Inherited Cardiomyopathies: From Genotype to Phenotype

The heart undergoes extensive morphological, metabolic, and energetic remodeling in response to inherited, or familial, hypertrophic cardiomyopathies (FHC). Myocyte contractile perturbations downstream of Ca2+, the so-called sarcomere-controlled mechanisms, may represent the earliest indicators of this remodeling. We can now state that the dynamics of cardiac contraction and relaxation during the progression of FHC are governed by downstream mechanisms, particularly the kinetics and energetics of actin and myosin interaction to drive the trajectory of pathological cardiac remodeling. This notion is unambiguously supported by elegant studies above linking inheritable FHC-causing mutations to cardiomyopathies, known to disturb contractile function and alter the energy landscape of the heart. Although studies examining the biophysical properties of cardiac myocytes with FHC-causing mutations have yielded a cellular and molecular understanding of myofilament function, this knowledge has had limited translational success. This is driven by a critical failure in elucidating an integrated and sequential link among the changing energy landscape, myofilament function, and initiated signaling pathways in response to FHC. Similarly, there continues to be a major gap in understanding the cellular and molecular mechanisms contributing to sex differences in FHC development and progression. The primary reason for this gap is a lack of a “unifying” or “central” hypothesis that integrates signaling cascades, energetics, sex and FHC

Estrogen Dependent Regulation of the Amp-Activated Protein Kinase Pathway

Sex differences exist in the progression of heart disease, as premenopausal women are protected from developing severe hypertension, aortic stenosis, myocardial infarction and hypertrophic cardiomyopathies. The susceptibility and progression of cardiovascular disease increases in post-menopausal women. This is at least partially underlined by a pronounced decrease in circulating estrogen levels. Estradiol (E2), the most abundant estrogen in premenopausal women, is known to be cardioprotective. Recently, AMP-activated protein kinase (AMPK) has emerged as a prominent player in the development of cardiac hypertrophy and heart failure. AMPK is central to the energetic metabolism of the cell and is activated in response to energy deprivation. E2 has been shown to activate AMPK, by yet an unknown mechanism. The first part of this dissertation focuses on describing the molecular mechanism behind this AMPK activation. We found that E2 activates AMPK through a non- genomic pathway and involves direct interaction of classical estrogen receptors (ERα and ERβ) with the α-catalytic subunit of AMPK. These receptors also associate with the upstream kinase LKB1, which is required for E2-dependent activation of AMPK. Furthermore, the two estrogen receptors play opposite roles, where ERα increases AMPK activation, and ERβ acts as a repressor, inhibiting AMPK phosphorylation. To translate our findings to heart disease, the next step was to determine the effect of ovarian failure, underlined by E2 loss, on AMPK signaling during the progression of cardiac hypertrophy. We hypothesized that ovarian failure decreases cardiac AMPK signaling, translating in worsening of hypertrophy. We found that the status of cardiac AMPK signaling depends on the nature of the hypertrophic stimulus and the timing of ovarian failure in relation to the onset of hypertrophy. Furthermore, we did not detect any differences in the development of cardiac hypertrophy between wild type mice and mice in ovarian failure, which most likely occur down the line. In summary we described a novel mechanism of AMPK activation by the hormone E2. We also explored the effect of estrogen loss on cardiac AMPK activity, and found that it is dependent on factors such as the pathological state of the heart and timing of the intervention. These findings add to our understanding of the molecular mechanisms behind sex differences in energy handling and in the future could be translated into better therapeutics for the treatment of cardiac pathologies.Release after 16-May-201

Estrogen receptors interact with the alpha catalytic subunit of AMP-activated protein kinase Running title: Estrogen receptors interact with AMPK

SYNOPSIS Normal and pathological stressors engage the AMP-activated protein kinase (AMPK) signaling axis to protect the cell from energetic pressures. Sex steroid hormones also play a critical role in energy metabolism and significantly modify pathological progression of cardiac disease, diabetes/obesity, and cancer. AMPK is targeted by 17β-estradiol (E2), the main circulating estrogen, but the mechanism by which E2 activates AMPK is currently unknown. Using an estrogen receptor α/β (ERα/β) positive (T47D) breast cancer cell line, we validated E2-dependent activation of AMPK that was mediated through ERα (not ERβ) by using three experimental strategies. A series of co-immunoprecipitation experiments showed that both ERs associated with AMPK in cancer and striated (skeletal and cardiac) muscle cells. We further demonstrated direct binding of ERs to the α-catalytic subunit of AMPK within the βγ-subunit binding domain. Finally, both ERs interacted with the upstream LKB1 kinase complex, which is required for E2-dependent activation of AMPK. We conclude that estradiol activates AMPK through ERα by direct interaction with the βγ-binding domain of AMPKα. Summary statement: We identified a novel interaction between the classical estrogen receptors (ERα and ERβ) and the catalytic subunit of AMPK in several cell types. In addition we demonstrate that estradiol activates AMPK through ERα and requires the upstream kinase complex LKB1

Using 4-vinylcyclohexene diepoxide as a model of menopause for cardiovascular disease

There is a sharp rise in cardiovascular disease (CVD) risk and progression with the onset of menopause. The 4-vinylcyclohexene diepoxide (VCD) model of menopause recapitulates the natural, physiological transition through perimenopause to menopause. We hypothesized that menopausal female mice were more susceptible to CVD than pre- or perimenopausal females. Female mice were treated with VCD or vehicle for 20 consecutive days. Premenopausal, perimenopausal, and menopausal mice were administered angiotensin II (ANG II) or subjected to ischemia-reperfusion (I/R). Menopausal females were more susceptible to pathological ANG II-induced cardiac remodeling and cardiac injury from a myocardial infarction (MI), while perimenopausal, like premenopausal, females remained protected. Specifically, ANG II significantly elevated diastolic (130.9 ± 6.0 vs. 114.7 ± 6.2 mmHg) and systolic (156.9 ± 4.8 vs. 141.7 ± 5.0 mmHg) blood pressure and normalized cardiac mass (15.9 ± 1.0 vs. 7.7 ± 1.5%) to a greater extent in menopausal females compared with controls, whereas perimenopausal females demonstrated a similar elevation of diastolic (93.7 ± 2.9 vs. 100.5 ± 4.1 mmHg) and systolic (155.9 ± 7.3 vs. 152.3 ± 6.5 mmHg) blood pressure and normalized cardiac mass (8.3 ± 2.1 vs. 7.5 ± 1.4%) compared with controls. Similarly, menopausal females demonstrated a threefold increase in fibrosis measured by Picrosirus red staining. Finally, hearts of menopausal females (41 ± 5%) showed larger infarct sizes following I/R injury than perimenopausal (18.0 ± 5.6%) and premenopausal (16.2 ± 3.3, 20.1 ± 4.8%) groups. Using the VCD model of menopause, we provide evidence that menopausal females were more susceptible to pathological cardiac remodeling. We suggest that the VCD model of menopause may be critical to better elucidate cellular and molecular mechanisms underlying the transition to CVD susceptibility in menopausal women.NEW & NOTEWORTHY Before menopause, women are protected against cardiovascular disease (CVD) compared with age-matched men; this protection is gradually lost after menopause. We present the first evidence that demonstrates menopausal females are more susceptible to pathological cardiac remodeling while perimenopausal and cycling females are not. The VCD model permits appropriate examination of how increased susceptibility to the pathological process of cardiac remodeling accelerates from pre- to perimenopause to menopause.12 month embargo; published online: 28 May 2020This item from the UA Faculty Publications collection is made available by the University of Arizona with support from the University of Arizona Libraries. If you have questions, please contact us at [email protected]