Effect of Cell Seeding Density and Inflammatory Cytokines on Adipose Tissue-Derived Stem Cells: an in Vitro Study

Adipose tissue-derived stem cells (ASCs) are known to be able to promote repair of injured tissue via paracrine factors. However, the effect of cell density and inflammatory cytokines on the paracrine ability of ASCs remains largely unknown. To investigate these effects, ASCs were cultured in 8000 cells/cm(2), 20,000 cells/cm(2), 50,000 cells/cm(2), and 400,000 cells/cm(2) with and without 10 or 20 ng/ml tumor necrosis factor alpha (TNF alpha) and 25 or 50 ng/ml interferon gamma (IFN gamma). ASC-sheets formed at 400,000 cells/cm(2) after 48 h of culture. With increasing concentrations of TNF alpha and IFN gamma, ASC-sheets with 400,000 cells/cm(2) had increased production of angiogenic factors Vascular Endothelial Growth Factor and Fibroblast Growth Factor and decreased expression of pro-inflammatory genes TNFA and Prostaglandin Synthase 2 (PTGS2) compared to lower density ASCs. Moreover, the conditioned medium of ASC-sheets with 400,000 cells/cm(2) stimulated with the low concentration of TNF alpha and IFN gamma enhanced endothelial cell proliferation and fibroblast migration. These results suggest that a high cell density enhances ASC paracrine function might beneficial for wound repair, especially in proinflammatory conditions.