Citations

BACKGROUND: Vascular large conductance Ca 2+ -activated K + (BK) channel, composed of the α-subunit (BK-α) and the β1-subunit (BK-β1), is a key determinant of coronary vasorelaxation and its function is impaired in diabetic vessels. However, our knowledge of diabetic BK channel dysregulation is incomplete. The Sorbs2 (Sorbin homology [SoHo] and Src homology 3 [SH3] domains-containing protein 2), is ubiquitously expressed in arteries, but its role in vascular pathophysiology is unknown. METHODS: The role of Sorbs2 in regulating vascular BK channel activity was determined using patch-clamp recordings, molecular biological techniques, and in silico analysis. RESULTS: Sorbs2 is not only a cytoskeletal protein but also an RNA-binding protein that binds to BK channel proteins and BK-α mRNA, regulating BK channel expression and function in coronary smooth muscle cells. Molecular biological studies reveal that the SH3 domain of Sorbs2 is necessary for Sorbs2 interaction with BK-α subunits, while both the SH3 and SoHo domains of Sorbs2 interact with BK-β1 subunits. Deletion of the SH3 or SoHo domains abolishes the Sorbs2 effect on the BK-α/BK-β1 channel current density. Additionally, Sorbs2 is a target gene of the Nrf2 (nuclear factor erythroid-2-related factor 2), which binds to the promoter of Sorbs2 and regulates Sorbs2 expression in coronary smooth muscle cells. In vivo studies demonstrate that Sorbs2 knockout mice at 4 months of age display a significant decrease in BK channel expression and function, accompanied by impaired BK channel Ca 2+ -sensitivity and BK channel-mediated vasodilation in coronary arteries, without altering their body weights and blood glucose levels. Importantly, Sorbs2 expression is significantly downregulated in the coronary arteries of db/db type 2 diabetic mice. CONCLUSIONS: Sorbs2, a downstream target of Nrf2, plays an important role in regulating BK channel expression and function in vascular smooth muscle cells. Vascular Sorbs2 is downregulated in diabetes. Genetic knockout of Sorbs2 manifests coronary BK channelopathy and vasculopathy observed in diabetic mice, independent of obesity and glucotoxicity.

Introduction: Although the prevalence of atrial fibrillation (AF) is higher in men than in women, the onset of AF appears to diminish the survival advantage in women. The pathophysiologic factors underlying survival outcome in AF are not well understood. Methods: A cohort of 3480 Olmsted County, Minnesota residents (mean age 72.8±14 years, 51.8% female) with incident AF who underwent transthoracic echocardiography within 30 days of AF diagnosis [median (IQR) 1, 1-6 days] were retrospectively studied. We examined the association of clinical parameters and survival according to gender. Kaplan-Meier and Cox proportional hazards models were used to compare overall survival in males and females. Results: Females were older and slimmer than males and more frequently had hypertension, prior stroke, and had higher CHA 2 DS 2 vasc score, pulse pressure and effective arterial elastance. Women also had smaller LV end diastolic dimension with higher LV ejection fraction, LA volume index, E/e prime ratio, and PASP (43.6±13.8 vs 39.1±13.3; <.001). During a mean follow-up of 7.4±6.2 years (median 6.6 years), 2569 patients died. Women had a lower 5-year survival compared to men. The probability of 1- and 5-year survival rates for women vs men were 80.2% vs 84.1% at 1 year and 54.4% vs 63.7% at 5 years (log-rank P <.001). In multivariable Cox regression analysis, after accounting for potential confounders, the independent predictors of mortality were age (HR 1.07 [95% CI 1.06-1.08]), PASP (per 5 unit increment) (HR 1.11 [95% CI 1.09-1.12]), effective arterial elastance (HR 1.06 [95% CI 1.01-1.12]), previous heart failure (HR 1.34 [95% CI 1.20-1.49]), diabetes (HR 1.33 [95% CI 1.20-1.48]), prior stroke (HR 1.26 [95% CI 1.13-1.41]), smoking (HR 1.13 [95% CI 1.06-1.20]), chronic lung disease (HR 1.32 [95% CI 1.20-1.45]), peripheral arterial disease (HR, 1.32 [95% CI 1.18-1.48]), and chronic kidney disease (HR, 1.52 [95% CI 1.38-1.69]). Conclusions: In a community-based cohort of patients with incident AF, markers of arterial stiffness and diastolic dysfunction were more prevalent in women compared to men and were strongly predictive of mortality. Our findings suggest that vascular stiffness may underlie the increased risk of mortality observed in women after a new diagnosis of AF.

