Mechanisms of wound closure following acute arm injury in Octopus vulgaris

Background: Octopoda utilise their arms for a diverse range of functions, including locomotion, hunting, defence, exploration, reproduction, and grooming. However the natural environment contains numerous threats to the integrity of arms, including predators and prey during capture. Impressively, octopoda are able to close open wounds in an aquatic environment and can fully regenerate arms. The regrowth phase of cephalopod arm regeneration has been grossly described; however, there is little information about the acute local response that occurs following an amputation injury comparable to that which frequently occurs in the wild. Methods: Adult Octopus vulgaris caught in the Bay of Naples were anaesthetised, the distal 10 % of an arm was surgically amputated, and wounded tissue was harvested from animals sacrificed at 2, 6, and 24 h post-amputation. The extent of wound closure was quantified, and the cell and tissue dynamics were observed histologically, by electron microscopy, as well as using ultrasound. Results: Macroscopic, ultrasonic and ultrastructural analyses showed extensive and significant contraction of the wound margins from the earliest time-point, evidenced by tissue puckering. By 6 h post amputation, the wound was 64.0 +/- 17.2 % closed compared to 0 h wound area. Wound edge epithelial cells were also seen to be migrating over the wound bed, thus contributing to tissue repair. Temporary protection of the exposed tip in the form of a cellular, non-mucus plug was observed, and cell death was apparent within two hours of injury. At earlier time-points this was apparent in the skin and deeper muscle layers, but ultimately extended to the nerve cord by 24 h. Conclusions: This work has revealed that O. vulgaris ecologically relevant amputation wounds are rapidly repaired via numerous mechanisms that are evolutionarily conserved. The findings provide insights into the early processes of repair preparatory to regeneration. The presence of epithelial, chromatophore, vascular, muscle and neural tissue in the arms makes this a particularly interesting system in which to study acute responses to injury and subsequent regeneration.