Differentiation and maintenance of mast cells from CD34(+) human cord blood cells.
OBJECTIVE
Establishment of a stable umbilical cord blood CD34(+) (UCB CD34(+)) cell culture system and identification of the cells in the cobblestone area differentiated from UCB CD34(+) long-term culture cells.


METHODS
Human UCB CD34(+) cells were cultured on MS-5 mouse stroma cells in the presence of stem cell factor (SCF), flt-3 ligand (FL), and thrombopoietin (TPO) for 4 to 16 weeks. Cells in the culture medium and in the cobblestone area were collected and characterized by flow cytometry and microscopy.


RESULTS
CD34(+) cells were stably expanded by culturing on MS-5 stroma cells in the presence of SCF, FL, and TPO for more than 4 months. Cells highly expressing CD117 (c-kit) appeared in the cobblestone area after 2 weeks and stably expanded. Isolation of cells highly expressing CD117 by fluorescence-activated cell sorter (FACS) revealed the cells were tryptase-positive and Fc epsilon receptor 1-negative mast cells. They showed typical mast cell morphology and released histamine upon stimulation by substance P or compound 48/80 in vitro.


CONCLUSIONS
Human UCB CD34(+) cells were stably expanded on MS-5 stroma cells in the presence of SCF, FL, and TPO. Under this condition, multipotent CD34(+) cells and mast cells differentiated from UCB CD34(+) cells were expanded in the cobblestone area. The expanded mast cells showed histamine release after substance P or compound 48/80 stimulation. These human mast cells will be useful as a source of human cells for evaluating the allergic effects of drugs.
Pharmaceutical Research Department III and Preclinical Research Department II, Chugai Pharmaceutical Co., Ltd, Kamakura, Japan. yoshikubotks@chugai-pharm.co.jp
Takashi
T
Yoshikubo
Tomoaki
T
Inoue
Mizuho
M
Noguchi
Hisafumi
H
Okabe
D000818:Animals; D018952:Antigens, CD34 / Q000276:immunology*; D002454:Cell Differentiation; D018920:Coculture Techniques; D005312:Fetal Blood / Q000166:cytology* / Q000276:immunology; D005434:Flow Cytometry; D006801:Humans; D016130:Immunophenotyping; D008407:Mast Cells / Q000166:cytology*; D051379:Mice; D016688:Mice, Inbred NOD; D016513:Mice, SCID
D018952:Antigens, CD34