After myocardial infarction, the adult mammalian heart fails to regenerate, while newborn mammals and adult zebrafish regenerate their damaged heart through robust cardiomyocyte proliferation. One of the factors that determines the regenerative competence of the heart is the DNA content (ploidy) of the cardiomyocytes. During postnatal growth, most cardiomyocytes in mammals become polyploid. Only a small fraction of cardiomyocytes remains diploid in adults. This population reactivates after injury, although its contribution is insufficient to compensate for the loss of millions of cardiomyocytes. By contrast, zebrafish cardiomyocytes remain diploid and regeneration-competent even in adulthood. We have previously demonstrated that experimental polyploidization of the myocardium dampens the regenerative competence of the zebrafish heart. However, the genetic mechanisms that regulate the transition from diploid to polyploid myocardium remain largely unknown.

Previous work in mice identified the myocardial-specific kinase Tnni3k as a gene involved in cardiomyocyte polyploidization. Mice lacking Tnni3k expression have an increased proportion of diploid cardiomyocytes and an augmented proliferative response after injury. Conversely, overexpression of mouse Tnni3k in zebrafish increased nuclear ploidy and impaired regeneration. Since the zebrafish genome includes a highly conserved copy of tnni3k, we sought to determine whether species-specific differences in this gene are responsible for the differences in ploidy between zebrafish and mice. Using fluorescent in situ hybridization, we first learned that tnni3k is expressed in the adult zebrafish myocardium and upregulated in response to injury. Next, we found that overexpression of either mouse or zebrafish tnni3k resulted in a modest increase in the proportion of polyploid cardiomyocytes. In contrast to previous reports, we did not detect a reduction in cardiomyocyte proliferation after injury in animals overexpressing tnni3k. However, histological analysis revealed that overexpression of tnni3k increased endocardial fibrosis at 7 days post-injury (dpi) and increased fibrosis retention at 60 dpi. This fibrotic response was associated with an elevation in the levels of phospho-p38, a stress kinase previously associated with fibrosis in the heart. Conversely, deletion of the tnni3k locus resulted in a reduction in phospho-p38 levels and fibrosis deposition after injury. Our results suggest that tnni3k determine regenerative competence in zebrafish by regulating fibrosis rather than affecting cardiomyocyte ploidy.