RESUMEN
Dexamethasone (dexa) is commonly used to stimulate osteogenic differentiation of mesenchymal stem/stromal cells (MSCs) in vitro. However, it is paradoxical that glucocorticoids (GCs) such as dexa lead to bone loss and increased fracture risk in patients undergoing glucocorticoid therapy, causing glucocorticoid-induced osteoporosis (GIOP). In a recent publication, we demonstrated that osteogenic differentiation of progenitor cells isolated from jaw periosteal tissue (JPCs) does not depend on dexa, if the medium is supplemented with human platelet lysate (hPL) instead of fetal bovine serum (FBS). This allows the in vitro conditions to be much closer to the natural situation in vivo and enables us to compare osteogenic differentiation with and without dexa. In the present study, we demonstrate that the absence of dexa did not reduce mineralization capacity, but instead slightly improved the osteogenic differentiation of jaw periosteal cells. On the other hand, we show that dexa supplementation strongly alters the gene expression, extracellular matrix (ECM), and cellular communication of jaw periosteal cells. The secretome of periosteal cells previously treated with an osteogenic medium with and without dexa was used to investigate the changes in paracrine secretion caused by dexa. Dexa altered the secretion of several cytokines by jaw periosteal cells and strongly induced osteoclast differentiation of peripheral blood mononuclear cells (PBMCs). This study demonstrates how dexa supplementation can influence the outcome of in vitro studies and highlights a possible role of periosteal cells in the pathogenesis of glucocorticoid-induced osteoporosis. The methods used here can serve as a model for studying bone formation, fracture healing, and various pathological conditions such as (glucocorticoid-induced) osteoporosis, osteoarthritis, bone cancer, and others, in which the interactions of osteoblasts with surrounding cells play a key role.
RESUMEN
Previously, we detected a higher degree of mineralization in fetal calf serum (FCS) compared to serum-free cultured jaw periosteum derived osteoprogenitor cells (JPCs). By Raman spectroscopy, we detected an earlier formation of mineralized extracellular matrix (ECM) of higher quality under serum-free media conditions. However, mineralization potential remained too low. In the present study, we aimed to investigate the biochemical composition and subsequent biomechanical properties of the JPC-formed ECM and minerals under human platelet lysate (hPL) and FCS supplementation. JPCs were isolated (n = 4 donors) and expanded under FCS conditions and used in passage five for osteogenic induction under both, FCS and hPL media supplementation. Raman spectroscopy and Alizarin Red/von Kossa staining were employed for biochemical composition analyses and for visualization and quantification of mineralization. Osteocalcin gene expression was analyzed by quantitative PCR. Biomechanical properties were assessed by using atomic force microscopy (AFM). Raman spectroscopic measurements showed significantly higher (p < 0.001) phosphate to protein ratios and in the tendency, lower carbonate to phosphate ratios in osteogenically induced JPCs under hPL in comparison to FCS culturing. Furthermore, higher crystal sizes were detected under hPL culturing of the cells. With respect to the ECM, significantly higher ratios of the precursor protein proline to hydroxyproline were detected in hPL-cultured JPC monolayers (p < 0.001). Additionally, significantly higher levels (p < 0.001) of collagen cross-linking were calculated, indicating a higher degree of collagen maturation in hPL-cultured JPCs. By atomic force microscopy, a significant increase in ECM stiffness (p < 0.001) of FCS cultured JPC monolayers was observed. The reverse effect was measured for the JPC formed precipitates/minerals. Under hPL supplementation, JPCs formed minerals of significantly higher stiffness (p < 0.001) when compared to the FCS setting. This study demonstrates that hPL culturing of JPCs leads to the formation of an anorganic material of superior quality in terms of biochemical composition and mechanical properties.