REPLYING TO C. Vicinanza et al. Nature 555 https://doi.org/10.1038/nature25771 (2018)
The accompanying Comment by Vicinanza et al.1 does not agree with our previous study2 regarding the utility of c-Kitpos (also known as Kitpos) cells when investigating regeneration in the heart. The authors claim that three previous publications2,3,4 using Kitcre knock-in lineage-tracing question their previous work that shows that tissue-specific c-Kitpos cardiac stem/progenitor cells (CSCs) are necessary and sufficient for cardiomyocyte regeneration and/or replenishment after injury5. However, the regenerative data within their previous manuscript did not address the role of endogenous c-Kitpos cells in the heart5. Instead, they relied on clonally derived c-Kitpos cells that were selected in culture and then injected into the circulatory system of mice and these cells remarkably homed to the heart and generated abundant new cardiomyocytes that repaired isoproterenol-mediated injury5.
The authors now claim that the number of c-Kitpos cells with cardiomyogenic-clone-producing potential is very low in the mouse heart (approximately 1% of total c-Kitpos cells)6. Indeed, the authors showed that broadly isolated primary cardiac c-Kitpos cells, that were not clonally selected, had almost no ability to generate new cardiomyocytes or repair the infarcted heart6. The authors’ new interpretation1 that only a very small portion of c-Kitlow cells can be made clonal with regenerative activity contradicts their published results from 2003 in which broadly isolated c-Kitpos cells regenerated 70% of the myocardium after infarction injury7, or published results from 2013 showing that the majority of endogenous c-Kitpos cells express Nkx2.5 (a cardiomyocyte-determining transcription factor)5.
The authors further claim that Kitcre knock-in lineage tracing approaches2,3,4 fail to track these rare c-Kitlow CSCs, because of poor recombination efficiency and heterozygosity in the Kit allele. While we have previously addressed this criticism and discussed data to the contrary8, we have recently published additional data showing that the majority of c-KitposCD45neg cells isolated from the hearts of Kitcre mice are correctly lineage-traced9, suggesting that this technique for tracking cells in mice is effective2,3,4. Indeed, a working group of established cardiac scientists recently published a consensus statement confirming that endogenous cardiac c-Kitpos cells are an unlikely source for meaningful heart regeneration10. This statement was also based on the known biology of the adult mammalian heart, which essentially lacks acute cardiomyocyte-forming regenerative capacity. However, it seems reasonable that one could select and isolate rare clonally derived c-Kitpos progenitor cell lines with ectopic cardiomyogenic activity, as suggested in the Comment1. However, the Comment does not address the role of endogenous c-Kitpos cells in the heart. The current negative results simply address whether clonally derived c-Kit-expressing cells can have cardiomyogenic capacity, which we believe lacks all in vivo relevance and in no way contests our previous observations.
Vicinanza, C. et al. Kitcre knock-ins fail to fate-map cardiac stem cells. Nature 555, https://doi.org/10.1038/nature25771 (2018)
van Berlo, J. H. et al. c-kit+ cells minimally contribute cardiomyocytes to the heart. Nature 509, 337–341 (2014)
Sultana, N. et al. Resident c-kit+ cells in the heart are not cardiac stem cells. Nat. Commun. 6, 8701 (2015)
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Ellison, G. M. et al. Adult c-kitpos cardiac stem cells are necessary and sufficient for functional cardiac regeneration and repair. Cell 154, 827–842 (2013)
Vicinanza, C. et al. Adult cardiac stem cells are multipotent and robustly myogenic: c-kit expression is necessary but not sufficient for their identification. Cell Death Differ. 24, 2101–2116 (2017)
Beltrami, A. P. et al. Adult cardiac stem cells are multipotent and support myocardial regeneration. Cell 114, 763–776 (2003)
Molkentin, J. D. Letter by Molkentin regarding article, “The absence of evidence is not evidence of absence: the pitfalls of Cre knock-ins in the c-Kit locus”. Circ. Res. 115, e21–e23 (2014)
Kanisicak, O., Vagnozzi, R. J. & Molkentin, J. D. Identity crisis for regenerative cardiac cKit+ cells. Circ. Res. 121, 1130–1132 (2017)
Eschenhagen, T. et al. Cardiomyocyte regeneration: a consensus statement. Circulation 136, 680–686 (2017)
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van Berlo, J., Kanisicak, O., Maillet, M. et al. van Berlo et al. reply. Nature 555, E18 (2018). https://doi.org/10.1038/nature25772
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