Attenuation of caspase-3-dependent apoptosis by Trolox post-treatment of X-irradiated MOLT-4 cells.

PURPOSE

The relationship between post-irradiation treatment with Trolox, an antioxidant that inhibits lipid peroxidation, and X-ray-induced apoptosis, with regard to signal transduction pathways, was examined in MOLT-4, a human leukaemia cell line.

MATERIALS AND METHODS

In MOLT-4 cells treated with Trolox after X-irradiation, viability, DNA fragmentation, expression of p53, BCL-2, BAX, active SAPK/JNK, active caspase-3 and the cleavage of PARP were measured by the trypan blue exclusion test, agarose gel electrophoresis and Western blotting.

RESULTS

Stained cells and ladder-like DNA cleavage were observed after X-irradiation. Cell death and DNA fragmentation were significantly inhibited by the post-irradiation treatment with Trolox. The expression of p53 and active SAPK/JNK was increased after X-irradiation, and fragments of PARP and the activated fragment of caspase-3 were produced. Post-irradiation treatment with Trolox attenuated the X-irradiation-induced expression, fragmentation or activation of these apoptosis-related biomolecules. The expression of BCL-2 and BAX, which would occur downstream from p53, was not changed by irradiation and Trolox treatment. Furthermore, cell death was associated with caspase-3 because the ladder-like DNA cleavage was completely inhibited by Ac-DEVD-CHO but not Ac-YVAD-CHO, TLCK and PMSF.

CONCLUSION

Post-irradiation events such as membrane damage induce caspase-3-dependent apoptosis, which might be mediated by the activation of SAPK/JNK and be independent of p53.