However in the case of large, critical-sized defects, these normal healing cascades are disrupted due to the shear size of the defect, as well as a number of confounding factors such as infection, mechanical instability, and absent sources of regenerative cues. Historically, tissue engineers have vastly improved upon many of these shortcomings with the application of novel scaffolds, a variety of clinically relevant cells, and growth factors to instruct cells. More recently, inflammatory mediators have Cefathiamidine gained interest in the field because of their important roles in normal healing processes. The goal of this study was to study the effects of TNF, a key inflammatory cytokine, on vascular assembly and osteogenesis. We demonstrate that TNF acts synergistically with PDGF another important Crolibulin component of the bone-healing cascade – to enhance vascularization within engineered osteogenic grafts. Initial studies tested the effects of TNF dose on ASC osteogenesis and vascular morphogenesis in separate cultures, demonstrating that TNF has a beneficial effect on both lineages when supplied at a low dose for 48 hours. While TNF induced a moderate increase in mineralization, this was concomitant with an increase in DNA content, suggesting that the increased mineralization may be due to proliferation. However, previous studies have indicated that TNF induces increased endogenous production of bone morphogenetic protein-2 in mesenchymal stem cells, suggesting that there may be a direct osteogenic effect on the cells. Not all pro-inflammatory cytokines elicit the same response, as we conducted a similar experiment with interleukin-1b that resulted in a significant increase in mineralization for all concentrations ranging from 0.1�C100 ng/mL, but a sharp dose-dependent inhibition of vascular assembly. In the in vitro model of vascularized bone development, acute treatment with TNF increased vessel length and interconnectivity. However, continuous exposure to the same TNF dose had an adverse effect on vascular network structure.This finding is supported by studies with endothelial cell cultures that have shown that continuous exposure to TNF can inhibit endothelial cell proliferation and growth factor responsiveness.