The paucity of mammalian adult cardiac myocytes (CM) proliferation following myocardial infarction (MI) and the remodeling of the necrotic tissue that ensues, result in non-regenerative repair. In contrast, zebrafish (ZF) can regenerate after an apical resection or cryoinjury of the heart. There is considerable interest in models where regeneration proceeds in the presence of necrotic tissue. We have developed and characterized a cautery injury model in the giant danio (GD), a species closely related to ZF, where necrotic tissue remains part of the ventricle, yet regeneration occurs. By light and transmission electron microscopy (TEM), we have documented four temporally overlapping processes: 1) a robust inflammatory response analogous to that observed in MI, 2) concomitant proliferation of epicardial cells leading to wound closure, 3) resorption of necrotic tissue and its replacement by granulation tissue, 4) regeneration of the myocardial tissue driven by 5-EDU and [3H]thymidine incorporating CMs. In conclusion, our data suggest that the GD possesses robust repair mechanisms in the ventricle, and can serve as an important model of cardiac inflammation, remodeling and regeneration.

The remarkable ability of the heart to regenerate has been demonstrated in the zebrafish and giant danio, two fish members of the cyprinid family. Here we use light and electron microscopy to examine the repair response in the heart of another cyprinid, the goldfish (Carassius auretus), following cautery injury to a small portion of its ventricular myocardium. We observed a robust inflammatory response in the first two weeks consisting primarily of infiltrating macrophages, heterophils, and melanomacrophages. These inflammatory cells were identified in the lumen of the spongy heart, within the site of the wound, and attached to endocardial cells adjacent to the site of injury. Marked accumulation of collagen fibers and increased connective tissue were also observed during the first and second week in a transition zone between healthy and injured myocardium as well as in adjacent sub-epicardial regions. The accumulation of collagen and connective tissue however did not persist. The presence of capillaries was also noted in the injured area during repair. The replacement of the cauterized region of the ventricle by myocardial tissue was achieved by 6 weeks. The presence of ethynyl deoxyuridine-positive cardiac myocytes and partially differentiated cardiac myocytes during repair suggest effective cardiac myocyte driven regeneration mechanisms also operate in the injured goldfish heart, and are similar to those observed in zebrafish and giant danio. Our data suggest the ability for cardiac regeneration may be widely conserved among cyprinids.

We have studied the characteristics of the heart in the adult Giant danio (Danio aequipinnatus), the wound repair response and the ability to regenerate following cautery injury. Electron microscopy studies revealed cardiac myocyte ultrastructure and organization similar to that observed in the zebrafish (Danio rerio). Apical cauterization resulted in injury to 25% of the ventricle. The injury initiated a robust inflammatory response with early recruitment of heterophils and mast cells, and the persistence of myeloperoxidase‐positive cells beyond the first week post‐injury. Bromodeoxyuridine and proliferating cell nuclear antigen‐positive cells, including endothelial cells and cardiac myocytes, were present in areas adjacent to the site of injury during the repair process, but primarily in adjacent subepicardial compact heart region by the second week. We observed marked collagen accumulation on day 14 that persisted through day 45. The collagen accumulation did not result in permanent fibrosis but appears to be resolved by 60 days when the ventricle was completely regenerated. Our data suggest that the Giant danio possesses robust repair mechanisms in the ventricle and may serve as an important model of cardiac inflammation, remodeling and regeneration.

Heart disease remains the number one cause of death in the United States. Mammalian and non‐mammalian models are used to study heart healing and regeneration following injury. In the teleost zebrafish, the removal of ventricular tissue by resection is fully replaced by thirty days with little to no scarring. We hypothesized that the goldfish (Carassius auratus) is able to regenerate its heart in a manner analogous to the zebrafish. We have performed cautery injury of 25% of the ventricle of goldfish that resulted in significant destruction of the ventricular wall and ventricular myocytes. We found an intense inflammatory response in the area of injury, followed the recruitment or proliferation of cells 3 days post injury. We also found a marked increased in collagen deposition at 3 days and 7 days that is resolved by 21 days. This observation is in contrast to the lack of collagen deposition reported in other models. Our data suggests that following cautery injury the goldfish ventricular wall is rebuilt by one week and a quasi complete healing is observed by 21 days. This is the first study showing healing following cautery injury in a fish heart.FundingDePauw University

A remarkable ability for heart regeneration has been demonstrated in the zebrafish and giant danio, two members of the cyprinids family. While regenerative capacities appear widely distributed within genera, they can vary significantly in closely related species. Here we describe using light and electron microscopy the injury response and repair mechanisms in the heart of another cyprinid, the goldfish, following cautery injury to a small portion of the ventricular myocardium. We observed a robust inflammatory response in the first week consisting of infiltrating macrophages and heterophils. These inflammatory cells were identified in the lumen of spongy heart and in the compact layer of myocardium both adjacent and remote to the injury. Marked accumulation of collagen was observed during the second week in a border zone between healthy and injured myocardium, in adjacent sub‐epicardial regions, and was concurrent with a robust angiogenic response. The replacement of the ablated region of the ventricle was achieved by six weeks post‐cautery. PCNA‐positive cardiac myocytes (CM), as well as apparent de‐differentiated cardiac CM were present during repair. This data suggest that effective regeneration mechanisms operate in the injured goldfish heart, similar to that demonstrated in zebrafish and giant danio, and that cardiac regeneration may be widespread among cyprinids.Grant Funding Source: none