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Type: Thesis
Title: Cellular and molecular mechanisms involved in bony tissue repair of injured growth plate cartilage in rats.
Author: Chung, Rosa
Issue Date: 2011
School/Discipline: School of Medical Sciences
Abstract: Being cartilage, the growth plate is often injury prone. This remains to be a significant problem particularly in children where, due to the dynamic nature of their skeletal growth, injury to the growth plate can result in orthopaedic problems including limb-length discrepancy and angulation deformity. Previous studies have identified these problems as a direct result of formation of bony repair tissue at the injury site. Although the sequential post-injury responses (namely the inflammatory, fibrogenic, osteogenic and remodelling phases) have been previously well documented histologically, the molecular and cellular events underlying the bony repair remain unclear. Using a well established rat growth plate injury model, this PhD project characterised presence of possible stromal progenitor cells within the mesenchymal infiltrate, roles of chemotactic growth factor PDGF-BB and protein kinase-D (PKD) in the fibrogenic response and subsequent bony repair events. Immunohistochemical analysis of tibial growth plates at different time points post-injury revealed cells immunopositive for alphasmooth muscle-actin (αSMA) or Activin-A Receptor Type II-like kinase- 3 (ALK-3) within the mesenchymal infiltrate, suggesting the potential presence of mesenchymal stem cell (MSC)-like cells. In addition, positive immunostaining of MSC-negative but endothelial cell-positive marker, von Willebrand Factor (vWF), also indicated that not all the cells within the infiltrate were MSC-like cells. Further analysis revealed that a portion of cells were immunopositive for osteogenic transcription factor core-binding factor-alpha 1 (cbf-α1) or chondrogenesis marker collagen-IIa, suggesting osteogenic and chondrogenic progenitors may also exist, respectively. Further studies are required for confirmation of MSC-like and progenitor cell existence within the infiltrate and their involvement in the bony repair. While the importance of the fibrogenic phase of repair is evident, the factors responsible for this cell influx are poorly studied. Previous studies have shown upregulation of the known key chemoattractant, PDGF-BB just prior to and during fibrogenic response. Studies in this project revealed that inhibition of PDGF signalling resulted in a significant delay in the healing responses in rats. Also in vitro studies found that PDGF-BB increased bone marrow stromal cell migration into an artificial “wound” site (P<0.005), which can be suppressed by the PDGF receptor inhibitor. These results suggest that PDGF signalling contributes to growth plate injury repair by promoting mesenchymal progenitor cell infiltration and subsequent tissue repair. Fibrogenic cells within the injury site can differentiate into bone or cartilage cells. However, what signals/ factors underlie these cell differentiation processes and bony repair remain unexplored. While osterix is one known important transcriptional factor for osteoblast maturation, and PKD is known to be involved in transcription of osterix, their potential roles in growth plate bony repair are unknown and were investigated in this project. Micro-CT and histology analysis of injury sites in rats treated with PKD inhibitor revealed significantly lower amount of bone formed after inhibiting PKD signalling (P<0.05). Consistently, inhibitor-treated animals showed decreased mRNA expression of bone-related genes (osterix and osteocalcin) and increased levels of cartilage-related genes (collagen-IIa and Sox9). In support, in vitro experiments showed that addition of PKD inhibitor during chondrogenic differentiation of rat primary bone marrow stromal progenitor cells resulted in a significant increase in collagen-IIa expression (P<0.05). These results suggest that PKD is an important factor for growth plate bony repair and blocking PKD activity after growth plate injury may result in partial suppression of osterix, less bone formation and potentially more desirable cartilage repair.
Advisor: Xian, Cory J.
Foster, Bruce Kristian
Dissertation Note: Thesis (Ph.D.) -- University of Adelaide, School of Medical Sciences, 2011
Keywords: growth plate injury; chondrocytes; osteoblasts; mesenchymal stem cells; chondrongenesis; osteogenesis; growth factors
Provenance: Copyright material removed from digital thesis. See print copy in University of Adelaide Library for full text.
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