Human adipose-derived stem cell isolation from fat tissues by membrane filtration method via nylon net filters having different pore size

• Main Authors: Hong-Ren Lin, 林泓任
• Other Authors: Akon Higuchi
• Format: Others
• Language: en_US
• Published: 2016
• Online Access: http://ndltd.ncl.edu.tw/handle/66418766352606571075

碩士 === 國立中央大學 === 化學工程與材料工程學系 === 104 === Human adult stem cells, such as human adipose-derived stem cells (hADSCs), are considered to be a more attractive source of stem cells than human embryonic stem cells (hESCs) and human induced pluripotent stem cells (hiPSCs). This is because human adult stem cells do not generate the ethical concerns that accompany hESCs. The hADSCs exhibit heterogeneous characteristics, indicating various genotypes and differentiation abilities. The isolated hADSCs can possess different purity levels and divergent properties depending on the purification methods used. A hybrid-membrane migration method was developed to purify hADSCs from a fat tissue solution with extremely high purity and pluripotency. A primary fat-tissue solution was permeated through the porous membranes with a pore size from 11 to 80 μm by the membrane filtration method, and the membranes were incubated in cell culture medium for 15-18 days to expand cell number and hADSCs as well as a primary fat-tissue solution were analyzed using flow-cytometry to evaluate mesenchymal stem cell surface markers. The migrated cells from the membranes were also analyzed using flow cytometry, after the membranes were cultivated in the cell culture medium after permeation of primary fat-tissue solution. The hADSCs that migrated from the membranes contained an extremely high percentage (e.g., >95%) of cells positive for mesenchymal stem cell markers. Compared with cells isolated by conventional culture method, the cells isolated by the membrane filtration method showed higher expression of several pluripotency genes (Oct4, Sox2, Klf4 and Nanog) and more calcium accumulation evaluated by alizarin red staining (ARS) and Von Kossa staining (VKS). hADSCs with high purity and pluripotency will be useful as a cell source for regenerative medicine. Especially, the effect of pore size of Nylon net filter membranes on purity and yield of the isolated hADSCs were investigated by the hybrid-membrane migration method and membrane filtration method in this study.