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Epicardial regeneration is guided by cardiac outflow tract and Hedgehog signalling.

Wang J, Cao J, Dickson AL, Poss KD - Nature (2015)

Bottom Line: Transplantation of Sonic hedgehog (Shh)-soaked beads at the ventricular base stimulates epicardial regeneration after bulbous arteriosus removal, indicating that Hh signalling can substitute for the influence of the outflow tract.Thus, the ventricular epicardium has pronounced regenerative capacity, regulated by the neighbouring cardiac outflow tract and Hh signalling.These findings extend our understanding of tissue interactions during regeneration and have implications for mobilizing epicardial cells for therapeutic heart repair.

View Article: PubMed Central - PubMed

Affiliation: Department of Cell Biology and Howard Hughes Medical Institute, Duke University Medical Center, Durham, North Carolina 27710, USA.

ABSTRACT
In response to cardiac damage, a mesothelial tissue layer enveloping the heart called the epicardium is activated to proliferate and accumulate at the injury site. Recent studies have implicated the epicardium in multiple aspects of cardiac repair: as a source of paracrine signals for cardiomyocyte survival or proliferation; a supply of perivascular cells and possibly other cell types such as cardiomyocytes; and as a mediator of inflammation. However, the biology and dynamism of the adult epicardium is poorly understood. To investigate this, we created a transgenic line to ablate the epicardial cell population in adult zebrafish. Here we find that genetic depletion of the epicardium after myocardial loss inhibits cardiomyocyte proliferation and delays muscle regeneration. The epicardium vigorously regenerates after its ablation, through proliferation and migration of spared epicardial cells as a sheet to cover the exposed ventricular surface in a wave from the chamber base towards its apex. By reconstituting epicardial regeneration ex vivo, we show that extirpation of the bulbous arteriosus-a distinct, smooth-muscle-rich tissue structure that distributes outflow from the ventricle-prevents epicardial regeneration. Conversely, experimental repositioning of the bulbous arteriosus by tissue recombination initiates epicardial regeneration and can govern its direction. Hedgehog (Hh) ligand is expressed in the bulbous arteriosus, and treatment with a Hh signalling antagonist arrests epicardial regeneration and blunts the epicardial response to muscle injury. Transplantation of Sonic hedgehog (Shh)-soaked beads at the ventricular base stimulates epicardial regeneration after bulbous arteriosus removal, indicating that Hh signalling can substitute for the influence of the outflow tract. Thus, the ventricular epicardium has pronounced regenerative capacity, regulated by the neighbouring cardiac outflow tract and Hh signalling. These findings extend our understanding of tissue interactions during regeneration and have implications for mobilizing epicardial cells for therapeutic heart repair.

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Context-specific effects of outflow tract on epicardial regenerationa, (Top) Following ex vivo epicardial ablation and BA removal, ventricles were co-cultured with 10 outflow tracts in a transwell assay and observed for regeneration. (Bottom) No evidence for epicardial regeneration was observed in these experiments (n = 9; behavior seen in all samples). b, (Left) Following ex vivo epicardial ablation and BA removal, a non-transgenic BA (labeled as donor OFT) was transplanted to the apex and observed for regeneration. (Right) No evidence for regeneration of EGFP+ epicardium from apex to base was observed in these experiments (n = 10; behavior seen in all samples). Red dashed lines in (a, b), epicardium. White dashed lines in (a, b), ventricle. Yellow dashed lines in (b), donor outflow tract. Scale bars, 50 μm.
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Figure 10: Context-specific effects of outflow tract on epicardial regenerationa, (Top) Following ex vivo epicardial ablation and BA removal, ventricles were co-cultured with 10 outflow tracts in a transwell assay and observed for regeneration. (Bottom) No evidence for epicardial regeneration was observed in these experiments (n = 9; behavior seen in all samples). b, (Left) Following ex vivo epicardial ablation and BA removal, a non-transgenic BA (labeled as donor OFT) was transplanted to the apex and observed for regeneration. (Right) No evidence for regeneration of EGFP+ epicardium from apex to base was observed in these experiments (n = 10; behavior seen in all samples). Red dashed lines in (a, b), epicardium. White dashed lines in (a, b), ventricle. Yellow dashed lines in (b), donor outflow tract. Scale bars, 50 μm.

Mentions: To test if outflow tract tissue is sufficient to stimulate epicardial regeneration, we ectopically positioned experimentally manipulated cardiac structures. Co-culture of several BAs in a transwell assay with an epicardially ablated ventricle did not restore regeneration in the absence of host BA (Extended Data Fig. 6a). Similarly, a BA graft placed at the ventricular apex showed no evidence of directing regeneration of basally located host epicardial cells toward the apex (Extended Data Fig. 6b). Thus, we could not detect BA effects requiring long-range diffusion through tissue or culture medium. Next, we transplanted an tcf21:nucEGFP+ epicardial cell patch to the apex of an ablated host ventricle, after which we grafted a wild-type BA to the apex (Fig. 3b). Remarkably, the apical BA was capable of stimulating apex-to-base regeneration from the nearby epicardial patch in a high proportion (21 of 32) of experiments, effectively reversing the stereotypic direction of recovery (Fig. 3c, d). Together, these experiments indicate that the cardiac outflow tract is necessary and sufficient for epicardial regeneration, and that this neighboring tissue provides a short-range influence(s) that directs regeneration from base to apex.


Epicardial regeneration is guided by cardiac outflow tract and Hedgehog signalling.

