INTRODUCING NON-ADHERENT PROGENITORS FROM ADIPOSE-DERIVED STEM CELLS (NAPADSCS): PROOF OF STEMNESS AND POSSIBLE FUTURE APPLICATIONS IN REGENERATIVE SURGERY

Abstract

Introduction: Mesenchymal Stromal Cells (MSCs) can be isolated from adipose tissue and are currently used in vitro for experimental studies. MSCs are often mislabeled as Adipose-derived Stem Cells (ADSCs) despite presenting a more differentiated phenotype. The isolation and expansion of non-adherent progenitors from adipose-derived stem cells (napADSCs), has been achieved in our laboratory and is currently under investigation. In this study we investigate: a) the proof of stemness of such progenitors, and b) the feasibility of napADSCs adhesion over Integra® for cell colonization and for future differentiation and engineering of semi-synthetic tissues. Material and Methods: Adipose tissue (20 cc) was extracted from lipoaspirate samples of 15 healthy donors following patients written consent. Following mechanical and enzymatic digestion, samples were plated in stem cell-specific enriched media and in no adherence conditions. Clonal expansion of a single cell was assessed by limiting dilution and asymmetric division was detected by PKH26 staining. Expanded cells were seeded within Integra® in 24-well plates. Results: NapADSCs represent an upstream line of mesenchymal progenitors compared to more differentiated, adherent, fibroblast-like MSCs. NapADSCs colonies defined as spheroids (polyclonal) and sphere-derived clonogenic cells (monoclonal) were visible in 1-3 weeks and their stemness confirmed in vitro by clonal expansion. NapADSCs adhesion to Integra® was achieved modifying culturing conditions, and was visible in 3-7 days with phenotype change from napADSCs to fibroblast-like MSCs. Conclusions: Our data suggests that the identification of napADSCs may dissipate the doubts on the stem-cell origin of the more differentiated and commonly used adherent MSCs. The ability of napADSCs to adhere to a clinically available dermal regenerative template, and grow within its three-dimensional structure, may prove useful in regenerative surgery. Further studies are warranted to assess whether such procedure may represent a first preliminary step toward clinical application of semi-synthetic tissues.