Ex Vivo Induction of Functional Human Neutrophils from Serum-Free Expanded Hematopoietic Stem Cells

• Main Authors: Yu-Shuo Hu, 胡育碩
• Other Authors: Chao-Ling Yao
• Format: Others
• Language: zh-TW
• Online Access: http://ndltd.ncl.edu.tw/handle/57928067987293837766

碩士 === 元智大學 === 化學工程與材料科學學系 === 101 === In recent years, the number of people who receive chemotherapy increased due to the rising trend of malignancy. Hematopoietic stem cells (HSCs) from bone marrow, peripheral blood and cord blood are widely used in transplantation after cancer chemotherapy. Among them, there are several limitations of cord blood hematopoietic stem cell transplantation, such as slow recoveries of neutrophils and platelets. The aim of this study is to develop a neutrophil induction medium to increase the number of neutrophils in vitro, to accelerate the recovery of autoimmune system of patients after cord blood HSC transplantation and to overcome the limitation of clinical application. In this study, HSCs were isolated from cord blood and were expand by a serum-free HSC expansion system (SF-HSC medium) that was developed previously by our lab. Then we screened five of cytokines (SCF, G-CSF, GM-SCF, IL-3 and IL-1) and optimized the concentration of cytokines that can effectively induce serum-free expanded HSCs into CD66+/MPO+ cells. In addition, we also checked CD66+/MPO+ cells by neutrophil-associated surface antigens, such as CD11b, CD13, CD15, CD16, CD33, CD64, TLR-2 and TLR-4, and neutrophil-specific functional assays, such as phagocytosis assay, chemotaxis assay and cellular reactive oxygen species (ROS) production assay, challenged with Pseudomonas aeruginosa after transplanted neutrophils in NOG mices. Finally, we confirmed induced cells were functional neutrophils. In this study, we have successfully established a neutrophil induction medium that can effectively induce HSCs into mature and functional neutrophils in vitro. We believe that combination of SF-HSC medium and neutrophil induction medium can generate a large amount of functional neutrophils and provide a promising cell source for basic research and clinical application in the future.