Effect of reactive nitrogen metabolite scavengers on sensitivity of human leukemia cells to immunotherapy

Abstract

Objective To investigate the effect of Tiopronin (TIP) on interleukin (IL)-2 immunotherapy of human leukemia KG-1 cells and its possible mechanism. Methods KG-1 cells in logarithmic growth phase were randomly divided into KG-1+ IL-2 group and KG-1+ IL-2+ TIP group. Methyl thiazolyl tetrazolium (MTT) assay and colony formation assay were used to detect the sensitivity and proliferation of KG-1 cells. The changes of reactive nitric metabolites (RNM) were detected with nitrate reductase method. The production of tumor necrosis factor (TNF)-β and interferon (IFN)-γ was detected with enzyme linked immunosorbent assay (ELISA). The expression of CD3ζ was detected with Western blot and real time polymerase chain reaction (RT-PCR). Results IL-2 and IL-2+ TIP could inhibit the growth of KG-1 cells. The inhibitory rate of KG-1+ IL-2+ TIP group was significantly higher than that of KG-1+ IL-2 group, and the sensitivity of KG-1 cells to IL-2 was 6.2 times higher. Both IL-2 and IL-2+ TIP group inhibited the colony formation of KG-1 cells. Compared to KG-1+ IL-2 group, KG-1+ IL-2+ TIP group inhibited the colony formation of KG-1 cells by 3.5 times. The RNM production of KG-1 + IL-2 group was (158.26±3.82) μmol/ml, which was significantly higher than (45.18±4.29) μmol/ml of KG-1 + IL-2 + TIP group (P<0.05). The levels of TNF-β and IFN-γ in KG-1+ IL-2+ TIP group were (253.28±7.84) pg/ml and (181.25±6.41) pg/ml, which was significantly higher than (98.45±6.43) pg/ml and (68.74±8.26) pg/ml of KG-1+ IL-2 group (P<0.05). The expression of CD3ζ in KG-1+ IL-2+ TIP group was significantly higher than that in KG-1+ IL-2 group. Conclusions Tiopronin can promote NK/T cell activity and increase the sensitivity of leukemia KG-1 cells to IL-2 by eliminating reactive nitrogen metabolites. Key words: Thiopronine/PD; Leukemia/TH; Immunotherapy