In vitro μελέτη της δράσης αυξητικών παραγόντων και κινάσης της τυροσίνης στην οργάνωση του εξωκυττάριου δικτύου κυτταρικών σειρών οστεοσαρκώματος

Abstract

Osteosarcoma is the most common bone tumor associated with childhood and adolescence. The main characteristics of the osteosarcoma are the malignant transformation of the osteoblasts and the excessive accumulation of the partially mineralized extracellular matrix (ECM). The abundant content of the ECM modulates the microenvironment of the bone tissue and affects the normal function of the osteoblasts as well as the differentiation of the malignant cells. Proteoglycans (PGs) are one of the major classes of macromolecules located both at the cell membrane and the ECM. They consist of a protein core to which one or more sulfated glycosaminoglycan (GAG) chains are covalently bound. GAG chains are linear polymers composed of repeating dissacharide units consisting of a hexosamine and an uronic acid. The specific physicochemical characteristics with which PGs are endowed enable them to affect the organization of the ECM, as well as to participate in the regulation of several cellular events, including cell proliferation, adhesion and migration. These roles of PGs are perpetrated through specific interactions with target macromolecules or growth factors and these interactions may involve both their GAG chains and/or protein cores. PGs are major non-collagen component of the bone ECM. They are synthesized and secreted by osteoblasts and participate in the regulation of bone calcification. Furthermore, it has been suggested that cell membrane (mainly heparan sulfate containing) as well as secreted PGs participate in cell-cell and cell-ECM interactions and play important roles in the in the formation and the renewal of the bone tissue. Both the physicochemical characteristics and the alternations in the composition of the osteosarcoma ECM as well as the association of these changes with its differentiation and prognosis have not been adequately studied. The role of PGs in the mechanisms of osteosarcoma development is suggested to be important. The fine structural analysis of the proteoglycans’ GAG chains may contribute to the better understanding of these glycocomplexes, which participate in the regulation of numerous cellular events as well as in the cell malignant transformation. Therefore, the aim of this study was to contribute to the understanding of the role specific PGs / GAGs have in the pathology of the osteosarcoma, which is one of the most important human primary bone tumors. MG-63 and Saos 2 human osteosarcoma cell lines, endowed with high and low metastatic capability, and differing in their differentiation level, being medium and well differentiated, respectively were utilized. The synthesis and the distribution of GAGs among the cell membrane and the culture medium by these cell lines were studied. The obtained results showed that both cell lines synthesized extracellular hyaluronan (HA) and both extracellular and cell-associated galactosaminoglycans (GalAGs) and heparin sulfate (HS). Even though both cell lines synthesized considerable amounts of PGs, the Saos 2 cells produced HA, GalAGs and HS at considerably lower rates than the MG-63 cells. The role of genistein on the synthesis of these macromolecules has also been studied. Genistein is a well known specific inhibitor of the protein tyrosine kinase (PTK) and affects the proliferation of both cancer and normal cells. The inhibitory effect of genistein on the synthesis of both extracellularly secreted and cell-associated GAGs / PGs in Saos 2 cells was found to be dose-dependent and mediated most probably through a PTK mechanism. The synthesis of GAGs / PGs by the MG-63 cells in the presence of genistein was dependent on their type and localization suggesting that a more complex mechanism regulates their PG synthesis. It has been shown that growth factors such as the transforming growth factor (TGF-β2), basic fibroblast growth factor (bFGF) and the platelet derived growth factor...