Role of prostanoids and lipid peroxides as mediators of the 12-O-tetradecanoylphorbol-13-acetate effect on cell growth.

Confluent cultures of guinea pig smooth muscle cells (SMC) or human fibroblasts (HNF) were treated with 12-tetradecanoylphorbol-13-acetate (TPA). Prostanoid levels were measured by the radioimmunoassay of 6-keto-PGF1 alpha and PGE2, and lipid peroxides were measured by the thiobarbituric acid test for malondialdehyde (MDA). Cells were seeded at low densities, and growth was calculated both from the cell count (Coulter Counter) and the colony number (image analysis). When confluent SMC and HNF were incubated in media alone, 6-keto-PGF1 alpha levels were a function of the TPA concentration, increasing to a maximum at 10(-8) M TPA and then decreasing at higher TPC concentrations. When confluent SMC and HNF were incubated in media containing exogenous arachidonic acid, 6-keto-PGF1 alpha levels again increased to a maximum at 10(-8) M TPA but decreased at higher TPA concentrations only with SMC. The increase in 6-keto-PGF1 alpha levels was much greater in HNF (1310%) than SMC (680%). SMC synthesized similar amounts of 6-keto-PGF1 alpha and PGE2, and the stimulatory effect of TPA was similar with 6-keto-PGF1 alpha and PGE2. Indomethacin (IM) blocked prostanoid synthesis at all TPA concentrations. TPA did not have a significant effect on MDA levels in either cell line. The lipid antioxidants alpha-tocopherol and alpha-tocopherylquinone blocked lipid peroxidation without affecting the stimulation of prostanoid synthesis with TPA. Cell number decreased to a minimum at 10(-8) M TPA in both cell lines. The decrease in cell number was much greater in HNF (72%) than SMC (30%). SMC colony number decreased at 10(-8) TPA and then increased at 10(-6) M TPA. IM did not block the TPA effect on cell number in either cell line.(ABSTRACT TRUNCATED AT 250 WORDS)