The P2Y2receptor mediates the epithelial injury response and cell migration

Abstract

Injury to epithelial cells results in the release of ATP and stimulation of purinergic receptors and is thought to alter cell migration and wound repair. Medium from the injured cells triggers Ca2+mobilization and phosphorylation of ERK, both of which are inhibited if the medium is pretreated with apyrase. To understand the wound repair mechanism that occurs with injury, our goal was to determine which purinergic receptor(s) was the critical player in the wound response. We hypothesize that the P2Y2receptor is the key player in the response of corneal epithelial cells to cell damage and subsequent repair events. Cells transfected with short interfering RNA to either P2Y2or P2Y4were stimulated either by injury or addition of UTP and imaged using fluo 3-AM to monitor changes in fluorescence. When cells with downregulated P2Y2receptors were injured or stimulated with UTP, the intensity of the Ca2+release was reduced significantly. However, when cells with downregulated P2Y4receptors were stimulated, only the UTP-induced Ca2+response was reduced significantly. In addition, downregulation of the P2Y2receptor inhibited wound closure compared with unstimulated cells or cells transfected with nontargeting sequence. This downregulation resulted also in an attenuation in phosphorylation of Src and ERK. Together, these data indicate that the P2Y2receptor plays a major biological role in the corneal injury response and repair mechanisms.