Connexins and cyclooxygenase-2 crosstalk in the expression of radiation-induced bystander effects

Background: Signalling events mediated by connexins and cyclooxygenase-2 (COX-2) have important roles in bystander effects induced by ionising radiation. However, whether these proteins mediate bystander effects independently or cooperatively has not been investigated. Methods: Bystander normal human fibroblasts were cocultured with irradiated adenocarcinoma HeLa cells in which specific connexins (Cx) are expressed in the absence of endogenous Cx, before and after COX-2 knockdown, to investigate DNA damage in bystander cells and their progeny. Results: Inducible expression of gap junctions composed of connexin26 (Cx26) in irradiated HeLa cells enhanced the induction of micronuclei in bystander cells (P<0.01) and reduced the coculture time necessary for manifestation of the effect. In contrast, expression of connexin32 (Cx32) conferred protective effects. COX-2 knockdown in irradiated HeLa Cx26 cells attenuated the bystander response due to connexin expression. However, COX-2 knockdown resulted in enhanced micronucleus formation in the progeny of the bystander cells (P<0.001). COX-2 knockdown delayed junctional communication in HeLa Cx26 cells, and reduced, in the plasma membrane, the physical interaction of Cx26 with MAPKKK, a controller of the MAPK pathway that regulates COX-2 and connexin. Conclusions: Junctional communication and COX-2 cooperatively mediate the propagation of radiation-induced non-targeted effects. Characterising the mediating events affected by both mechanisms may lead to new approaches that mitigate secondary debilitating effects of cancer radiotherapy.