ABSTRACT
Background: As adipose tissue-derived mesenchymal stem cells are becoming the tool of choice for many clinical applications; standardized cryopreservation protocols are necessary to deliver high-quality samples. For this purpose, the cryopreservation and thawing of native adipose tissue under GMP conditions could represent an extremely useful and powerful tool for the direct reinfusion of the tissue, and consequently, of its stromal vascular fraction. Methods: In this study, 19 samples of adipose tissue were cryopreserved and characterized before and after storage in liquid nitrogen vapors. Of these 19 samples, 14 were processed in research and 5 in a GMP-compliant environment. Storage with and without cryopreservation medium was also evaluated. After one week to three months of storage, samples were thawed, washed, enzymatically digested, and characterized with flow cytometry. Results: The results show that there is a loss of nearly 50% of total nucleated cells during the cryopreservation/thawing process. Non-GMP and GMP samples are comparable for all parameters analyzed. This study also allowed us to exclude the cryopreservation of adipose tissue without any cryopreservation medium. Conclusions: The data shown in this work are consistent with the idea that native adipose tissue, if properly processed and controlled, could be a useful source of cells for regenerative medicine, keeping in mind that there is a clear difference in the quality between fresh and thawed samples.
ABSTRACT
In recent years, there has been a shift toward tissue-engineering strategies using stem cells for plastic and reconstructive surgical procedures. Therefore, it is important to develop safe and reproducible protocols for the extraction of adipose-derived stromal cells (ASCs) to allow cells to be stored in liquid nitrogen for future needs. The aspirated liposuction obtained from healthy donors were immediately processed after the suction using a protocol developed in our laboratory. The resulting stromal vascular fraction (SVF) was then characterized by the presence of adipose-derived stromal cells, at later stage frozen in liquid nitrogen. After that, cells were thawed and again characterized by adipose-derived stromal cells, cellular survival, differentiation ability and Colony Forming Unit-Fibroblast like colonies (CFU-F). Extraction and freezing of cells contained in the stromal vascular fraction demonstrate that thawed cells maintain the full capability to grow and differentiate in culture. The advent of adipose-derived stromal cells use in tissue engineering will assume a wide role in esthetic restoration in plastic surgery. It is thus important to develop clinically translatable protocols for the preparation and storage of adipose-derived stromal cells. Our results show that adipose-derived stromal cells in serum free can easily be frozen and stored in liquid nitrogen with retention of 85% of cell viability and 180,890 cell/g yield plus normal proliferative capacity and differentiation potential compared with fresh controls. These observations set the basis for adipose-derived stromal cells banking.
