The radiation-induced bystander effect is defined as "the induction of biological effects in cells that are not directly traversed by a charged particle but are in close proximity to cells that are." Although these bystander effects have been demonstrated with a variety of biological endpoints in both human and rodent cell lines (as well as in 3D tissue samples), the mechanism of the phenomenon is not known. Although gap junction communication and the presence of soluble mediator(s) are both known to play important roles in the bystander response, the precise signaling molecules have yet to be identified. By using the Columbia University charged particle beam in conjunction with a strip dish design, we show here that the cyclooxygenase-2 (COX-2, also known as prostaglandin endoperoxide synthase-2) signaling cascade plays an essential role in the bystander process. Treatment of bystander cells with NS-398, which suppresses COX-2 activity, significantly reduced the bystander effect. Because the critical event of the COX-2 signaling is the activation of the mitogen-activated protein kinase pathways, our finding that inhibition of the extracellular signal-related kinase phosphorylation suppressed bystander response further confirmed the important role of mitogen-activated protein kinase signaling cascade in the bystander process. These results provide evidence that the COX-2-related pathway, which is essential in mediating cellular inflammatory response, is the critical signaling link for the bystander phenomenon.