The use of multipotent human mesenchymal stem cells (hMSCs) for tissue engineering has been a subject of extensive research. The donor variation in growth, differentiation and in vivo bone forming ability of hMSCs is a bottleneck for standardization of therapeutic protocols. In this study, we isolated and characterized hMSCs from 19 independent donors, aged between 27 and 85 years, and investigated the extent of heterogeneity of the cells and the extent to which hMSCs can be expanded without loosing multipotency. Dexamethasone-induced ALP expression varied between 1.2-and 3.7-fold, but no correlation was found with age, gender, or source of isolation. The cells from donors with a higher percentage of ALP-positive cells in control and dexamethasone-induced groups showed more calcium deposition than cells with lower percentage of ALP positive cells. Despite the variability in osteogenic gene expression among the donors tested, ALP, Collagen type 1, osteocalcin, and S100A4 showed similar trends during the course of osteogenic differentiation. In vitro expansion studies showed that hMSCs can be effectively expanded up to four passages (approximately 10-12 population doublings from a P0 culture) while retaining their multipotency. Our in vivo studies suggest a correlation between in vitro ALP expression and in vivo bone formation. In conclusion, irrespective of age, gender, and source of isolation, cells from all donors showed osteogenic potential. The variability in ALP expression appears to be a result of sampling method and cellular heterogeneity among the donor population. ß

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Tissue engineering of large bone defects is approached through implantation of autologous osteogenic cells, generally referred to as multipotent stromal cells or mesenchymal stem cells (MSCs). Animal-derived MSCs successfully bridge large bone defects, but models for ectopic bone formation as well as recent clinical trials demonstrate that bone formation by human MSCs (hMSCs) is inadequate. The expansion phase presents an attractive window to direct hMSCs by pharmacological manipulation, even though no profound effect on bone formation in vivo has been described so far using this approach. We report that activation of protein kinase A elicits an immediate response through induction of genes such as ID2 and FosB, followed by sustained secretion of bone-related cytokines such as BMP-2, IGF-1, and IL-11. As a consequence, PKA activation results in robust in vivo bone formation by hMSCs derived from orthopedic patients.bone tissue engineering ͉ osteogenesis ͉ PKA signaling

The cardinal feature of systemic lupus erythematosus is the formation of anti-nuclear antibodies. In recent years, it has become clear that the nucleosome is a major autoantigen that drives this T cell-dependent autoimmune response, as exemplified by the presence of nucleosome-specific T helper cells and the high prevalence of nucleosome-specific autoantibodies. The only way to generate nucleosomes in vivo is by the process of apoptosis. There is growing evidence that in systemic lupus erythematosus apoptosis is disturbed, leading to the release of nucleosomes. Moreover, apoptosis-induced modifications of these autoantigens may render them more immunogenic, especially if the removal of apoptotic cells is insufficient. The first indications for the impaired clearance of apoptotic cells in systemic lupus erythematosus are emerging. Nucleosomes are also important for mediating tissue lesions, especially glomerulonephritis. In lupus nephritis nucleosomes, nucleosome-specific antibodies and nucleosome/IgG complexes have been identified in the glomerular immune deposits. Via their cationic histone part nucleosomes mediate the binding of anti-nuclear antibodies to intrinsic constituents of the glomerular basement membrane, such as the anionic heparan sulfate and collagen IV. Appreciation of this binding mechanism may lead to new treatment strategies, as shown for non-coagulant heparinoids.