissue regeneration. First, CTGF was found to induce human bone marrow derived mesenchymal stem cells (hMSCs) to differentiate into fibroblasts, as evidenced by remarkable increases in proteins and mRNA of fibroblasts and concomitant reduction in hMSC stemness genes. Furthermore, hMSC-derived fibroblasts upon CTGF treatment fail to differentiate into other mesenchymal lineages including osteoblasts, chondrocytes or adipocytes. Single clones of the heterogeneous hMSCs population were capable of differentiating into not only osteoblasts, chondrocytes, and adipocytes, but also fibroblasts, providing experimental evidence of fibroblast differentiation from single stem cells in the heterogeneous hMSC population, rather than proliferation of existing fibroblasts in the cell population. In addition, microfilaments of CTGF-induced hMSC-derived fibroblasts were aligned in comparison with undifferentiated hMSCs. Together, these findings suggest that CTGF alone is necessary and sufficient to restrict fate selection of hMSCs exclusively towards fibroblasts.;A number of diseases and trauma are intimately related to fibroblasts. For example, ligament and tendon injuries are difficult to repair due to the paucity of fibroblasts. Periodontal disease is characterized by a breakdown of periodontal ligament which is primarily a fibroblast-predominant structure. The focus of the next experiment was related to craniosynostosis, which represents premature ossification of calvarial sutures in newborns and characterized by fate switch of fibroblasts to osteoblasts. In an ex vivo culture model of synostosing rat calvarial sutures, CTGF delivery reversed the fate of craniosynostosis and led to the restoration of fibrogenic/mesenchymal tissues in the calvarial suture.;Extending from the ex vivo data, CTGF was encapsulated in PLGA microspheres for controlled release to potentiate CTGF's bioactive effects. In a well characterized in vivo model of rat interfrontal suture synostosis, microencapsulated CTGF was delivered to determine whether an otherwise fate of synostosis can be reversed. At 4 wks post-op, control-released CTGF successfully restored the synostosing rat interfrontal suture with mesenchymal/fibrogenic tissues. Contrastingly, synostosing calvarial sutures with placebo CTGF-free microspheres were fully re-synostosis. Significant fibrogenesis was revealed in the regenerated calvarial sutures with CTGF delivery, but attenuated in synostosis calvarial sutures without CTGF delivery. Thus, CTGF delivery may transform a highly traumatic craniotomy in existing craniosynostosis patients to a localized suturectomy, and therefore may serve as a key element of minimally invasive biosurgery. CTGF/CCN2 may also have implications in wound healing, organ fibrosis, ligament and tendon repair, and periodontal ligament regeneration. |