Introduction: Doxorubicin (DOX) is widely used for chemotherapy, but cardiotoxicity and cardiomyopathy are major adverse effects. The canonical transient receptor potential 6 gene ( TRPC6) is a putative risk gene for cardiomyopathy. Recently, a TRPC6 N338S mutation was found in a patient with DOX-induced heart failure. However, the TRPC6 N338S mutant has not been characterized, and how it affects cardiac function is unknown. Hypothesis: We hypothesized that TRPC6 N338S is a gain-of-function mutation producing intracellular Ca 2+ overload in cardiomyocytes. Methods: Using patch clamp recording, molecular biological and molecular docking analyses, we assessed the function of TRPC6 N338S heterologously expressed in HL-1 cardiac cells. Results: TRPC6 N338S significantly increased the current densities of TRPC6 (defined as 1 μM BI-749327 sensitive components, a highly selective TRPC6 inhibitor) in HL-1 cells at baseline and after exposure to 50 μM 1-oleoyl 2-acetyl-sn-glycerol (OAG, an activator of TRPC6), compared to cells expressing TRPC6 WT. Moreover, a 24-h pretreatment with 0.5 μM DOX further potentiated the OAG effects on TRPC6 WT currents with a significantly greater response in the N338S currents. Ca 2+ imaging also showed enhanced intracellular Ca 2+ levels in cells expressing the N338S mutant with OAG and DOX treatment. Biological studies found that the cytoplasmic and surface expression of TRPC6 WT and N338S mutant were identical in HL-1 cells, but those of TRPC6 N338S were markedly upregulated after treatment with 0.5 μM DOX, compared to TRPC6 WT. Molecular docking and dynamics simulation analysis unraveled that OAG binds to a pocket constituted by the pore-helix, S5 and S6 domains of TRPC6 through polar interactions. OAG binds to the same site in TRPC6 N338S with shortened interfacial distances due to additional polar and hydrogen bonding between OAG and the N338S mutant. Conclusions: TRPC6 N338S mutation strengthens the interaction with OAG, stabilizes the OAG-TRPC6 N338S complex, and enhances the OAG effects. Treatment with DOX significantly increases the expression and function of TRPC6 N338S in HL-1 cells. Hence, the TRPC6 N338S is a gain-of-function mutation and may contribute to cardiotoxicity in patients with anthracycline treatment.

Background: Aldosterone is a critical pathological driver for cardiac and renal diseases. We recently discovered that mutant atrial natriuretic peptide (MANP), a novel atrial natriuretic peptide (ANP) analog, possessed more potent aldosterone inhibitory action than ANP in vivo. MANP and natriuretic peptide (NP)-augmenting therapy sacubitril/valsartan are under investigations for human hypertension treatment. Understanding the elusive mechanism of aldosterone inhibition by NPs remains to be a priority. Conflicting results were reported on the roles of the pGC-A (particulate guanylyl cyclase A receptor) and NP clearance receptor in aldosterone inhibition. Furthermore, the function of PKG (protein kinase G) and PDEs (phosphodiesterases) on aldosterone regulation are not clear. Methods: In the present study, we investigated the molecular mechanism of aldosterone regulation in a human adrenocortical cell line H295R and in mice. Results: We first provided evidence to show that pGC-A, not NP clearance receptor, mediates aldosterone inhibition. Next, we confirmed that MANP inhibits aldosterone via PDE2 (phosphodiesterase 2) not PKG, with specific agonists, antagonists, siRNA silencing, and fluorescence resonance energy transfer experiments. Further, the inhibitory effect is mediated by a reduction of intracellular Ca2+ levels. We then illustrated that MANP directly reduces aldosterone synthase CYP11B2 (cytochrome p450 family 11 subfamily b member 2) expression via PDE2. Last, in PDE2 knockout mice, consistent with in vitro findings, embryonic adrenal CYP11B2 is markedly increased. Conclusions: Our results innovatively explore and expand the NP/pGC-A/3',5', cyclic guanosine monophosphate (cGMP)/PDE2 pathway for aldosterone inhibition by MANP in vitro and in vivo. In addition, our data also support the development of MANP as a novel ANP analog drug for aldosterone excess treatment.

