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ISSN Online: 2379-1748
ISBN Flash Drive: 978-1-56700-517-2
5-6th Thermal and Fluids Engineering Conference (TFEC)
Novel Freezing Strategies to Retain the Stem Cell Reserves of Adipose Tissues
Аннотация
The subcutaneous adipose tissue obtained during liposuction procedures is a rich source of stromal vascular fraction (SVF) and stromal/stem cells (ASC), both of which exhibit regenerative properties. Furthermore, "fat grafting" as a preferred cosmetic and reconstructive surgical modality for the treatment of post-lumpectomy or post-mastectomy breast cancer patients. Despite its potential use in regenerative medicine, the vast majority of adipose tissue specimens generated during liposuction surgery is discarded as medical waste due to a lack of validated cryopreservation reagents, protocols, and closed-system collection/preservation devices. The aim of this study to develop a xenoprotein-free cryoprotective agent cocktail that will allow for short-term (up to 6 months) preservation of lipoaspirates tissue sections suitable for fat grafting and/or stromal/stem cell isolation when stored at achievable temperatures (−20 °C or −80 °C). In this study, the lipoaspirates from 3 different donors were suspended with 5 different proprietary cryoprotectant cocktails and frozen at -20 °C and -80 °C for at least a 2 months. After freezing, the lipoaspirates were thawed at 37 °C in water bath, washed with phosphate buffer saline (PBS) to remove the cryoprotectants. ASCs were isolated from the frozen-thawed lipoaspirates by treating them with collagenase. The cell viability was evaluated by fluorescence microscope after staining with acridine orange and ethidium bromide. The ASCs isolated from lipoaspirates frozen at -80 °C resulted in 80% cell viability with 3 of our proprietary cryoprotectant cocktails whereas the other two cryoprotectant combinations resulted in 40% and 60% viability respectively. The surface marker expression (CD90, CD29, CD34, CD146, CD31, and CD45) of ASCs from frozen lipoaspirates at -80 °C in different cryoprotectant media were also evaluated. Furthermore, the adipogenesis and osteogenesis were also studied by histochemical staining and gene expression by quantitative polymerase chain reaction (qPCR).
Video presentation
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