Non-dioxin-like polychlorinated biphenyls induce a release of arachidonic acid in liver epithelial cells: a partial role of cytosolic phospholipase A(2) and extracellular signal-regulated kinases 1/2 signalling

Abstract

Non-dioxin-like polychlorinated biphenyls (NDL-PCBs) have been shown to act as tumor promoters in liver; however, the exact mechanisms of their action are still only partially understood. One of the interesting effects of NDL-PCBs is the acute inhibition of gap junctional intercellular communication (GJIC), an effect, which has been often found to be associated with tumor promotion. As previous studies have suggested that NDL-PCB-induced disruption of lipid signalling pathways might correspond with GJIC inhibition, we investigated effects of PCBs on the release of arachidonic acid (AA) in the rat liver epithelial WB-F344 cell line, a well-established model of liver progenitor cells. We found that both 2,2',4,4'-tetrachlorobiphenyl (PCB 47) and 2,2',4,4',5,5'-hexachlorobiphenyl (PCB 153), but not the dioxin-like, non-ortho-substituted, 3,3',4,4',5-pentachlorobiphenyl (PCB 126), induce a massive release of AA. The AA release, induced by PCB 153, was partially inhibited by extracellular signal-regulated kinases 1/2 (ERK1/2) signalling inhibitor, U0126, and by cytosolic phospholipase A(2) (cPLA(2)) inhibitor, AACOCF(3). Although PCB 153 induced both ERK1/2 and p38 activation, the specific p38 kinase inhibitor, SB203580, had no effect on AA release. Inhibitors of other phospholipases, including phosphatidylcholine-specific phospholipase C or phosphatidylinositol-specific phospholipase C, were also without effect. Taken together, our findings suggest that the AA release, induced by non-dioxin-like PCBs in liver progenitor cell line, is partially mediated by cytosolic PLA(2) and regulated by ERK1/2 kinases. Our results suggest that more attention should be paid to cell signalling pathways regulated by AA or eicosanoids after PCB exposure, which might be involved in their toxic effects.

Figure 1

Effects of PCB 47, 153 and 126 on AA release from WB-F344 cells. Cells were exposed to indicated concentrations of PCBs for 30 min or 3 h, and AA levels in medium were determined by HPLC as described in Materials and Methods. Data are expressed as means ± S.D. of three independent experiments. * A significant difference between control (DMSO) and treated samples (p < 0.05). ** A significant difference between control (DMSO) and PCB-treated samples (p < 0.01).

Figure 2

AACOCF₃, inhibitor of cPLA₂, partially inhibits the PCB 153-induced release of AA. Cells were pre-treated with inhibitors and then exposed to 50 μM PCB 153 for 3 h, and AA levels in medium were determined by HPLC as described in Materials and Methods. The concentrations and respective specificities of inhibitors are summarized in Materials and Methods. Data are expressed as means ± S.D. of three independent experiments. * A significant difference between PCB 153 and PCB 153 + inhibitor (p < 0.05).

Figure 3

PCB 153 induces release of AA partly through activation of ERK1/2 signalling. (A) Rapid induction of ERK1/2 and p38 activation by PCB 153. Cells were treated with 50 μM PCB153, DMSO (control), or with EGF as a positive control for ERK1/2 activation. Cell lysates were prepared and Western blotting performed as described in Materials and Methods. The results shown here are representative of three independent experiments. Detection of total ERK1/2 and p38 expression has been performed as described in Materials and Methods to verify equal loading – no differences were observed (data not shown). (B) U0126, inhibitor of ERK1/2 activation, partially inhibits the PCB 153-induced release of AA. Cells were pre-treated with U0126 (10 μM) or SB203580 (10 μM) and then exposed to 50 μM PCB 153 for 3 h, and AA levels in medium were determined by HPLC as described in Materials and Methods. Data are expressed as means ± S.D. of three independent experiments. * A significant difference between PCB 153 and PCB 153 + inhibitor (p < 0.05).