Journal
METHODS
Volume 190, Issue -, Pages 55-62Publisher
ACADEMIC PRESS INC ELSEVIER SCIENCE
DOI: 10.1016/j.ymeth.2020.06.002
Keywords
Stereology; Cardiomyocyte proliferation; Cell cycle; Binucleation; Cardiomyocyte number; Cardiac regeneration
Funding
- European Regional Development Fund (ERDF) through COMPETE 2020, Portugal 2020
- FCT (Fundacao para a Ciencia e Tecnologia) [POCI-01-0145-FEDER-030985]
- FCT/Ministerio da Ciencia, Tecnologia e Inovacao [SFRH/BD/111799/2015, PD/BD/127997/2016, SFRH/BD/144769/2019, CEECINST/00091/2018]
- i3S Scientific Platform Advanced Light Microscopy [POCI-01-0145-FEDER-022122]
- Dutch Heart Foundation [NHS2015T066]
- Fundação para a Ciência e a Tecnologia [SFRH/BD/111799/2015, PD/BD/127997/2016, SFRH/BD/144769/2019, CEECINST/00091/2018] Funding Source: FCT
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Cardiovascular diseases continue to be the leading cause of death, with limited regenerative capacity of the adult mammalian heart. Neonatal mammals have demonstrated the ability to regenerate heart muscle. An optimized protocol has been proposed to assess cardiomyocyte proliferation and quantity, aiming to improve cardiac response to injury.
Cardiovascular diseases remain the leading cause of death, largely due to the limited regenerative capacity of the adult mammalian heart. Yet, neonatal mammals were shown to regenerate the myocardium after injury by increasing the proliferation of pre-existing cardiomyocytes. Re-activation of cardiomyocyte proliferation in adulthood has been considered a promising strategy to improve cardiac response to injury. Notwithstanding, quantification of cardiomyocyte proliferation, which occurs at a very low rate, is hampered by inefficient or unreliable techniques. Herein, we propose an optimized protocol to unequivocally assess cardiomyocyte proliferation and/or cardiomyocyte number in the myocardium. Resorting to a stereological approach we estimate the number of cardiomyocytes using representative thick sections of left ventricle fragments. This protocol overcomes the need for spatial-temporal capture of cardiomyocyte proliferation events by focusing instead on the quantification of the outcome of this process. In addition, assessment of cardiomyocyte nucleation avoids overestimation of cardiomyocyte proliferation due to increased binucleation. By applying this protocol, we were able to previously show that apical resection triggers proliferation of preexisting cardiomyocytes generating hearts with more cardiomyocytes. Likewise, the protocol will be useful for any study aiming at evaluating the impact of neomyogenic therapies.
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