Abstract
: It is important to standardize methods to quantify the purity of adipose tissue-derived cells for regenerative medicine. We developed a simple and robust tool to discriminate fibroblasts and adipose stem cells (ASCs) by testing release of specific growth factors. ASCs and dermal fibroblasts (DFs) were isolated from human donors (n = 8). At passage 4, cultures were prepared with progressive ASC/DF ratios of 100%/0%, 75%/25%, 50%/50%, 25%/75%, and 0%/100% for each donor and incubated in hypoxic chambers at 0.1% and 5% O(2) and hyperglycemia at 1.0 and 4.5 g/l. After incubation for 24 hours, cell survival, proliferation, and growth factor release (vascular endothelial growth factor [VEGF], hepatocyte growth factor [HGF], insulin-like growth factor 1 [IGF-1], stromal cell-derived factor 1α [SDF-1α], and basic fibroblast growth factor [bFGF]) were assessed for each condition. The proliferation and viability of ASCs and DFs were not impacted by the oxygen tension conditions. No significant difference in HGF, IGF-1, bFGF, and keratinocyte growth factor secretome was found across the various ASC/DF ratios. Interestingly, a negative relation for VEGF secretion was found when ASCs were contaminated by fibroblasts, especially when cells were exposed to 4.5 g/l glucose and 0.1% O(2) (R = -0.521; p < .001). In contrast, secretion of SDF-1α was positively correlated with the fibroblast ratio, more prominently in low glucose and low oxygen tension (r = .657; p < .001). Above and beyond these previously unreported metabolic features, these results (a) allow us to discriminate fibroblasts and ASCs specifically and (b) allow new tools be developed for the rapid testing (a response within 24 hours) for the release of ASC-based therapies. SIGNIFICANCE: In order to provide direction to academia, industry, and regulatory authorities regarding purity assessment for adipose tissue-derived cells, this report describes a simple tool to facilitate development of international standards based on reproducible parameters and endpoints that may systematize cellular products across boundaries and accelerate the delivery of safe and effective adipose stem cell (ASC)-based tools to the medical community and the patients it serves. This tool (a) can discriminate specifically fibroblasts and ASCs and (b) can be rapidly implemented and performed before the release of the ASC-based therapy (a response within 24 hours).