Effects of GM-CSF, IL-3, and GM-CSF/IL-3 fusion protein on apoptosis of human myeloid leukemic cell line Tf-1 induced by irradiation.

Aim: To observe the effects of three cytokines on the apoptosis of Tf-1 cells induced by gamma irradiation and investigate the relationship between apoptosis and caspase-3 activity.

Methods: Different cytokines GM-CSF, IL-3 and GM-CS/IL-3 fusion protein were added into the irradiated Tf-1 cells. MTT assay, morphology, flow cytometry, and DNA fragmentation assay were used to observe the effects of cytokines on apoptosis. The caspase-3 activity was determined with a fluorocytometer.

Results: Irradiated Tf-1 cells showed typical morphological characteristic of apoptosis demonstrated by transmission electron microscopy and were accumulated in G0/G1 phase. In the groups treated with growth factors after irradiation, three cytokines significantly increased the viability rate, distinctly decreased the apoptosis rate and the proportion of DNA fragmentation. When Tf-1 cells were irradiated by gamma irradiation, caspase-3 activity was increased at different time points. In comparison with the control group in which no growth factor was added after the cells were irradiated, the caspase-3 activity of irradiated Tf-1 cells was significantly inhibited by addition of the above cytokines. Thirty-six hours after irradiation, in the control group, GM-CSF, IL-3, GM-CSF and IL-3 in combination, and two GM-CSF/IL-3 fusion protein groups, the apoptosis rate was 73 %, 11 %, 15 %, 13 %, 12 %, and 13 %. The percent of fragmented DNA was 36 %, 19 %, 18 %, 14 %, 13 %, and 14 %. The fluorescence intensity was 16923, 5529, 6581, 5322, 5426, and 5485.

Conclusion: GM-CSF, IL-3, and GM-CSF/IL-3 fusion protein could protect Tf-1 cells from apoptosis induced by gamma irradiation. After Tf-1 cells were irradiated, the caspase-3 activity was significantly increased but was dramatically decreased by the above cytokines. The remarkable inhibition of caspase-3 activity may be one of the mechanisms of these hematopoietic growth factors exerting their anti-apoptotic effects.