Cardiomyocytes are polyploid, but how this polyploidy is established during development is unclear. In a new paper in Development, Samantha Swift and colleagues reveal variation in cardiomyocyte polyploidy between mouse strains and identify candidates that regulate this phenotype. We caught up with first author, Samantha Swift, and corresponding author, Michaela Patterson (Assistant Professor at the Medical College of Wisconsin, USA), to learn more about their research.

Samantha Swift (L) and Michaela Patterson (R)

Michaela, can you give us your scientific biography and the questions your lab is trying to answer?

MP: During my undergraduate days, I was exposed to the field of developmental biology and tissue regeneration, and I have truly never looked back. I'm captivated by the observation that a single cell, the fertilized egg, can divide and differentiate and morph through an incredibly intricate process to form a fully functioning organism. But, more exciting – can we leverage our knowledge of development to stimulate the process for tissue regeneration purposes? Since my undergraduate days, my specific focus has jumped around a bit – as a research technician, I was involved in cell transplantation studies of pluripotent stem cell derivatives for neurodegenerative disorders; as a graduate student, I took a step back to better understand the differentiation process and ask whether in vitro pluripotent stem cell differentiation modelled in vivo human development; as a postdoc, and now as an independent investigator, I have moved into the realm of endogenous regeneration – but my overarching interests have remained.

Samantha, how did you come to work in Michaela's lab and what drives your research today?

SS: I was drawn to the Medical College of Wisconsin for my graduate training largely because of the incredibly collaborative nature of the research groups here. I could tell instantly that this was an institution of researchers that worked with and supported each other. I completed my very last rotation of four in Michaela's lab at the suggestion of a friend. Her lab was brand new at the institution and my research interests were largely neurobiology based, so the rotation was a bit of a shot in the dark. It couldn't have been more than a handful of days before I knew that this was the person I wanted to train me as a scientist. Her research was fascinating, and from the very first conversation she valued my input – considered me as a whole person, not just a student – and brought me into important conversations that shaped the way I approach scientific questions. I always encourage students to choose the lab in which they will receive their PhD by prioritizing compatible mentorship styles. One can always adjust/tweak their field of study, but a healthy lab environment is everything when you're learning to be a scientist. My research interests now are largely driven by a desire to understand the genetic underpinnings of cell biology. I think using disease models (such as heart disease) to understand these genetic components is absolutely fascinating.

What was known about cardiomyocyte ploidy variation before your work?

MP: Very little. We knew that inbred mouse strains have diverse displays of cardiomyocyte ploidy in adulthood and that polyploidization in the mammalian heart is predominantly a postnatal phenomenon. That's about it. We're really just scratching the surface here – there is so much more we don't understand.

Can you give us the key results of the paper in a paragraph?

SS: In this paper, we both expand upon previous findings and make entirely novel discoveries about the dynamics of ploidy in distinct inbred mouse strains. We found that C57Bl/6J mice follow the accepted and understood developmental time course in terms of cardiomyocyte expansion, cell cycle activity and polyploidization. Conversely, we found that A/J mice deviate from this known and understood progression to cardiomyocyte polyploidization and show evidence for cell division at a time far later in development than would have been expected previously. We go on to show that Tnni3k, which was already known to influence the end-state ploidy classes in adult A/J mice, also plays a role in the developmental phenotypes we characterized. Additionally, we pulled out and validated one more candidate gene, Runx1, as regulating these ploidy and cell cycle phenotypes.

Were you surprised that Tnni3k knockout was insufficient to recapitulate the phenotype in A/J mice?

MP: Not at all, in fact, and for a couple reasons. We knew from the original assessment of cardiomyocyte ploidy in adult hearts across inbred strains that the distribution of the phenotype was continuous, which in genetics suggests a complex trait, or multiple genes are at play. We also knew that knocking out Tnni3k on its own in a C57BL/6J mouse only raised the mononuclear diploid cardiomyocyte (MNDCM) numbers from ∼2% to ∼5%. Meanwhile, in A/J mice MNDCM numbers are up around 9-10%, so this one gene was only responsible for about 35% of the complete phenotypic difference between A/J and C57BL/6. We fully anticipated that if Tnni3k was also influencing this unique dynamic ploidy event we uncovered here in this story, it would likely only be able to partially recapitulate the process.

How do you propose Runx1 regulates ploidy in the cardiomyocytes of A/J mice?

