Inducing the Anti-Leukemia Effects of CD8⁺ T Cytotoxic Lymphocytes Derived from Umbilical Cord Cell.

Objective: The previous studies of our research group have proved that specific cytotoxic T lymphocytes(CTLs) can be generated from umbilical cord cells. The purpose of this study is to further demonstrate whether CD8⁺CTLs can be generated from cord blood (CB) and kill leukemia cells specifically ex vivo. And whether the leukemia cytotoxicity of CD8⁺CTLs and CD8⁻CTLs are different.

Method: First we induce adherent cells collecting from CB samples to generate dendritic cells (DCs) by culturing with GM-CSF, IL-4 and load lysate antigen of U937 cell to immature DCs. On day 6^th, TNF-αand PGE₂ was added to accelerate the maturation of DCs. After co-culturing DCs with the CTLs from the same CB, then we used MidiMACS to isolate CD8⁺ and CD8⁻ T cell. The cytotoxicity of the CD8⁺ CTLs and CD8⁻⁻CTLs to U937 cell line was evaluated with Methyl thiazolyl tetrazolium(MTT) assay.

Result: The purity of CD8⁺CTL was(95.73±1.50)%. Among the CD8⁻CTLs, the percentage of CD4⁺CTLs was (65.01±9.29)% and CD8⁺T lymphocytes was (4.9±4.46)%. When E:T ratios were 40:1, 20:1 and 10:1, the mean cytotoxic rate (MCR)of CD8⁺ CTLs to U937 cell were (66.36±12.43)%,(35.38±9.64)% and(19.04±6.15)% respectively; the MCR of CD8⁻CTLs to U937 cell were (34.47±8.19)%,(21.85±7.06)% and(12.26±4.87)% and the MCR of T cell from CB to U937 cell were(15.79±4.64)%,(9.6±3.71)% and (5.69±3.14)%. The MCR of CD8⁺ CTLs to U937 cell were significant higher than those of CD8⁻CTLs and T cell groups at the same E:T ratios(P<0.05). When E:T ratios was 40:1,the MCR of CD8⁺CTLs and CD8⁻CTLs to K562 cell line were (36.77±10.24)% and (26.95±3.06)% respectively, the cytotoxicity effect of CD8⁺ and CD8⁻CTLs to K562 cell are no difference (P>0.05).

Conclusions: The cytotoxicity of CD8⁺CTLs against U937 cell lines is more potent than that of CD8⁻CTLs, and this effect is specific.