Abstract Objective . Arterial dispersion ultrasound vibrometry (ADUV) relies on the use of guided waves in arterial geometries for shear wave elastography measurements. Both the generation of waves through the use of acoustic radiation force (ARF) and the techniques employed to infer the speed of the resulting wave motion affect the spectral content and accuracy of the measurement. In particular, the effects of the shape and location of the ARF beam in ADUV have not been widely studied. In this work, we investigated how such variations of the ARF beam affect the induced motion and the measurements in the dispersive modes that are excited. Approach. The study includes an experimental evaluation on an arterial phantom and an in vivo validation of the observed trends, observing the two walls of the waveguide, simultaneously, when subjected to variations in the ARF beam extension ( F / N ) and focus location. Main results. Relying on the theory of guided waves in cylindrical shells, the shape of the beam controls the selection and nature of the induced modes, while the location affects the measured dispersion curves (i.e. variation of phase velocity with frequency or wavenumber, multiple modes) across the waveguide walls. Significance. This investigation is important to understand the spectral content variations in ADUV measurements and to maximize inversion accuracy by tuning the ARF beam settings in clinical applications.

AbstractAimsThe MicraTM transcatheter pacing system (TPS) (Medtronic) is the only leadless pacemaker that promotes atrioventricular (AV) synchrony via accelerometer‐based atrial sensing. Data regarding the real‐world experience with this novel system are scarce. We sought to characterize patients undergoing MicraTM‐AV implants, describe percentage AV synchrony achieved, and analyze the causes for suboptimal AV synchrony.MethodsIn this retrospective cohort study, electronic medical records from 56 consecutive patients undergoing MicraTM‐AV implants at the Mayo Clinic sites in Minnesota, Florida, and Arizona with a minimum follow‐up of 3 months were reviewed. Demographic data, comorbidities, echocardiographic data, and clinical outcomes were compared among patients with and without atrial synchronous ventricular pacing (AsVP) ≥ 70%.ResultsSixty‐five percent of patients achieved AsVP ≥ 70%. Patients with adequate AsVP had smaller body mass indices, a lower proportion of congestive heart failure, and prior cardiac surgery. Echocardiographic parameters and procedural characteristics were similar across the two groups. Active device troubleshooting was associated with higher AsVP. The likely reasons for low AsVP were small A4‐wave amplitude, high ventricular pacing burden, and inadequate device reprogramming. Importantly, in patients with low AsVP, subjective clinical worsening was not noted during follow‐up.ConclusionWith the increasing popularity of leadless pacemakers, it is paramount for device implanting teams to be familiar with common predictors of AV synchrony and troubleshooting with MicraTM‐AV devices.

