Phorbol ester (12-O-tetradecanoylphorbol 13-acetate) prevents ornithine decarboxylase inhibition and apoptosis and L1210 leukemic cells exposed to TGF-beta 1.

Previous studies have shown that growth suppression and apoptosis of leukemic cells exposed to TGF-beta 1 is associated with the inhibition of ornithine decarboxylase (ODC)--the key enzyme of polyamine pathway. The aim of the present study was to evaluate the influence of 12-O-tetradecanoylphorbol 13-acetate (TPA)--a potent ODC inducer on antiproliferative and apoptotic effects of TGF-beta 1 in L1210 leukemic cells. Cells were incubated in 2% FCS/RPMI-1640 medium, supplemented with TGF-beta 1 (2 ng/ml). TPA (100 ng/ml) or alpha-difluoromethylornithine (DFMO) (5 mM). Cell proliferation, apoptosis and necrosis were evaluated using [methyl-3H] thymidine, electron microscopy, electrophoresis of DNA and trypan blue exclusion. Expression and activity of ODC were determined by RT-PCR and measurement of 14CO2 release from L-1-14C ornithine, respectively. TGF-beta 1 inhibited proliferation and induced apoptotic and necrotic cell death in L1210 leukemic cells. The above effects were associated with the inhibition of ODC expression and activity, measured 2 and 4 hr after TGF-beta 1 administration, respectively. The presence of DFMO, an irreversible inhibitor of ODC, led to apoptotic fragmentation of DNA, similar to that observed in TGF-beta 1-treated cultures. Administration of TPA simultaneously with TGF-beta 1 significantly reduced antiproliferative, apoptotic and necrotic effects of TGF-beta 1, and prevented its inhibitory action of ODC expression and activity. It is concluded that: down-regulation of ODC expression may be one of the early events associated with TGF-beta 1-evoked suppression of growth and apoptosis; ODC is involved in the mechanism of protective action of TPA on TGF-beta 1-related growth inhibition of L1210 leukemic cells.