Induction of Intracellular Calcium Concentration by Environmental Benzo(a)pyrene Involves a β2-Adrenergic Receptor/Adenylyl Cyclase/Epac-1/Inositol 1,4,5-Trisphosphate Pathway in Endothelial Cells

Polycyclic aromatic hydrocarbons (PAHs) such as benzo-(a)pyrene (B(a)P) are widely distributed environmental contaminants, known as potent ligands of the aryl hydrocarbon receptor (AhR). These chemicals trigger an early and transient increase of intracellular calcium concentration ([Ca2+](i)), required for AhR-related effects of PAHs. The mechanisms involved in this calcium mobilization were investigated in the present study. We demonstrated that B(a)P-mediated [Ca2+](i) induction was prevented in endothelial HMEC-1 cells by counteracting beta 2-adrenoreceptor (beta 2ADR) activity using pharmacological antagonists, anti-beta 2ADR antibodies, or siRNA-mediated knockdown of beta 2ADR expression; by contrast, it was strongly potentiated by beta 2ADR overexpression in human kidney HEK293 cells. B(a)P was shown, moreover, to directly bind to beta 2ADR, as assessed by in vitro binding assays and molecular modeling. Pharmacological inhibition and/or siRNA-mediated silencing of various signaling actors acting downstream of beta 2ADR in a sequential manner, such as G protein, adenylyl cyclase, Epac-1 protein, and inositol 1,4,5-trisphosphate (IP3)/IP3 receptor, were next demonstrated to prevent B(a)P-induced calcium signal. Inhibition or knockdown of these signaling elements, as well as the use of chemical beta-blockers, were finally shown to counteract B(a)P-mediated induction of cytochrome P-450 1B1, a prototypical AhR target gene. Taken together, our results show that B(a)P binds directly to beta 2ADR and consequently utilizes beta 2ADR machinery to mobilize [Ca2+](i),through activation of a G protein/adenylyl cyclase/cAMP/Epac-1/IP3 pathway. This beta 2ADR-dependent signaling pathway activated by PAHs may likely be crucial for PAH-mediated up-regulation of AhR target genes, thus suggesting a contribution of beta 2ADR to the health-threatening effects of these environmental pollutants.