The experimental study on inducing and expanding T/NK cells from mononuclear cells of human umbilical cord blood.
- Author:
Ge-yu CHEN
1
;
Shao-liang HUANG
;
Dun-hua ZHOU
;
Yan-feng WU
;
Jing WEI
;
Qin CHEN
Author Information
- Publication Type:Journal Article
- MeSH: Cell Differentiation; Cell Division; Cytotoxicity, Immunologic; Fetal Blood; cytology; Humans; Infant, Newborn; Interleukin-2; pharmacology; Killer Cells, Natural; cytology; Leukocytes, Mononuclear; cytology; Receptors, Antigen, T-Cell, gamma-delta; analysis; T-Lymphocytes; cytology; Tumor Necrosis Factor-alpha; pharmacology
- From: Chinese Journal of Hematology 2003;24(11):576-579
- CountryChina
- Language:Chinese
-
Abstract:
OBJECTIVETo explore the most efficient culture system which can induce cord blood (CB)-mononuclear cells (MNC) to differentiate into mature T/NK cells in vitro.
METHODSThe CB MNCs were cultured in six culture systems respectively for 4 weeks. The T/NK cell surface phenotypes were analyzed by flow cytometry and the absolute numbers of nucleated cells (NCs) were counted at each time point. Moreover, cell morphology was identified by Giemsa-Wright staining, and cytotoxicity of the cultured cells to K562 and Raji tumor cells was also evaluated by MTT method.
RESULTSCultured in the cytokine cocktail of SCF + FLT-3L + IL-7 + IL-15 + TNF-alpha + IL-2, the NCs were (20 approximately 26) x 10(6)/ml in numbers at day 22. The percentage of lymphocytes in the NCs and that of CD(3)(+) T cells in the lymphocytes both exceeded 90% at the same time. Most of the CD(3)(+) T cells were CD(3)(+)CD(8)(+) and the percentage of CD(3)(+)CD(4)(+) T cells declined gradually. The percentage of CD(3)(+)CD(56)(+) NKT cells and gamma delta(+)T cells in the lymphocytes arised from lower than 2% to 30% approximately 40% and 10% approximately 15%, respectively. CD(3)(-)CD(56)(+) NK cells were not expanded. The cytotoxic activity of the cultured cells to K562 and Raji cells at an effector:target (E:T) ratio of 50:1 was over 75% and about 32% approximately 65%, respectively.
CONCLUSIONThe most efficient culture system which can induce CB MNC to differentiate into mature T/NK cells in vitro is the cytokines cocktail of SCF + FLT-3L + IL-7 + IL-15 + TNF-alpha + IL-2, and the optimum culture time is 22 days.