Item – Theses Canada

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OCLC number
465562212
Link(s) to full text
LAC copy
LAC copy
Author
Gordon, Jonathan A. R.(Jonathan Alexander Robert),1974-
Title
Cellular interaction with bone sialoprotein regulates osteoblast differentiation and cancer cell migration.
Degree
Ph. D. -- University of Western Ontario, 2006
Publisher
Ottawa : Library and Archives Canada = Bibliothèque et Archives Canada, [2008]
Description
3 microfiches
Notes
Includes bibliographical references.
Abstract
Bone sialoprotein (BSP) is an acidic glycoprotein that is normally expressed in mineralized tissues and aberrantly expressed in metastatic osteotropic tumours. BSP belongs to the SIBLING family (Small Integrin-Binding Ligand N-linked Glycoprotein) of RGD-containing matrix proteins, several members of which have been shown to affect cell differentiation. The role of BSP expression and its interaction with physiological relevant cell types were investigated. The attachment and migration stimulation properties of several RGD-motif mutants of rat recombinant BSP were examined in osteoblasts, breast and prostate cancer cell lines. Alteration of the RGD motif reduced or abolished attachment and migration activity in all cells suggesting that BSP interacts with cells through the RGD motif. The effect of BSP expression in osteoblasts was examined using complementary strategies of adenoviral-mediated overexpression and gene silencing by siRNA. BSP overexpression increased the expression of osterix, Runx2 and osteocalcin mRNA, increased alkaline phosphatase activity and mineralized nodule formation. Conversely, inhibition of BSP expression decreased these markers of osteoblast differentiation. BSP-overexpressiog cells demonstrated increased expression of [alpha]v[beta]3 integrins and elevated focal adhesion kinase (FAK) activity, suggesting a mechanism for promoting osteoblast differentiation. To further delineate the molecular mechanisms of BSP-mediated osteoblast differentiation, overexpression of functional domain mutants of BSP demonstrated that increases in osteoblast markers are mediated by the integrin-binding RGD motif. BSP mediated osteoblast differentiation through FAK as the expression of dominant-negative kinase-dead mutant of FAK abolished effects of BSP. Western blotting for extracellular signal regulated kinase (ERK) and the BMP-related signaling molecules Smad1/5 demonstrated that BSP can activate these two mediators of osteoblastic differentiation. The role of BSP in cancer cells was examined. Expression of BSP but not BSP-RGD mutants in cancer cells increased levels of [alpha]v-containing integrins and the number of mature focal adhesions. BSP also stimulated FAK and ERK phosphorylation, which was significantly increased upon EGF or TGF-[beta]1 stimulation. This expression also resulted in increased cancer cell invasion and cell survival, which were both further increased by addition of EGF or TGF-[beta]1. These results suggest that BSP activates intracellular signaling pathways that have a role in stimulating osteoblast differentiation and regulating motility in cancer cells. 'Keywords'. Bone sialoprotein, osteoblasts, differentiation, cancer migration, focal adhesion kinase
ISBN
9780494303597
049430359X