No abstract

Inhibition of gap junctional intercellular communication (GJIC) and the activation of intracellular mitogenic pathways are common hallmarks of epithelial derived cancer cells. We previously determined that the 1-methyl and not the 2-methyl isomer of anthracene, which are prominent cigarette smoke components, activated extracellular receptor kinase, and inhibited GJIC in WB-F344 rat liver epithelial cells. Using these same cells, we show that an immediate upstream response to 1-methylanthracene was a rapid (<1 min) release of arachidonic acid. Inhibition of phosphatidylcholine-specific phospholipase C prevented the inhibition of GJIC by 1-methylanthracene. In contrast, inhibition of phosphatidylinositol specific phospholipase C, phospholipase A 2 , diacylglycerol lipase, phospholipase D, protein kinase C, and tyrosine protein kinases had no effect on 1-methylanthracene-induced inhibition of GJIC. Inhibition of protein kinase A also prevented inhibition of GJIC by 1-methylanthracene. Direct measurement of phosphatidylcholine-specific phospholipase C and sphingomyelinase indicated that only phosphatidylcholinespecific phospholipase C was activated in response to 1-methylanthracene, while 2-methylanthracene had no effect. 1-methylanthracene also activated p38-mitogen activated protein kinase; however, like extracellular kinase, its activation was not involved in 1-methylanthracene-induced regulation of GJIC, and this activation was independent of phosphatidylcholine-specific phospholipase C. Although mitogen activated protein kinases were activated, Western blot analyzes indicated no change in connexin43 phosphorylation status. Our results indicate that phosphatidylcholinespecific phospholipase C is an important enzyme in the induction of a tumorigenic phenotype, namely the inhibition of GJIC; whereas mitogen activated protein kinases triggered in response to 1-methylanthracene, were not involved in the deregulation of GJIC. (Cancer Sci 2008; 99: 696-705)

Dysregulation of gap junctional intercellular communication (GJIC) has been associated with different pathologies, including cancer; however, molecular mechanisms regulating GJIC are not fully understood. Mitogen Activated Protein Kinase (MAPK)-dependent mechanisms of GJIC-dysregulation have been well-established, however recent discoveries have implicated phosphatidylcholine-specific phospholipase C (PC-PLC) in the regulation of GJIC. What is not known is how prevalent these two signaling mechanisms are in toxicant/toxin-induced dysregulation of GJIC, and do toxicants/toxins work through either signaling mechanisms or both, or through alternative signaling mechanisms. Different chemical toxicants were used to assess whether they dysregulate GJIC via MEK or PC-PLC, or both Mek and PC-PLC, or through other signaling pathways, using a pluripotent rat liver epithelial oval-cell line, WB-F344. Epidermal growth factor, 12-O-tetradecanoylphorbol-13-acetate, thrombin receptor activating peptide-6 and lindane regulated GJIC through a MEK1/2-dependent mechanism that was independent of PC-PLC; whereas PAHs, DDT, PCB 153, dicumylperoxide and perfluorodecanoic acid inhibited GJIC through PC-PLC independent of Mek. Dysregulation of GJIC by perfluorooctanoic acid and R59022 required both MEK1/2 and PC-PLC; while benzoylperoxide, arachidonic acid, 18β-glycyrrhetinic acid, perfluorooctane sulfonic acid, 1-monolaurin, pentachlorophenol and alachlor required neither MEK1/2 nor PC-PLC. Resveratrol prevented dysregulation of GJIC by toxicants that acted either through MEK1/2 or PC-PLC. Except for alachlor, resveratrol did not prevent dysregulation of GJIC by toxicants that worked through PC-PLC-independent and MEK1/2-independent pathways, which indicated at least two other, yet unidentified, pathways that are involved in the regulation of GJIC. In conclusion: the dysregulation of GJIC is a contributing factor to the cancer process; however the underlying mechanisms by which gap junction channels are closed by toxicants vary. Thus, accurate assessments of risk posed by toxic agents, and the role of dietary phytochemicals play in preventing or reversing the effects of these agents must take into account the specific mechanisms involved in the cancer process.