Modified mRNA directs the fate of heart progenitor cells and induces vascular regeneration after myocardial infarction

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Abstract

In a cell-free approach to regenerative therapeutics, transient application of paracrine factors in vivo could be used to alter the behavior and fate of progenitor cells to achieve sustained clinical benefits. Here we show that intramyocardial injection of synthetic modified RNA (modRNA) encoding human vascular endothelial growth factor-A (VEGF-A) results in the expansion and directed differentiation of endogenous heart progenitors in a mouse myocardial infarction model. VEGF-A modRNA markedly improved heart function and enhanced long-term survival of recipients. This improvement was in part due to mobilization of epicardial progenitor cells and redirection of their differentiation toward cardiovascular cell types. Direct in vivo comparison with DNA vectors and temporal control with VEGF inhibitors revealed the greatly increased efficacy of pulse-like delivery of VEGF-A. Our results suggest that modRNA is a versatile approach for expressing paracrine factors as cell fate switches to control progenitor cell fate and thereby enhance long-term organ repair.

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Figure 1: Highly efficient, transient gene transfer in vivo using modRNA.
Figure 2: VEGF-A modRNA enhanced formation of functional, nonleaky vessels.
Figure 3: VEGF-A modRNA improved outcome in a mouse myocardial infarction model.
Figure 4: VEGF-A modRNA reduced scar area and apoptosis and increased capillary density and WT1+ cells proliferation after myocardial infarction in a KDR-dependent manner.
Figure 5: VEGF-A modRNA induced WT1+ epicardial progenitor proliferation and shifted differentiation toward the endothelial lineage.
Figure 6: VEGF-A modRNA promoted differentiation of EPDCs toward the cardiovascular lineage in vivo.
Figure 7: Suggested model for the role of VEGF-A modRNA on EPDCs differentiation in vivo.

Change history

  • 19 September 2013

    In the version of this article initially published online, W.T.P. was not included in the correspondence line. Three sentences have also been added to Author Contributions. The omissions have been corrected for the print, PDF and HTML versions of this article.

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Acknowledgements

This work was funded by US National Institutes of Health U01H100408 (K.R.C.), U01HL098166 (K.R.C.), U01JL100401 (W.T.P.), R01HL094683 (W.T.P.), RC1HL099618 (K.R.C., W.T.P.) and UO1HL100402 (A.J.W.). K.O.L. held a Croucher Foundation Fellowship and A.J.W. is an Early Career Scientist of the Howard Hughes Medical Institute. We thank R. Liao, J. Guan, J. Truelove, L. Bu, M. Stachel, K. Buac, V. Priestly, R. Gazit, K. Ketman, N. Barteneva, A. He, S. Stevens, B. Zhou and L.Warren for all their help in this project. Adult cardiomyocytes were a kind gift from R. Liao (Biological and Biomaterial Science, Department of Medicine, Brigham and Women's Hospital, Harvard Medical School).

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Authors

Contributions

L.Z. (lzangi@enders.tch.harvard.edu) worked in the Rossi, Chien and Pu laboratories, and designed and carried out most of the experiments, analyzed most of the data, and wrote the manuscript. K.O.L. in the Chien lab designed and performed experiments and analyzed the qRT-PCR and immunostaining data, and wrote the manuscript. Her contribution is similar in significance to the contributions of L.Z. A.v.G. performed and analyzed the Wt1-related experiments. Q.M. and R.G. carried out myocardial infarction experiments. W.E. carried out plasmid preparation. L.M.P. performed blinded analysis of imaging data and wrote the manuscript. D.S. performed and analyzed skeletal muscle in vivo transfection. H.X. performed isolation of neonatal mouse cardiomyocytes. M.T. performed and analyzed in vitro transfection of mouse adult myotubes. B.S. carried out and analyzed the MRI experiment. M.N., D.M.B., R.A.L. and A.J.W. designed experiments, analyzed data, and revised the manuscript. D.J.R. (Derrick.Rossi@childrens.harvard.edu) designed in vitro cardiomyocyte experiments and revised the manuscript. K.R.C. (kchien@harvard.edu; kenneth.chien@ki.se) conceived the initial project and experimental studies, and with W.T.P. (wpu@enders.tch.harvard.edu) designed further experiments, analyzed data, and wrote the manuscript. The initial discovery of VEGF-A as a cell fate switch for heart progenitors in general, and its effects in myocardial infarction on the expansion of epicardial heart progenitors along with vascular regeneration was made in the Chien lab (L.Z., K.O.L. and K.R.C.). The Chien lab (L.Z., K.R.C.) and the Pu lab (A.v.G., W.T.P.) worked together to extend and expand these initial results. Address correspondence on ModRNA reagents, delivery and protocols to K.R.C. and on mouse models to W.T.P.

Corresponding authors

Correspondence to William T Pu or Kenneth R Chien.

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Competing interests

K.R.C. and D.J.R. are co-founders of Moderna Therapeutics, a Cambridge, Massachusetts company that is developing therapeutics based on modified mRNA. K.R.C. is an advisor to AstraZeneca, which has an interest in cardiovascular therapeutic applications of modRNA.

Supplementary information

Supplementary Text and Figures

Supplemental Appendix, Supplementary Tables 1-6, and Supplementary Figures 1-12 (PDF 24781 kb)

41587_2013_BFnbt2682_MOESM6_ESM.mov

Cine MRI of sham-operated heart, 21 days after operation (MOV 915 kb)

Supplementary video 1

Cine MRI of sham-operated heart, 21 days after operation (MOV 915 kb)

41587_2013_BFnbt2682_MOESM7_ESM.mov

Cine MRI of MI heart, vehicle treated heart, 21 days after operation (MOV 836 kb)

Supplementary video 2

Cine MRI of MI heart, vehicle treated heart, 21 days after operation (MOV 836 kb)

41587_2013_BFnbt2682_MOESM8_ESM.mov

Cine MRI of MI heart, VEGF-A modRNA treated, 21 days after operation (MOV 871 kb)

Supplementary video 3

Cine MRI of MI heart, VEGF-A modRNA treated, 21 days after operation (MOV 871 kb)

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Zangi, L., Lui, K., von Gise, A. et al. Modified mRNA directs the fate of heart progenitor cells and induces vascular regeneration after myocardial infarction. Nat Biotechnol 31, 898–907 (2013). https://doi.org/10.1038/nbt.2682

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