Investigating the mechanisms underlying myocardial regeneration in the Mexican cavefish

Following myocardial infarction (MI), the adult human heart is unable to regenerate and replace the lost cardiac muscle with new functioning cardiomyocytes. Instead, it forms a permanent non-contractile scar that impairs cardiac function, which may later result in heart failure. In contrast, the adult zebrafish and neonatal mouse heart possess an innate ability to regenerate their heart after injury, with studies revealing myocardial proliferation in the border zone next to the injury site as a key driver to the regenerative process.
The Astyanax mexicanus is a unique cardiac regeneration fish model that enables the comparison of adult regeneration with human-like scarring all within the same species. In this model, the Astyanax surface fish regenerate their heart after injury, while their Pachón cave-dwelling counterparts cannot, and similar to humans, form a permanent scar. These opposing regenerative capacities coincide with contrasting myocardial proliferation profiles, which are characterised by initiation of cardiomyocyte proliferation in both Pachón and surface fish, but no observable cardiomyocyte expansion in Pachón.
To determine the capability of the Astyanax model to detect novel genes in regeneration and decipher the mechanisms behind these contrasting myocardial regeneration capacities, comparative bulk and single cell transcriptomics were employed. Bulk transcriptome analysis identified upregulation of lrrc10, a novel gene to regeneration, in the surface fish compared to Pachón heart. CRISPR mediated knockout of lrrc10 in zebrafish impaired heart regeneration, with lrrc10-/- zebrafish displaying a Pachón-like phenotype, confirming the ability of the Astyanax model to detect novel genes.
Targeted analysis of the cardiomyocytes using single cell transcriptomics revealed myocardial heterogeneity in Astyanax hearts, with a transcriptionally distinct myocardial border zone after injury. This highly proliferative border zone generally possessed characteristics consistent with reduced maturity, dedifferentiation, cell proliferation and metabolic reprogramming in both surface fish and Pachón. However, differential expression of cell cycle genes between surface fish and Pachón border zone cardiomyocytes alongside cardiomyocyte DNA quantification suggested cell cycle arrest and polyploidisation in Pachón proliferating cardiomyocytes at the expense of meaningful cardiomyocyte expansion. The finding of polyploidisation in Pachón coincided with lack of upregulation of glucose uptake and glycolytic genes as observed in the surface fish myocardial border zone. In contrast, evidence suggested increased fatty acid uptake/utilisation and metabolic inflexibility after injury in the Pachón heart – all characteristics consistent with the diabetic heart. However, the insra P211L mutation, which is responsible for insulin resistance in Pachón, does not have a negative impact on cardiac regeneration.
The Astyanax is a model capable of enhancing our understanding of regeneration and identification of the above metabolic differences provides the initial strides into establishing the metabolic fingerprint required for successful cardiomyocyte proliferation during cardiac regeneration.

Keywords: Zebrafish
Astyanax Mexicanus
Metabolism
Cavefish
Cardiomyocyte
Heart regeneration
Advisor: Mommersteeg, M
Riley, P