Osteogenic differentiation of mesenchymal stem cells (MSC) is important in the field of bone tissue engineering. The identification of biological factors that influence osteogenesis is vital for developing a broader understanding of how complex microenvironments play a role in differentiation. The aim of this study was to demonstrate that adipose-derived stem cell (ADSC) osteogenesis is enhanced through interaction with extracellular matrices (ECM) secreted by ADSC undergoing osteogenesis. ADSC were obtained from human patients following elective abdominoplasty. Cells were selected for plastic adherence, characterized, and induced to differentiate using osteogenic supplements (OS; dexamethasone, ascorbic acid, and beta-glycerol phosphate). Cells were removed at several time points during osteogenesis and the secreted ECM was isolated. Undifferentiated cells were re-seeded onto the cell secreted ECMs and induced to differentiate with OS. At several time points, cells cultured on ECMs or tissue culture plastic controls (i.e. uncoated surface) were collected and RNA isolated. QPCR and gene array analysis revealed enrichment of osteogenic markers and more rapid progression through osteogenic maturational phases in cells seeded onto ECM secreted at the midpoint in differentiation (ca. 15 days). Our results demonstrate that the cumulative deposition of ECM reaches a critical point at approximately 15 days, before which there appear to be no definitive osteogenic cues from the matrix, and after which, strong drivers of osteogenesis are present. The creation of microenvironments that contain essential morphogenic matrix signals is an important step towards methods of growing and differentiating MSC in a rapid effective manner, particularly for bone-related clinical applications.

A BSTRACT : Hematopoiesis is maintained by "fine-tuned" regulation among cytokines, neuropeptides, neurotransmitters, and neurotrophic factors. (1-4) can be partly explained by the induction of suppressive cytokines (TGF-␤ , TNF-␣ , and INF-␥ ). Use of antagonists and screening of a dodecapeptide expression library determined that the effects of SP(1-4) were mediated by NK-1. These results show that PPT-I peptides and their endopeptidase-derived fragments may add to the fine-tuned regulation on hematopoiesis. Furthermore, PPT-I may be exerting autoregulation to protect hematopoietic stem cells. These studies have relevance to stem cell protection and BM transplant. Neurotransmitters, derived from PPT-I exert immune and hematopoietic regulation. PPT-I is also expressed locally in bone marrow (BM) stromal cells. PPT-I peptides induce the production of cytokines in BM cells, resulting in regulation of both committed progenitors (CFU-GM) and primitive hematopoietic progenitors (CAFC). Both types of progenitors are regulated differently by the two major PPT-I peptides, SP and NK-A. Endopeptidases, present in BM cells, can digest SP to produce SP(1-4) and SP(4-11). In this study, we investigated the hematopoietic effects of these fragments on CFU-GM and CAFC. Similar to the two major intact PPT-I peptides (SP and NK-A), we observed different hematopoietic effects by SP(1-4) and SP(4-11). Whereas SP(1-4) exerted inhibitory effects on CFU-GM and CAFC, SP(4-11) mediated stimulatory effects. Similar to NK-A, the inhibitory effects of SP