Improved GMP compliant approach to manipulate lipoaspirates, to cryopreserve stromal vascular fraction, and to expand adipose stem cells in xeno-free media

Stem Cell Research & Therapy
Francesco AgostiniMario Mazzucato

Abstract

The stromal vascular fraction (SVF) derived from adipose tissue contains adipose-derived stromal/stem cells (ASC) and can be used for regenerative applications. Thus, a validated protocol for SVF isolation, freezing, and thawing is required to manage product administration. To comply with Good Manufacturing Practice (GMP), fetal bovine serum (FBS), used to expand ASC in vitro, could be replaced by growth factors from platelet concentrates. Throughout each protocol, GMP-compliant reagents and devices were used. SVF cells were isolated from lipoaspirates by a standardized enzymatic protocol. Cells were cryopreserved in solutions containing different albumin or serum and dimethylsulfoxide (DMSO) concentrations. Before and after cryopreservation, we analyzed: cell viability (by Trypan blue); immunophenotype (by flow cytometry); colony-forming unit-fibroblast (CFU-F) formation; and differentiation potential. ASC, seeded at different densities, were expanded in presence of 10% FBS or 5% supernatant rich in growth factors (SRGF) from platelets. The differentiation potential and cell transformation grade were tested in expanded ASC. We demonstrated that SVF can be obtained with a consistent yield (about 185 × 103 cells/ml lipoaspirate)...Continue Reading

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Citations

Jun 25, 2019·Frontiers in Cell and Developmental Biology·Letizia SenesiAntonio Gigante
Jul 1, 2018·International Journal of Molecular Sciences·Paola PalumboMaurizio Giuliani
Nov 12, 2020·Microcirculation : the Official Journal of the Microcirculatory Society, Inc·V Morgan JonesAdam J Katz
Apr 4, 2021·International Journal of Molecular Sciences·Baoqiang GuoMark Slevin
Jun 25, 2020·Cells, Tissues, Organs·Jesper Dyrendom SvalgaardLea Munthe-Fog

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Methods Mentioned

BETA
Electrophoresis
flow cytometry

Software Mentioned

CytoVision®
BD Diva
Motic Images Plus
MATLAB®

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