Mechanistic studies on the DDT-induced inhibition of intercellular communication

Two structurally unrelated compounds, 1,1'-(2,2,2-trichloroethylidene) bis(4-chlorobenzene) (DDT) and 12-O-tetradecanoylphorbol-13-acetate (TPA), are both potent inhibitors of cell-cell communication in vitro as well as tumour promoters in vivo. There is evidence that TPA acts via a specific receptor mechanism involving activation of protein kinase C (pkC). The mechanism of action of DDT has been discussed in terms of membrane perturbation, increased intracellular calcium, interaction with calmodulin and decreased cAMP levels. In the present study the objective was to examine the potential role of pkC activation in DDT-induced inhibition of intercellular communication in cultured cells. The V79 metabolic cooperation assay was used for measuring intercellular communication. Furthermore, the effects of DDT on the activity of partially purified pkC from V79 cells was measured, as was the interaction of DDT with the phorbol ester/DAG-binding site on the pkC enzyme. Results from the biochemical studies showed that DDT neither activates pkC nor binds to the phorbol ester/DAG-binding site, as measured by displacement of PDBU binding. Using the metabolic cooperation assay it was demonstrated that pretreatment with TPA made cells refractory, i.e. a second application of TPA did not inhibit cell-cell communication. DDT added to cells down-regulated with TPA inhibited cell-cell communication, even though these cells were refractive to TPA. This result further supports the hypothesis that DDT and TPA inhibit intercellular communication primarily by different pathways. At non-cytotoxic concentrations, pkC inhibitors (H7, W7 and palmitoyl carnitine) did not affect the TPA- or DDT-induced inhibition of cell-cell communication in the V79 metabolic cooperation assay. Quercetin, a pkC inhibitor which has been reported to eliminate DDT- or TPA-induced inhibition of intercellular communication, was investigated in an in vivo study that measured promotion of enzyme-altered foci in DEN-treated rat liver. Quercetin co-administered with DDT did not act as an antipromoter.