Objective To investigate the association of mitral regurgitation (MR) on thromboembolic risk of patients with non-valvular atrial fibrillation (NVAF) undergoing transoesophageal echocardiography (TEE)-guided cardioversion. Methods Data for consecutive patients who underwent TEE-guided cardioversion for NVAF between 2000 and 2012 were analysed. MR severity was assessed by Doppler echocardiography and classified as ≤mild, moderate or severe. Left atrial appendage emptying velocities were averaged for five consecutive cycles. Multivariable regression models were used to identify independent predictors of left atrial appendage thrombus (LAAT) and stroke. Results 2950 patients (age, 69.3±12.2 years, 67% men) were analysed. 2173 (73.7%) had ≤mild MR; 631 (21.4%), moderate MR; and 146 (4.9%), severe MR. Patients with moderate (age, 72.4±10.7 years) and severe (age, 72.8±12.1 years) MR were older than those with ≤mild MR (age, 68.2±12.5 years). The prevalence of LAAT was 1.5% (n=43). CHA 2 DS 2 -VASc scores (≤mild MR, 3.0±1.6; moderate MR, 3.5±1.5; severe MR, 3.9±1.5; p<0.001) and heart failure frequency (≤mild MR, 38.4%; moderate MR, 48.0%; severe MR, 69.2%; p<0.001) were increasingly higher with greater MR severity. Multivariable logistic regression analysis showed no association of moderate MR (OR 0.77, 95% CI 0.38 to 1.56) or severe MR (OR 0.55, 95% CI 0.21 to 1.49) with LAAT. During a mean follow-up of 7.3±5.1 years (median 7.5, IQR, 2.7–10.9), 216 patients had an ischaemic stroke. Adjusted Cox regression analysis showed no significant association of moderate MR (HR 1.22, 95% CI 0.88 to 1.68) or severe MR (HR 0.73, 95% CI 0.31 to 1.46) with stroke. Conclusions Among patients with NVAF, the presence or severity of MR was not associated with a decreased risk of LAAT or stroke.

Aims: The MicraTM transcatheter pacing system (TPS) (Medtronic) is the only leadless pacemaker that promotes atrioventricular (AV) synchrony via accelerometer-based atrial sensing. Data regarding the real-world experience with this novel system are currently lacking. We sought to characterize patients undergoing MicraTM -AV implants, describe percentage AV synchrony achieved, and analyze the causes for suboptimal AV synchrony. Methods: In this retrospective cohort study, electronic medical records from 56 consecutive patients undergoing MicraTM -AV implants at the Mayo Clinic sites in Minnesota, Florida, and Arizona with a minimum follow-up of 3 months were reviewed. Demographic data, comorbidities, echocardiographic data, and clinical outcomes were compared among patients with and without atrial synchronous-ventricular pacing (AsVP) ≥70%. Results: Fifty-six percent of patients achieved AsVP ≥70%. Patients with adequate AsVP had smaller body mass indices, a lower proportion of congestive heart failure and pulmonary hypertension. Echocardiographic parameters and procedural characteristics were similar across the two groups. Active device troubleshooting was associated with higher AsVP. The likely reasons for low AsVP were persistent atrial arrhythmias, small A4-wave amplitude, and inadequate device reprogramming. Importantly, in patients with low AsVP, subjective clinical worsening was not noted during follow-up. Conclusion: With the increasing popularity of leadless PM, it is paramount for device implanting teams to be familiar with common predictors of AV synchrony and troubleshooting with MicraTM -AV devices.

