A conserved HH-Gli1-Mycn network regulates heart regeneration from newt to human

Bhairab N Singh,Cyprian V Weaver,Naoko Koyano-Nakagawa,Satyabrata Das,Mary G Garry,Jop H Van Berlo,Wuming Gong,Elizabeth Braunlin,Ivan P Moskowitz,Daniel J Garry

doi:10.1038/s41467-018-06617-z

Bhairab N Singh, Cyprian V Weaver + Show 8 more

Open Access

https://doi.org/10.1038/s41467-018-06617-z

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Abstract

The mammalian heart has a limited regenerative capacity and typically progresses to heart failure following injury. Here, we defined a hedgehog (HH)-Gli1-Mycn network for cardiomyocyte proliferation and heart regeneration from amphibians to mammals. Using a genome-wide screen, we verified that HH signaling was essential for heart regeneration in the injured newt. Next, pharmacological and genetic loss- and gain-of-function of HH signaling demonstrated the essential requirement for HH signaling in the neonatal, adolescent, and adult mouse heart regeneration, and in the proliferation of hiPSC-derived cardiomyocytes. Fate-mapping and molecular biological studies revealed that HH signaling, via a HH-Gli1-Mycn network, contributed to heart regeneration by inducing proliferation of pre-existing cardiomyocytes and not by de novo cardiomyogenesis. Further, Mycn mRNA transfection experiments recapitulated the effects of HH signaling and promoted adult cardiomyocyte proliferation. These studies defined an evolutionarily conserved function of HH signaling that may serve as a platform for human regenerative therapies.

Highlights

The mammalian heart has a limited regenerative capacity and typically progresses to heart failure following injury
Having described the role of HH signaling in the regulation of cardiomyocyte proliferation and regeneration in both newt and mouse (Figs. 1–7), we examined the conserved function of HH signaling using human-induced pluripotent stem cell-derived cardiomyocytes
Ccnd[2], Ccne[1], and Cdkn1b (p27) following lentiviral Gli[1] overexpression or knockdown in the postnatal day 1 (P1) cardiomyocytes. g qPCR analysis of Mycn transcripts using RNA isolated from P1 and P28 wild-type heart tissue (n = 3). h qPCR analysis of Mycn transcripts using RNA isolated from control, SAG, and CyA treated isolated neonatal cardiomyocytes (n = 3 replicates from each group). i Schematic of Mycn overexpression and knockdown experiments in the P1 cardiomyocytes. j, k Immunostaining (j) and quantification of α-Actinin+-EdU+ cells (k) from control, Mycn, and shMycn lentiviral infected neonatal cardiomyocytes

Summary

Introduction

The mammalian heart has a limited regenerative capacity and typically progresses to heart failure following injury. Mycn mRNA transfection experiments recapitulated the effects of HH signaling and promoted adult cardiomyocyte proliferation These studies defined an evolutionarily conserved function of HH signaling that may serve as a platform for human regenerative therapies. Lower vertebrates such as the adult newt and zebrafish can achieve complete heart regeneration following injury by activating developmental regulatory networks[1,2,3,4,5] In these organisms, adult cardiomyocytes undergo dedifferentiation to re-enter the cell cycle and, differentiation to facilitate tissue regeneration[6,7]. Efforts have focused on the role of pathways and factors that promote cardiomyocyte proliferation and tissue regeneration in the adult mammalian heart that can prevent the progression of heart failure and premature death following cardiac injury. We define a novel HH-Gli1-Mycn gene regulatory network that regulates cardiomyocyte proliferation and promotes heart regeneration

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