Host: The Japan Radiation Research Society, Chairman of the 52nd Annual Meeting, Toshiteru Okubo (Radiation Effects Research Foundation)
Purpose: The thermo-enhancement effects of the sesquiterpene lactone parthenolide (PTL), which targets transcription factor NF-κB, with hyperthermia at 40, 42 and 44degreeC on the human lung adenocarcinoma A549 cell line were investigated in vitro. The expressions of p53 and hsp72 proteins following application of PTL, hyperthermia at 44degreeC, and those in combination were examined to investigate whether p53 and hsp72 participated in apoptosis induction via NF-κB signal pathway. The kinetics of apoptosis induction was also determined.
Methods: PTL was dissolved in culture medium to an appropriate final concentration (0.2microM) prior to being used for treatment. Kinetic assessments of the sensitivity of A549 cells to PTL and hyperthermia were carried out using colony forming assays. The accumulation of p53 and hsp72 proteins in cells after the application of hyperthermia, PTL, and those in combination were examined by western blot analysis. The induction of apoptosis in cells was analyzed under a fluorescence microscope, and the percentage of typical apoptotic cells was calculated.
Results: Thermotherapy using a combined treatment with PTL prior to hyperthermia showed synergistic thermo-enhancement effects toward A549 cells. The p53 was increased slightly by PTL. That after combination therapy with PTL and hyperthermia at 44degreeC was a little increase in comparison with hyperthermia alone. The hsp72 after treatment with PTL alone was little increase as same as a control, and also for combination therapy was not induced or reduced significantly as compared with hyperthermia alone. PTL showed remarkable cytotoxicity at S phase of cell cycle on any concentrations as well as hyperthermia. That is, PTL reduced the cells of proliferation phase. PTL alone did not inhibit NF-κB activation in the cells stimulated with TNF-alpha, unexpectedly. While heating alone inhibited NF-κB for early interval time after treatment, but faded for long time of interval. However, the combined treatments with PTL and hyperthermia inhibited NF-κB activation completely.
Conclusions: Thermotherapy of combined treatment with PTL and hyperthermia in A549 cells showed synergistic thermo-enhancement effects. It was suggested that the PTL-induced apoptosis in A549 cells due to suppress the activity of NF-κB directly, and via p53-independent and hsp72-independent manner on NF-κB signaling.
