Diastolic contributors in cardiomyocytes of a cardiometabolic HFpEF-like mouse model.

Three main contributors to cardiomyocyte diastolic stiffness are (1) passive sarcomere stiffness, (2) microtubules, and (3) diastolic crossbridges (XBs), the XBs that are present in the diastolic phase. However, the relative contributions of these key determinants in heart failure with preserved ejection fraction (HFpEF) conditions are unclear. We quantify the relative contributions of passive sarcomere stress, the microtubule network, and diastolic XB activity to overall diastolic stress in intact cardiomyocytes isolated from two-hit mice, a cardiometabolic HFpEF-like model, in both sexes. The stretch-release protocol was used to obtain the diastolic stress-sarcomere length relation. XB inhibitor and colchicine treatment were used to determine the contributions of diastolic XBs and microtubules, respectively. Passive sarcomere stress was measured in cells treated with both colchicine and the XB inhibitor. Male HFpEF-like cardiomyocytes exhibit increases in both passive sarcomere stress (by 70%) and diastolic XBs (by 52%), whereas female HFpEF-like cardiomyocytes show an increase in passive sarcomere stress alone (by 55%). The microtubule network contributes to diastolic stress by augmenting the extent of diastolic XBs in males. The elevated diastolic XBs in male HFpEF-like mice are accompanied by altered Ca2+ transient, suggesting that remodeled Ca2+ handling accounts, in part, for the enhanced diastolic XB activity in males. These findings imply that, in males, both increased passive sarcomere stiffness and diastolic XB activity represent potential therapeutic targets for reducing cardiomyocyte diastolic stiffness, whereas in females, diastolic stiffness is predominantly driven by elevated passive sarcomere stiffness.