The effects of platelet lysate on adipose-derived stem cell sheet formation and its potential wound healing capability

• Main Authors: Ning Hsu Lee, 李寧栩
• Other Authors: Tai-Horng Young
• Format: Others
• Language: en_US
• Published: 2019
• Online Access: http://ndltd.ncl.edu.tw/handle/246ch8

碩士 === 國立臺灣大學 === 醫學工程學研究所 === 107 === Adipose-derived stem cells (ASCs) holds a valuable future in regenerative medicine. ASC sheet has been suggested to promote tissue healing and has been applied as possible treatment in a variety of disease models. However, the use of animal-derived serum supplemented growth media may lead to concerns regarding to clinical complication. In recent years, human platelet lysate (HPL) provides an attractive alternative to fetal bovine serum (FBS) for the ex-vivo expansion of mesenchymal stem cells for clinical use. In this study, we compare ASC sheets that have been cultured in growth media supplemented with either FBS or HPL. ASCs were cultured in Dulbecco’s modified Eagle’s medium (DMEM)-high glucose supplemented with either FBS or HPL. Cell sheet formation was further induced by the addition of ascorbic acid 2-phosphate. We performed quantitative PCR, western blotting, ELISA and proteomic analysis to further compare ASC sheets culture with FBS and HPL. In this study, we found that ASC sheets cultured with HPL acquired a higher population doubling and better osteogenic differentiation, though adipogenic differentiation was reduced relative to ASC sheet cultured with FBS. Proteomic analysis of sheet extracellular matrix showed a much more abundant extracellular matrix deposition. ASC sheets cultured in HPL-supplemented growth medium exhibit enhancing effects on the proliferation and migration of dermal fibroblasts and endothelial cell. Conditioned medium collected from ASC sheets cultured in with HPL showed significantly higher proliferation and migration rates when used to cultured fibroblasts cell line HS68. It also results in more tube formation in the in-vitro HUVEC angiogenesis assay when compared to medium collected from ASC sheets cultured with FBS. Further analysis revealed that HPL-supplemented culture medium significantly enhanced expression of hepatocyte growth factor (HGF) and chemokine (C-C motif) ligand 5, and reduced expression of IL-6 in ASC sheets, which have all been shown to play important roles in angiogenesis and wound healing process. Overall, Supplementation of HPL promoted a faster and more robust ASC sheet formation, and the secretion of angiogenic paracrine factors from ASC sheets was also enhanced. ASC sheet cultured with HPL has been shown to promote in vitro fibroblasts and endothelial cells proliferation and migration. Though the mechanism behind these results is yet to be confirmed, these in-vitro test results demonstrated a potential wound healing capability of HPL-cultured ASC sheet in addition to circumventing the untoward side effects for using FBS for cell culture. Therefore, HPL-cultured ASC sheet exhibits great potential in future clinical application to promote better wound healing.