| Background A chromosomal abnormality affecting the 11q23 chromosome region is common in hemotopoietic tumors, seen in 40-70% of infant acute leukemia(IAL). It specifically results in the rearrangement of the mixed-lineage leukemia (MLL) gene, including translocation, deletion and duplicate. Most pediatric leukemias with MLL rearrangement have a high malignancy, remarkably short latency and allo-hemopoietic stem cell transplantation is not likely to improve the poor prognosis. TRAIL (tumor necrosis factor-related apoptosis-inducing ligand) is a member of the tumor necrosis factor family. After binding with its death receptors, the apoptosis signaling pathway is activated. TRAIL is believed to be a promising candidate for cancer therapy because of its selectively cytotoxity against tumor cells. However, leukemias with 11q23 abnormality are resistant to TRAIL-induced apoptosis.In recent years, the endoplasmic reticulum stress inducers, such as tunicamycin, promoting the apoptosis of tumor cells has become a hot spot of research. Researchers have conducted large amount of research about the combination treatment of TRAIL and ionic rays, antimycotic agents, chemotherapy, genototoxicity agents and cytokines and have obtained helpful results. However, there still lacks evidence of whether the combination of tunicamycin and TRAIL can enhance the sensitivity of leukemia with 11q23 abnormality.Objective To assess the effect of tunicamycin alone and with TRAIL on the apotosis of leukemia cell line with 11q23 abnormality, discuss the possible apoptosis pathway, so as to provide theoretical foundation of the two agents.Methods t(11;19) cell line KOCL44 was used in the study. 1640 culture medium was added in the blank group, different concentration of tunicamycin or TRAIL alone was added in the negative control groups, while tunicamycin and TRAIL were both added in the experimental groups. The cell survival rate was measured by typan blue staining. Early apoptosis was detected with flow cytometry by FITC labeled Annexin-V/PI staining. The cell surface expression of death receptors was detected with flow cytometry. The protein level of Bip, CHOP and caspase-3 was detected by Western blotting so as to discuss the apoptosis pathway.Results The cell growth was inhibited by neither tunicamycin nor TRAIL alone and P>0.05 when compared with control groups. Cell growth were inhibited by combination of tunicamycin with TRAIL and P<0.05 when compared with control groups. The inhibition was dose dependent and most obvious with 2ug/ml tunicamycin plus 50ng/ml TRAIL, indicating synergistic effect of the two agents. Annexin-V/PI staining found the same tendency. Cell surface expression of DR4 and DR5 was not induced by neither tunicamycin nor TRAIL alone. DR5, but not DR4 was obviously induced by the combination of tunicamycin and TRAIL. Upon western blotting, Bip and CHOP were activated after treatment with tunicamycin plus TRAIL and the expression was time dependent. No activation of caspase-3 was found.Conclusion 1. KOCL44 is resistant to the apoptosis induced by tunicamycin or TRAIL alone.2. Tunicamycin up-regulates the cell surface expression of death receptor DR5 through Bip mediated UPR, as well as CHOP apoptosis pathway, so as to enhances the sensitivity of KOCL44 to TRAIL induced apoptosis... |
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