Ex vivo expansion of CD3depletedcord blood-MNCs in the presence of bone marrow stromal cells; an appropriate strategy to provide functional NK cells applicable for cellular therapy.
Considering umbilical cord blood (UCB) as a rich source of hematopoietic stem cells, we introduced a cost-effective approach to expand CD3depletedUCB-MNCs into functional NK cells. CD3depletedUCB-MNCs were expanded in the presence or absence of a feeder [bone marrow stem cells (BMSCs) or osteoblasts], with or without cytokines and their differentiation into NK cells was determined by flow cytometry. NK cell function was quantified by LAMP-1/CD107a expression, TNF-α/IFN-γ release, and LDH release/PI staining in targets. Higher expansion of NK cells was observed after two weeks in the presence of BMSCs and cytokines (104±15) compared to osteoblasts and cytokines (84±29, p<0.05). On day 14, CD3depletedUCB-MNCs in the presence of BMSCs and cytokines showed lower expression of CD3, CD19, CD14, CD15 and CD69 as well as higher expression of CD2 and CD7, which were suggestive of cell differentiation into mature NK cell lineage. Strong cytotoxicity of expanded cells was also identified with higher LDH release and PI% in targets. Significant upregulation of LAMP-1 with decreased release of IFN-γ and TNF-α from effectors were observed. We demonstrate an effective expansion of UCB-NK cells that maintained their functional capabilities applicable for cellular therapies.
Blood Transfusion Research Center, High Institute for Research and Education in Transfusion Medicine, Tehran, Iran. Electronic address: e.hosseini10@yahoo.com.au.
Ehteramolsadat
E
Hosseini
Blood Transfusion Research Center, High Institute for Research and Education in Transfusion Medicine, Tehran, Iran.
Mehran
M
Ghasemzadeh
Blood Transfusion Research Center, High Institute for Research and Education in Transfusion Medicine, Tehran, Iran.
Maedeh
M
Kamalizad
Department of Hematology and Oncology, Eastern School, Monash University, Melbourne, Australia.
Anthony P
AP
Schwarer
D000954:Antigens, Surface / Q000235:genetics / Q000378:metabolism; D001854:Bone Marrow Cells / Q000166:cytology*; D017252:CD3 Complex / Q000235:genetics* / Q000378:metabolism; D002454:Cell Differentiation / Q000187:drug effects; D019070:Cell Lineage; D049109:Cell Proliferation / Q000187:drug effects; D064987:Cell- and Tissue-Based Therapy; D002478:Cells, Cultured; D018920:Coculture Techniques; D016207:Cytokines / Q000494:pharmacology; D005312:Fetal Blood / Q000166:cytology*; D015870:Gene Expression / Q000187:drug effects; D006801:Humans; D007694:Killer Cells, Natural / Q000166:cytology / Q000276:immunology / Q000378:metabolism*; D007963:Leukocytes, Mononuclear / Q000166:cytology* / Q000378:metabolism; D059630:Mesenchymal Stem Cells / Q000166:cytology* / Q000378:metabolism; D010006:Osteoblasts / Q000166:cytology / Q000378:metabolism
D000954:Antigens, Surface; D017252:CD3 Complex; D016207:Cytokines
BMSCs; CD3(depleted) mononuclear cells; Cytotoxicity; NK cell; Osteoblastic cells; Umbilical cord blood