49 - Inhibition of Amputation-Induced Reactive Oxygen Species Blocks Salamander Tail Regeneration

Abstract

Very little is known about signaling mechanisms that initiate and sustain tissue regeneration responses after injury. Here, we investigated the requirement of reactive oxygen species (ROS) and NOX activity for tail regeneration using a Mexican axolotl (Ambystoma mexicanum) embryo model. In vivo imaging using dihydroethidium (DHE) revealed a marked increase of ROS and superoxide anions O2●– only minutes after tail amputation. Surprisingly, ROS production was observed at 48 hours post amputation (hpa), long after wound healing. To establish that ROS production is dependent upon early NAPDH oxidase activity, we treated embryos with NOX inhibitor VAS2870 and observed a significant reduction in ROS at 6 hpa. We also observed that VAS2870 strongly inhibited tail regeneration at 72 hpa and 168 hpa. Our findings show that axolotl tail amputation induces ROS production at the amputation site and NOX activity is required for successful tail regeneration. Given the requirement of ROS production for Xenopus tadpole tail and zebrafish fin regeneration, our results suggest that ROS is a conserved mechanism of appendage regeneration.