Expansion of alpha-galactosylceramide-stimulated Valpha24+ NKT cells cultured in the absence of animal materials.
Valpha24+ NKT is an innate lymphocyte with potential antitumor activity. Clinical applications of Valpha24+ natural killer (NK) T cells, which are innate lymphocytes with potential antitumor activity, require their in vitro expansion. To avoid the potential dangers posed to patients by fetal bovine serum (FBS), the authors evaluated non-FBS culture conditions for the selective and efficient expansion of human Valpha24+ NKT cells. Mononuclear cells (MNCs) and plasma from the peripheral blood of normal healthy donors were used before and after G-CSF mobilization. MNCs and plasma separated from apheresis products were also used. MNCs were cultured for 12 days in AIM-V medium containing alpha-galactosylceramide (alpha-GalCer) (100 ng/mL) and IL-2 (100 U/mL) supplemented with FBS, autologous plasma, or autologous serum. The cultured cells were collected and their surface markers, intracellular cytokines, and cytotoxicity were evaluated. The highest expansion ratio for Valpha24+ NKT cells was obtained from G-CSF-mobilized MNCs cultured in medium containing 5% autologous plasma. Cultures containing MNCs and autologous plasma obtained before and after G-CSF mobilization had approximately 350-fold and 2,000-fold expansion ratios, respectively. These results suggest that G-CSF mobilization conferred a proliferative advantage to Valpha24+ NKT cells by modifying the biology of cells and plasma factors. Expanded Valpha24+ NKT cells retained their surface antigen expression and production of IFN-gamma and exhibited CD1d-independent cytotoxicity against tumor cells. Valpha24+ NKT cells can be efficiently expanded from G-CSF-mobilized peripheral blood MNCs in non-FBS culture conditions with alpha-GalCer and IL-2.
Harada
Yukie
Y
Hematopoietic Stem Cell Transplant/Immunotherapy Unit, National Cancer Center Hospital, Tokyo 104-0045, Japan.
Imataki
Osamu
O
Heike
Yuji
Y
Kawai
Hiroyuki
H
Shimosaka
Akihiro
A
Mori
Shin-ichiro
S
Kami
Masahiro
M
Tanosaki
Ryuji
R
Ikarashi
Yoshinori
Y
Iizuka
Akira
A
Yoshida
Mitsuji
M
Wakasugi
Hiro
H
Saito
Shigeru
S
Takaue
Yoichi
Y
Takei
Masao
M
Kakizoe
Tadao
T
D001781:Blood Component Removal; D018929:Cell Culture Techniques; D049109:Cell Proliferation; D003470:Culture Media / Q000737:chemistry / Q000494:pharmacology; D016207:Cytokines / Q000097:blood / Q000378:metabolism; D003601:Cytotoxicity Tests, Immunologic; D005699:Galactosylceramides / Q000276:immunology*; D016179:Granulocyte Colony-Stimulating Factor / Q000494:pharmacology; D006801:Humans; D016219:Immunotherapy, Adoptive; D007694:Killer Cells, Natural / Q000166:cytology* / Q000276:immunology / Q000378:metabolism; D007958:Leukocyte Count; D007963:Leukocytes, Mononuclear / Q000166:cytology / Q000187:drug effects / Q000378:metabolism; D018655:Lymphocyte Count; D016693:Receptors, Antigen, T-Cell, alpha-beta / Q000276:immunology*; D018121:Receptors, Cytokine / Q000378:metabolism; D016176:T-Lymphocyte Subsets / Q000166:cytology* / Q000276:immunology / Q000378:metabolism; D013601:T-Lymphocytes / Q000187:drug effects / Q000276:immunology / Q000378:metabolism
D003470:Culture Media; D016207:Cytokines; D005699:Galactosylceramides; D016693:Receptors, Antigen, T-Cell, alpha-beta; D018121:Receptors, Cytokine; C493154:alpha-galactosylceramide; D016179:Granulocyte Colony-Stimulating Factor