Background:

Hypertrophic cardiomyopathy (HCM) is a genetic disease characterized by myocardial hypertrophy and progressive heart failure. Current therapeutic strategies are limited, ranging from symptomatic heart failure treatment to ventricular septal myomectomy. Therapeutic endeavors are currently limited by both the heterogeneity of disease severity, as well as the poorly-defined molecular underpinnings of HCM. Over 50% of HCM-associated mutations are found in the MYBPC3 gene, which encodes the sarcomere protein Myosin Binding Protein C3 (MyBPC3). We found that Calcium/calmodulin-dependent kinase II (CaMKII) - a known key regulator of ventricular remodelling and malignant arrhythmias - is overactive in the hearts of MyBPC3 knock-in mice (KI) that carry a human c.772G>A mutation. These findings are consistent with studies demonstrating an up-regulation of CaMKII in human ventriculoseptal HCM biopsies.

AIM: To unravel the role of CaMKII in the pathogenesis of HCM in vivo

Methods and Results:

Homozygous cardiomyocyte-specific CaMKIId/CaMKIIg double knockout (CaMKII DKO) mice were crossed with homozygous MyBPC3 KI mice. Early prenatal genetic deletion of CaMKII via Cre-recombinase driven by the αMHC promoter reduced the survival of CaMKII DKO/MYBPC3 KI mice dramatically to only 28 weeks compared with approximately 105 weeks in MyBPC3 KI mice. Echocardiographic measurements revealed a massive reduction in cardiac ejection fraction (EF) in CaMKII DKO/MYBPC3 KI mice (30.93 % vs. 67.12% in MyBPC3 KI) mice already at postnatal day (PND) 5. In contrast, cardiomyocyte-specific induction of CaMKII DKO in adult MyBPC3 KI mice via the MerCreMer system had no worsening effect on survival and cardiac function, indicating that CaMKII plays an adaptive role solely in the early development of HCM. qPCR analysis from cardiac tissue revealed an upregulation of proliferation markers in MyBPC3 KI mice during the first 2 postnatal weeks compared with WT mice, i.e. an approximately 2-fold increase in the cell cycle Cyclin Dependent Kinase (CDK) encoding genes Ccna2, Ccnb1, CDK4, E2F2, Cdc25c. Moreover, immunostainings for the proliferation marker H3S10ph indicated a proliferation rate of 1,89% cardiomyocytes of MyBPC3 KI mice versus 0,57% in cardiomyocytes of WT mice at PND5, suggesting cardiomyocyte hyperplasia in the early phase of HCM. In CaMKII DKO/MYBPC3 KI mice up-regulation of these pro-proliferative markers was markedly diminished by 50% compared with MyBPC3 KI (in Ccna2, Ccnb1, Cdc20 and Cdc25c ), indicating a CaMKII-dependent activation of proliferation genes. To assess whether increased cardiomyocyte proliferation plays an adaptive or detrimental role in MyBPC3 KI mice, we administered the proliferation inhibitor Rapamycin (2 μg/g/d) in MyBPC3 mice and WT mice during the first postnatal week. Echocardiographic measurements revealed an 18% decrease in EF on PND 7 in MyBPC3 KI mice injected with Rapamycin compared with vehicle-treated KI mice, indicating that cardiomyocyte proliferation is an adaptive feature during the early development of HCM in vivo. 

Conclusion:

Our in vivo data from genetic mouse models suggest that CaMKII plays an adaptive role in the setting of HCM. Mechanistically, CaMKII appears to control postnatal cardiomyocyte proliferation, which is likely a relevant adaptive feature in the early development of HCM. We therefore present a previously unknown role of CaMKII as a cardioprotective mediator of postnatal proliferation.