Abstract Body: Background:
In the mammalian heart, most CM are polyploid, having >2N DNA content. Using the Hybrid Rat Diversity Panel (HRDP), the Patterson Lab recently showed, hyperpolyploid CM (≥8N) positively correlates with LV dilation and negatively with ejection fraction (EF), suggesting CM ploidy may influence cell dimension and contractility. In the present study, We characterize cellular relationships and underlying mechanisms.
Methods:
Using HRDP, data was collected from 13 rat strains. CMs (3–4 weeks old) were isolated via Langendorff digestion, fixed, and immunostained (α-actinin/DAPI). NIS Elements quantified ploidy and dimensions, while IonOptix (1Hz, 20V) measured contractility. Finally, 3-week-old rats received Mavacamten (1.25-2.5 mg/kg/day, 9 days) followed by echocardiography and ploidy analysis.
Results:
Rat strains exhibit a range of hyperpolyploid CM (≥8N) spanning from 0.8-20.7%. Dimension (area, length & width) of each CM, considering ploidy (e.g. 2N, 4N, 8N) unveiled that all strains showed a significant stepwise increase in area across ploidy classes (p=1.0e-16), suggesting a relationship between ploidy and cell size. Length vs width analysis further established eccentric growth with genome doublings. Finally, while holding ploidy constant, some strains displayed larger CMs, suggesting genetics independently influence cell size (p=1.11e-18).
As per observation, EF declined as ploidy increases, next assessed if contractility differences were observed at the cellular level. Mononuclear CMs showed increased contractility (Δ sarcomere length: 0.05±0.001 µm) than the binucleated (0.115±0.003 µm; p=0.0001). To directly assess whether contractility modulates polyploidy, SHR rats were treated with 2.5 mg/kg Mavacamten, significantly reduced EF by 25% and increased the ≥8N CMs (25.12±0.91% vs. 12.33±2.91% vehi; p=0.004). A lower dose (1.25 mg/kg) produced a 15% EF drop with an increase in ≥8N CMs (17.5±0.65% vs. 12.33±1.20% veh; p=0.021). In contrast, identical dosing in M520 caused a marked 60–70% EF reduction without altering CM ploidy. Mechanistically, Mava-treated CMs exhibited significant reductions in P21 and Lamin B2 expression, suggesting involvement of cell-cycle regulation and nuclear envelope remodeling.
Conclusion:
CM ploidy correlates with contractility and CM eccentric growth, though size is partly genetically independent of ploidy. Pharmacological reduction in contractility suggests a direct effect on ploidy, thereby moving past correlation.