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A systematic analysis of neonatal mouse heart regeneration after apical resection

https://doi.org/10.1016/j.yjmcc.2014.12.011Get rights and content

Highlights

  • Regeneration occurs after apical resection in neonatal mice.

  • The extensiveness of scarring after apical resection is related to resection size.

  • Apical resection results in increased cardiomyocyte cell cycle activity.

  • Surgical retraction of the ventricle increases cardiomyocyte cell cycle activity.

Abstract

The finding that neonatal mice are able to regenerate myocardium after apical resection has recently been questioned. We determined if heart regeneration is influenced by the size of cardiac resection and whether surgical retraction of the ventricular apex results in an increase in cardiomyocyte cell cycle activity. We performed moderate or large apical ventricular resections on neonatal mice and quantified scar infiltration into the left ventricular wall at 21 days post-surgery. Moderately resected hearts had 15 ± 2% of the wall infiltrated by a collagen scar; significantly greater scar infiltration (23 ± 4%) was observed in hearts with large resections. Resected hearts had higher levels of cardiomyocyte cell cycle activity relative to sham hearts. Surgically retracting the ventricle often resulted in fibrosis and induced cardiomyocyte cell cycle activity that were comparable to that of resected hearts. We conclude that apical resection in neonatal mice induces cardiomyocyte cell cycle activity and neomyogenesis, although scarring can occur. Surgical technique and definition of approach to assessing the extent of regeneration are both critical when using the neonatal mouse apical resection model.

Introduction

The discovery by Porrello et al. in 2011 that neonatal mice have the potential to regenerate resected myocardium generated much excitement in cardiovascular biology [1]. A recent study by Andersen et al. questioned the utility of the apical resection model [2]. Andersen et al. found that extensive scarring occurred in apically resected hearts at 21 days post-surgery and found limited evidence for neomyogenesis [2]. Understandably, the conflicting results produced by this study has generated some confusion and controversy as several laboratories have produced data that the neonatal mouse heart does have regenerative capabilities and is able to undergo neomyogenesis after sustaining myocardial injury [3]. Much speculation has occurred as to why the Porrello and Anderson studies appear to conflict, and it is possible that technical considerations as well as determining how regeneration is defined are important [3], [4], [5].

We systematically examined how technical considerations influence this important experimental model. These considerations included the size of apical resection, which was posited to be one cause for the conflicting findings, as well as the mechanical fixation (surgical retraction) of the ventricle during surgery [3]. We aimed to systematically and quantitatively clarify the extent of regeneration, neomyogenesis, and scarring that occur in this model. We show that new myocardial formation clearly occurs after apical resection, but that this regeneration is often accompanied by some scarring at 21 days post-resection (dpr). The extent of scarring is related to resection size. We also show that there is an increase in cardiomyocyte cell cycle activity, but fibrosis and an increase in cardiomyocyte cell cycle activity occur when sham operations include surgical retraction; the surgical retraction effect in control hearts can mask the induction of cardiomyocyte cell cycle activity in resected hearts. These data clarify the neonatal mouse apical resection model.

Section snippets

Surgical procedures

Neonatal mice were anesthetized for 4 min on ice. We performed thoracotomy followed by resection of either 10 or 20% of the ventricle. Our typical sham operation did not involve mechanically fixing the apex of the left ventricle. For non-retracted sham operations, we performed thoracotomy without resection. For surgical retraction experiments, we gently fixed the left ventricle with a microneedle holder after open thoracotomy.

Statistical analysis

Data are presented as mean ± SEM. A one-way ANOVA followed by a pairwise

Scarring often accompanies new heart muscle formation after apical resection and is related to resection size

To study the impact of resection size, we measured the relative size of the resected myocardium (Supplemental Fig. 1A). As intended, large resections were approximately twice as large as moderate resections (Supplemental Fig. 1B). After quantifying the weights of hearts at 3 h post-surgery, we found that moderate and large resections removed about 10% and 20% of the ventricular myocardium respectively (Fig. 1A). Thus, we were able to quantitatively distinguish that resection size, and the sizes

Funding sources

This work was supported by an HHMI Gilliam Fellowship (D.M.B.), an NIH National Research Service Award (C.C.O.), and grants from the NIH (AG040019 and HL117986 to R.T.L.).

Disclosure statement

None.

References (5)

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