Wang J, Cao J, Dickson AL, Poss KD - Nature (2015)

Context-specific effects of outflow tract on epicardial regenerationa, (Top) Following ex vivo epicardial ablation and BA removal, ventricles were co-cultured with 10 outflow tracts in a transwell assay and observed for regeneration. (Bottom) No evidence for epicardial regeneration was observed in these experiments (n = 9; behavior seen in all samples). b, (Left) Following ex vivo epicardial ablation and BA removal, a non-transgenic BA (labeled as donor OFT) was transplanted to the apex and observed for regeneration. (Right) No evidence for regeneration of EGFP+ epicardium from apex to base was observed in these experiments (n = 10; behavior seen in all samples). Red dashed lines in (a, b), epicardium. White dashed lines in (a, b), ventricle. Yellow dashed lines in (b), donor outflow tract. Scale bars, 50 μm.
© Copyright Policy - permission
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC4494087&req=5

Figure 10: Context-specific effects of outflow tract on epicardial regenerationa, (Top) Following ex vivo epicardial ablation and BA removal, ventricles were co-cultured with 10 outflow tracts in a transwell assay and observed for regeneration. (Bottom) No evidence for epicardial regeneration was observed in these experiments (n = 9; behavior seen in all samples). b, (Left) Following ex vivo epicardial ablation and BA removal, a non-transgenic BA (labeled as donor OFT) was transplanted to the apex and observed for regeneration. (Right) No evidence for regeneration of EGFP+ epicardium from apex to base was observed in these experiments (n = 10; behavior seen in all samples). Red dashed lines in (a, b), epicardium. White dashed lines in (a, b), ventricle. Yellow dashed lines in (b), donor outflow tract. Scale bars, 50 μm.
Mentions: To test if outflow tract tissue is sufficient to stimulate epicardial regeneration, we ectopically positioned experimentally manipulated cardiac structures. Co-culture of several BAs in a transwell assay with an epicardially ablated ventricle did not restore regeneration in the absence of host BA (Extended Data Fig. 6a). Similarly, a BA graft placed at the ventricular apex showed no evidence of directing regeneration of basally located host epicardial cells toward the apex (Extended Data Fig. 6b). Thus, we could not detect BA effects requiring long-range diffusion through tissue or culture medium. Next, we transplanted an tcf21:nucEGFP+ epicardial cell patch to the apex of an ablated host ventricle, after which we grafted a wild-type BA to the apex (Fig. 3b). Remarkably, the apical BA was capable of stimulating apex-to-base regeneration from the nearby epicardial patch in a high proportion (21 of 32) of experiments, effectively reversing the stereotypic direction of recovery (Fig. 3c, d). Together, these experiments indicate that the cardiac outflow tract is necessary and sufficient for epicardial regeneration, and that this neighboring tissue provides a short-range influence(s) that directs regeneration from base to apex.

Bottom Line: Transplantation of Sonic hedgehog (Shh)-soaked beads at the ventricular base stimulates epicardial regeneration after bulbous arteriosus removal, indicating that Hh signalling can substitute for the influence of the outflow tract.Thus, the ventricular epicardium has pronounced regenerative capacity, regulated by the neighbouring cardiac outflow tract and Hh signalling.These findings extend our understanding of tissue interactions during regeneration and have implications for mobilizing epicardial cells for therapeutic heart repair.

View Article: PubMed Central - PubMed

Affiliation: Department of Cell Biology and Howard Hughes Medical Institute, Duke University Medical Center, Durham, North Carolina 27710, USA.

ABSTRACT
In response to cardiac damage, a mesothelial tissue layer enveloping the heart called the epicardium is activated to proliferate and accumulate at the injury site. Recent studies have implicated the epicardium in multiple aspects of cardiac repair: as a source of paracrine signals for cardiomyocyte survival or proliferation; a supply of perivascular cells and possibly other cell types such as cardiomyocytes; and as a mediator of inflammation. However, the biology and dynamism of the adult epicardium is poorly understood. To investigate this, we created a transgenic line to ablate the epicardial cell population in adult zebrafish. Here we find that genetic depletion of the epicardium after myocardial loss inhibits cardiomyocyte proliferation and delays muscle regeneration. The epicardium vigorously regenerates after its ablation, through proliferation and migration of spared epicardial cells as a sheet to cover the exposed ventricular surface in a wave from the chamber base towards its apex. By reconstituting epicardial regeneration ex vivo, we show that extirpation of the bulbous arteriosus-a distinct, smooth-muscle-rich tissue structure that distributes outflow from the ventricle-prevents epicardial regeneration. Conversely, experimental repositioning of the bulbous arteriosus by tissue recombination initiates epicardial regeneration and can govern its direction. Hedgehog (Hh) ligand is expressed in the bulbous arteriosus, and treatment with a Hh signalling antagonist arrests epicardial regeneration and blunts the epicardial response to muscle injury. Transplantation of Sonic hedgehog (Shh)-soaked beads at the ventricular base stimulates epicardial regeneration after bulbous arteriosus removal, indicating that Hh signalling can substitute for the influence of the outflow tract. Thus, the ventricular epicardium has pronounced regenerative capacity, regulated by the neighbouring cardiac outflow tract and Hh signalling. These findings extend our understanding of tissue interactions during regeneration and have implications for mobilizing epicardial cells for therapeutic heart repair.

Show MeSH
Related in: MedlinePlus