Osteoporosis is a metabolic disease of the bone, characterized by low bone mineral density, microarchitectural alterations and increased risk for an osteoporotic fracture. Due to its polygenic and multifactorial nature, despite intensive research, the key genes involved in the pathogenesis of osteoporosis still have not been revealed. Osteoblasts are bone cells with a central role in bone metabolism. Their main role is the formation of bone tissue, however they also regulate osteoclast differentiation and activity and participate in response to mechanical loading of the bone.We used an innovative approach in order to discover new genes participating in the pathogenesis of osteoporosis. Genome wide DNA microarrays, capable of identification of new molecules on the field of bone biology, were used to analyze the gene expression in primary cultures of osteoblasts, obtained from osteoporotic and non-osteoporotic bone tissue. We found 1606 genes, which were differentially expressed between osteoblasts from osteoporotic and non-osteoporotic bone tissue. 352 genes exceeded the common 2-fold threshold. Based on the analysis of function of differentially expressed genes, we suspect that the osteoblasts differ in activity, proliferation rate and degree of differentiation, since many of the differentially expressed genes encode ribosomal proteins and proteins involved in the cell cycle. Based on our results, we further analyzed the involvement in osteoporosis of 4 genes and one biochemical process on the level of bone tissue and human osteosarcoma cell line. Genome wide analysis of osteoblast transcriptomes suggests important roles of cxcl2, ibsp, ptn and col15a1 in osteoporosis. Therefore we used quantitative polymerase chain reaction (qPCR) to measure the expression of the above mentioned genes in bone tissue of a larger group of patients, to assess their clinical applicability. We found downregulation of ptn and col15a1 and upregulation of ibsp (p < 0,05) in osteoporotic bone tissue. Upregulation of cxcl2 in bone tissue was not statistically significant. Genome wide analysis of osteoblast transcriptomes also showed the importance of oxidative stress in osteoporosis. Therefore, we designed an in vitro experiment, where human osteosarcoma cells were exposed to simulated conditions of oxidative stress. Cells were treated with hydrogen peroxide and the expression of 12 genes was monitored by qPCR. The experiment confirmed the differential expression of most of the selected genes, of which we would like to highlight txnrd1, aox1, gsr and mt1g. By using genome wide gene expression microarrays we found a large number of differentially expressed genes and functional gene sets. Our results suggest upregulation of cxcl2, downregulation of ptn, increased demand for protein synthesis, decreased cell proliferation rate and upregulation of genes, involved in response to reactive oxygen species, indicating increased exposure of osteoblasts to oxidative stress in osteoporotic tissue. Ptn, cxcl2, col15a1, ibsp, aox1, mt1g, gsr and txnrd1 are the most important candidate genes for further studies of the genetic background of osteoporosis. Furthermore, our results offer novel suggestions for the study of new molecules and biochemical pathways in osteoblasts and bone tissue.
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