Aldosterone is a critical driver for cardiovascular disease (CVD). We recently discovered that MANP, a novel atrial natriuretic peptide (ANP) analog, possessed more potent aldosterone inhibitory action than ANP. MANP is currently entering clinical trials for hypertension and thus understanding its aldosterone suppressing mechanism is important. The mechanism of aldosterone inhibition by natriuretic peptides (NPs) remains to be clearly defined. Conflicting results were reported on the roles of particulate guanylyl cyclase A receptor (pGC-A) and NP clearance receptor (NPRC) in aldosterone inhibition. Furthermore, the functions of protein kinase G (PKG) and phosphodiesterases (PDE) on aldosterone regulation are not clear. Herein, we investigated the molecular mechanism of aldosterone regulation in the human adrenocortical cell line H295R and in mice. We firstly showed that pGC-A mediates aldosterone inhibition. In contrast, with NPRC agonist and antagonist, we showed that NPRC did not inhibit aldosterone. Next, we confirmed that MANP inhibits aldosterone via PDE2, not PKG, with specific agonists, antagonists, siRNA silencing, and fluorescence resonance energy transfer (FRET) experiments. Specifically, MANP suppressed ANGII mediated activation of aldosterone (fold change) MANP+ANGII 3.2±0.1* vs. ANGII 3.8±0.1 (*p<0.05) with IBMX, a PDEs inhibitor and the PDE2 antagonist Bay 60-7550 reversed MANP-mediated aldosterone suppression (IBMX+MANP+ANGII 3.9±0.2 and Bay+MANP+ANGII 4.1±0.1). With PKG agonists and inhibitors, aldosterone levels were not changed. In PDE2 activity FRET studies, aldosterone control was 3.7±0.4 and with MANP 0.9±0.2* supporting PDE2 activation by MANP. Further, the inhibitory effect of PDE2 is mediated by a reduction of intracellular Ca2+ concentration (~22%). We then showed that MANP directly reduced aldosterone synthase CYP11B2 expression in vitro. Lastly, in PDE2 knockout mice (embryonic lethal), embryonic adrenal CYP11B2 expression is markedly increased (wild type: 1±0.2, KO: 2.8±0.5*). Our findings innovatively elucidate the pGC-A/cGMP/PDE2 pathway in aldosterone inhibition by MANP in vitro and in vivo. Additionally, our data also support the development of MANP as a novel ANP analog drug for CVD.

BK channels, abundantly expressed in vascular smooth muscle cells (VSMCs), are the key determinant of vascular physiology. BK channel activity is tightly regulated by its accessory β 1 (BK-β 1 ) subunits. BK-β 1 protein expression is down-regulated in diabetic vessels, contributing to diabetic vascular complications including hypertension, stroke and coronary heart disease. However, BK-β 1 mRNA is not reduced in diabetic vessels suggesting increased BK-β 1 turnover. Since ubiquitin-proteasome system (UPS) accounts for 80~90% of intracellular protein degradations in mammalian cells, we hypothesize that muscle ring finger-1 (MuRF-1) protein, the muscle specific E3 ligase, accelerates vascular BK-β 1 degradation in diabetes mellitus. In this study, we found that BK-β 1 protein level was decreased by 1.95±0.14 fold, accompanied by a 2.83±0.16 fold increase of MuRF-1 expression in streptozotocin-induced diabetic mouse vessels (n=3, p<0.05 vs. control). Moreover, response of vascular BK channel activation by DHS-1 (a specific BK-β 1 activator) was absent in diabetic mouse vascular smooth muscle cells (VSMCs). Similar results were observed in human VSMCs cultured with 22 mM glucose (HG) compared to those cultured with 5 mM glucose. Knockdown of MuRF-1 by siRNA (100 nM) produced a 4.2-fold decrease of MuRF-1 protein level and a 1.8-fold increase in BK-β 1 expression in HG-cultured VSMCs. In addition, the total BK-β 1 expression was 2.58±0.04 fold lower but the ubiquitinated BK-β 1 protein was 5.18±0.04 fold higher in HEK293 cells 48-h after transfection with MuRF-1, compared to those with MuRF-1ΔR (an inactive mutant with the Ring structure deletion) transfection (n=3, p<0.05). Immunoprecipitation experiments confirmed that the N-terminus of BK-β 1 was physically associated with MuRF-1. Inhibition of UPS activity by 10 μM MG-132 (a proteasome inhibitor) enhanced BK-β 1 expression by 1.70±0.01 fold (n=3, p<0.05 vs. control). Our results indicate that MuRF-1 participated in ubiquitin-mediated vascular BK-β 1 protein proteasomal degradation and delineated a fundamental mechanism underlying vasculopathy in diabetes.