[Recombinant soluble CD40 ligand enhances Wogonin-induced antitumor activity].

OBJECTIVE

To investigate the effect of recombinant soluble CD40 ligand (rsCD40L) on Wogonin mediated antitumor activity in cancer cells and the underlying molecular mechanisms.

METHODS

Cell death was detected based on the release of lactate dehydrogenase (LDH) using a cytotoxicity detection kit. For morphological study of cell death, cells were stained with 50 microg/mL of acridine orange and 50 microg/mL of ethidium bromide and observed and photographed under a fluorescence microscope. Activation of apoptosis pathway was evaluated by Western blot. The effects of pan-caspase inhibitor Z-VAD-FMK and tumor necrosis factor alpha (TNF-alpha) neutralizing antibody on cell death induced by rsCD40L and Wogonin co-treatment were also investigated.

RESULTS

rsCD40L significantly enhanced Wogonin-induced cell death of ovarian cancer cells SKOV3. A dose-dependent synergism was found with a fixed rsCD40L dose (1 microg/mL) and increased concentrations of Wogonin (5 micromol/L-15 micromol/L). rsCD40L and Wogonin co-treated cells showed typical apoptotic morphologies and enhanced activation of caspases pathway. As expected, the pan-caspase inhibitor Z-VAD-FMK inhibited synergistic cell death of rsCD40L and Wogonin co-treated SKOV3 cells. Interestingly, the TNF-alpha neutralizing antibody that blocks TNF-alpha binding to its receptor also significantly suppressed the cell death enhancing effect, indicating that autocrine TNF-alpha played a role of sensitization.

CONCLUSIONS

rscCD40L sensitizes cancer cells to wogonin-mediated apoptosis, which may involve autocrine of TNF-alpha, and the combination of rsCD40L and Wogonin may have a potential for cancer therapy.