Regenerative potential of ASC-derived conditioned medium (CM)

OBJECTIVE: Adipose-derived Stem Cells (ASCs) are multipotent progenitors able to participate in damaged tissue regeneration and to modulate immune response. Their regenerative and protective action might be mainly due to the release of soluble factors and/or extracellular vesicles (EVs) which might mediate immunomodulation, proliferation and differentiation of the surrounding cells. We have found that hASC-conditioned medium (CM) counteracts neuropathic pain symptoms in preclinical mouse models. Here we show the ASC secretome effects in vitro on primary cells and the biophysical characterization of its EVs. MATERIALS AND METHODS: ASC were isolated from subcutaneous adipose tissue and cultured in cDMEM. After a 72-hours-starving period, ASC CM was collected, concentrated through Amicon Ultra-15 Centrifugal Filter Unit (Merck-Millipore) and its protein content quantified by Bradford assay. The trophic effect was tested on terminally differentiated cells (human osteoblasts, OBs, and human chondrocytes, CHs) and progenitors (ASCs) by Alamar Blue and MTT assays. Micro-vesicles (MVs) of 500-1000 nm and MVs-depleted CM (CM-MVs) were fractionated by centrifugation at 10,000xg, while exosomes (80-100 nm) and smaller micro-vesicles (