Effects of 1,25-dihydroxyvitamin D3 and growth hormone on osteoblast-like cells

Sammanfattning: Bone remodeling, which occurs throughout life in both cancellous and cortical bone, involves mainly the osteoblasts and osteoclasts. The osteoblast is responsible for the production of matrix, which later becomes mineralized. The osteoclast is a large, multinuclear cell, which is responsible for bone resorption. Bone remodeling is controlled by systemic hormones and cytokines and is an integral part of the calcium homeostasis system. An imbalance between bone formation and bone resorption may result in diseases such as osteoporosis. The aims of this thesis were to study growth hormone (GH) and 1alpha,25-dihydroxyvitamin D3 (1,25(OH)2D3) intracellular signaling in the osteoblast and to initiate the establishment of an in vitro system to differentiate human adult mesenchymal stem cells along an osteblastic lineage. In the first paper, we focused on whether cells of the UMR 106 cell line (a rat clonal osteosarcoma cell line with osteoblast-like phenotypic properties and endogenous expression of growth hormone receptor), express a GH-response through the JAK2/STAT5 pathway. We demonstrated the expression and GH-induced activation of JAK2 and STAT5A/B and showed that this cell line is a suitable experimental model to study unique GH effects in osteoblasts mediated by STAT5. In the second paper, we demonstrated that pretreatment of UMR 106 cells with 1,25(OH)2D3 prolonged GH signaling via JAK2/STAT5 pathway. Pretreatment of cells with 1,25(OH)2D3 was also necessary in order to detect GH-induced STAT5 transcriptional response and also rendered the cells capacity to respond to repetitive GH stimulation. Pretreatment with 1,25(OH)2D3 also inhibited GH-induced expression of SOCS-3 and CIS, proposing that 1,25(OH)2D3 had an inhibitory effect on negative regulatory pathways acting on JAK2 and/or STAT5 in UMR 106 cells. In the third paper of this thesis we investigated the effects of 1,25(OH)2D3 and GH on proliferation and apoptosis in UMR 106 cells. We found that 1,25(OH)2D3 and GH, separate or in combination, inhibited apoptosis. GH also had profound effects on cell cycle distribution and proliferation. In addition, pretreatment of cells with 1,25(OH)2D3 was necessary in order to detect GH induced MAPK activation. We hypothesize that these hormones, separately or in combination, regulate the processes of apoptosis and proliferation, which may be important for the osteoblast cell number. Finally, in paper four we investigated the effect of 1,25(OH)2D3 on the differentiation of human adult mesenchymal stem cells. Treatment with 1,25(OH)2D3 affected the processes of apoptosis and differentiation. These findings may have important implications for the future usage of stem cells in the treatment of bone disorders.