SS: We really don't know yet. If I had to give a broad hypothesis based on the literature and knowing its function as a transcription factor, I imagine it could activate the expression of genes relevant to the ‘dedifferentiation’ of cardiomyocytes allowing them to both re-enter – and complete – the cell cycle, while also repressing expression of maturation factors. This would be an exciting and relevant question to investigate in the future, which I hope the lab pursues. It could have profound effects on the distribution of ploidy classes.

Variation in cardiomyocyte polyploidy. Single-cell ventricular suspension stained for cardiac troponin T (cTnT) (magenta) and DAPI (cyan). Identified cardiomyocyte ploidy classes are labelled on the image.

Variation in cardiomyocyte polyploidy. Single-cell ventricular suspension stained for cardiac troponin T (cTnT) (magenta) and DAPI (cyan). Identified cardiomyocyte ploidy classes are labelled on the image.

What implications will your study have for the importance of considering mouse strain when studying heart development and regeneration?

MP: I hope people will start to pay more attention to genetic backgrounds and appreciate that the conclusions one might draw from a single study are not necessarily blanket conclusions, but instead incredibly context dependent.

SS: My grand hope is that, as we acquire more information on how genetic backgrounds influence phenotypes like those discussed here in mice, eventually we will be able to extrapolate some of this understanding to diverse outcomes in human populations and optimize treatment plans based on a person's unique genetic make-up.

When doing the research, did you have any particular result or eureka moment that has stuck with you?

SS: I would say, with this paper, every single piece of data felt like a tiny eureka moment. I tend to approach science and results with a fairly large amount of scepticism. The initial discovery of dynamic ploidy states in A/J mice felt like such an unbelievable result that every single time our team got another result that confirmed what we were seeing was actually happening the ‘eureka’ moment was building.

With this paper, every single piece of data felt like a tiny eureka moment

And what about the flipside: any moments of frustration or despair?

SS: There were equally as many of these as there were tiny victories and eureka moments. Given this research makes up the bulk of my thesis work, every single study and experiment was a learning experience for me, especially being the first student in a new lab. Every single test we wanted to run or experiment we wanted to do required troubleshooting and optimization, and came with a fairly large amount of failure for a PhD student transitioning from a neuroscience background to a cardiac field. The support I received from my lab, mentor, and collaborating labs helped to make these moments transient so I could get back to the bench and try that tricky stain just one more time, until we had beautifully functioning protocols. Also, A/Js are TERRIBLE breeders.

Samantha, what is next for you after this paper?

SS: I have a few more techniques I'd like to learn and a couple more short stories to tell before I wrap up my PhD and start searching for post-doctoral positions! But, very soon I will be defending my thesis, leaving the ‘nest’ (Medical College of Wisconsin) and taking the next steps in my career.

Michaela, where will this story take your lab next?

MP: We continue to employ forward and reverse genetics techniques to explore additional genes and loci that contribute to cardiomyocyte ploidy. We're seeing some really dramatic and unexpected phenotypes with these new candidates and, more exciting to me, we're seeing some really cool effects when we combine genes together. From a grander perspective, we as a field still don't understand why mammalian cardiomyocytes become polyploid in the first place. All the reasons that have been offered by the field are still just supposition. I hope in the coming years that my lab can help contribute to our understanding of how polyploidization influences heart physiology and function.

Finally, let's move outside the lab – what do you like to do in your spare time?

SS: I love to get outside! Milwaukee is a great place to be, but especially so in the summer, which is almost upon us! I'll either be exploring the woods near the city or hanging out in a beer garden with my close friends – that is, when I'm not in lab of course!

MP: I'm a mom to two young kids. Together with my scientist husband, we love to be outside – hikes, beach trips, sledding in the winter, playing in the yard, even just a walk around the neighbourhood. I'm also quite passionate about a new programme I'm building across the state of Wisconsin that aims to partner our phenomenal primarily undergraduate institutions with the Medical College of Wisconsin. In the long run, we hope it will expose more undergraduates to scientific principles, cutting-edge science and ultimately inspire them to pursue careers in research. Time will tell, but for now it certainly inspires me daily.

Medical College of Wisconsin, Milwaukee, WI 53226, USA.

E-mail: [email protected]

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Cardiomyocyte ploidy is dynamic during postnatal development and varies across genetic backgrounds