Factors affecting growth, differentiation and apoptosis of osteoblastic and osteosarcoma cells

Abstract: Osteoblasts play a fundamental role in determining bone structure and function. These cells originate from mesenchymal stem cells (MSCs) and through proliferation and differentiation develop into preosteoblasts and then into mature cells. Most of these cells undergo apoptosis before reaching their terminal differentiated stages of either osteocytes or bone lining cells. These processes, i.e. proliferation, differentiation, and apoptosis, are affected by systemic hormones and local factors. In addition, there are exogenous regulators, which can either be natural substances or synthetic compounds. This thesis describes investigations of the effects of several selected factors on proliferation, differentiation, and apoptosis of osteoblastic cells. The thesis is based on four papers: In the first paper, the effects of Sirt1 regulators, resveratrol (RSV), nicotinamide (NAM), and isonicotinamide (INM), on the commitment ofmesenchymal stem cells (MSC) were studied. We found that the Sirt1 activators, RSV and INM, inhibited adipocyte formation and enhanced osteoblast differentiation, while the inhibitor NAM had the opposite effect. In the second paper, osteoblastic cells from different origins, mouse, rat, and human, were treated with 1alpha,25(OH)2D3 and its analogue, 2-methylene-19-nor-(20S)- 1alpha,25(OH)2D3 (2MD). Species-dependent effects on cell growth and alkaline phosphatase (ALP) activity were clearly seen. In the third paper, we found that the expression of Interleukin-6 (IL-6) receptor increased during osteoblast differentiation. IL-6 acted as a differentiation accelerator in the early stage and an apoptosis inducer at late mature stage. In the forth paper, the effects of Sirt1 activators, RSV and INM, on proliferation and apoptosis of human osteosarcoma (OS) cells were studied. We found an inhibitory effect of Sirt1 activators on OS cells and showed a synergism between RSV and L-asparaginase (ASNase), which is a selective nutritional restrictor. In summary, the work presented in this thesis provides new information about the effects of two osteoblast differentiation regulators, 1alpha,25(OH)2D3 and IL-6. Additionally, certain compounds affecting Sirt1 activity were found to influence osteoblast differentiation; RSV and INM which increase Sirt1 activity also had a profoundly negative effect on growth of OS cells in vitro.