Sorbs2 Deficiency and Vascular BK Channelopathy in Diabetes.

BACKGROUND: Vascular large conductance Ca2+-activated K+ (BK) channel, composed of the α-subunit (BK-α) and the ß1-subunit (BK-ß1), 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-ß1 subunits. Deletion of the SH3 or SoHo domains abolishes the Sorbs2 effect on the BK-α/BK-ß1 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 Ca2+-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.

A real-world experience of atrioventricular synchronous pacing with leadless ventricular pacemakers.

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 scarce. 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: Sixty-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. CONCLUSION: With 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.

CRRL269.

RATIONALE: Acute kidney injury (AKI) has a high prevalence and mortality in critically ill patients. It is also a powerful risk factor for heart failure incidence driven by hemodynamic changes and neurohormonal activation. However, no drugs have been approved by the Food and Drug Administration. Endogenous pGC-A (particulate guanylyl cyclase A receptor) activators were reported to preserve renal function and improve mortality in AKI patients, although hypotension accompanied by pGC-A activators have limited their therapeutic potential. OBJECTIVE: We investigated the therapeutic potential of a nonhypotensive pGC-A activator/designer natriuretic peptide, CRRL269, in a short-term, large animal model of ischemia-induced AKI and also investigated the potential of uCNP (urinary C-type natriuretic peptide) as a biomarker for AKI. METHODS AND RESULTS: We first showed that CRRL269 stimulated cGMP generation, suppressed plasma angiotensin II, and reduced cardiac filling pressures without lowering blood pressure in the AKI canine model. We also demonstrated that CRRL269 preserved glomerular filtration rate, increased renal blood flow, and promoted diuresis and natriuresis. Further, CRRL269 reduced kidney injury and apoptosis as evidenced by ex vivo histology and tissue apoptosis analysis. We also showed, compared with native pGC-A activators, that CRRL269 is a more potent inhibitor of apoptosis in renal cells and induced less decreases in intracellular Ca2+ concentration in vascular smooth muscle cells. The renal antiapoptotic effects were at least mediated by cGMP/PKG pathway. Further, CRRL269 inhibited proapoptotic genes expression using a polymerase chain reaction gene array. Additionally, we demonstrated that AKI increased uCNP levels. CONCLUSIONS: Our study supports developing CRRL269 as a novel renocardiac protective agent for AKI treatment.

Regulation of KCNMA1 transcription by Nrf2 in coronary arterial smooth muscle cells.

The large conductance Ca2+-activated K+ (BK) channels, composed of the pore-forming α subunits (BK-α, encoded by KCNMA1 gene) and the regulatory ß1 subunits (BK-ß1, encoded by KCNMB1 gene), play a unique role in the regulation of coronary vascular tone and myocardial perfusion by linking intracellular Ca2+ homeostasis with excitation-contraction coupling in coronary arterial smooth muscle cells (SMCs). The nuclear factor erythroid 2-related factor 2 (Nrf2) belongs to a member of basic leucine zipper transcription factor family that regulates the expression of antioxidant and detoxification enzymes by binding to the antioxidant response elements (AREs) of these target genes. We have previously reported that vascular BK-ß1 protein expression was tightly regulated by Nrf2. However, the molecular mechanism underlying the regulation of BK channel expression by Nrf2, particularly at transcription level, is unknown. In this study, we hypothesized that KCNMA1 and KCNMB1 are the target genes of Nrf2 transcriptional regulation. We found that BK channel protein expression and current density were diminished in freshly isolated coronary arterial SMCs of Nrf2 knockout (KO) mice. However, BK-α mRNA expression was reduced, but not that of BK-ß1 mRNA expression, in the arteries of Nrf2 KO mice. Promoter-Nrf2 luciferase reporter assay confirmed that Nrf2 binds to the ARE of KCNMA1 promoter, but not that of KCNMB1. Adenoviral expression and pharmacological activation of Nrf2 increased BK-α and BK-ß1 protein levels and enhanced BK channel activity in coronary arterial SMCs. Hence, our results indicate that Nrf2 is a key determinant of BK channel expression and function in vascular SMCs. Nrf2 facilitates BK-α expression through a direct increase in gene transcription, whereas that on BK-ß1 is through a different mechanism.

F-box protein-32 down-regulates small-conductance calcium-activated potassium channel 2 in diabetic mouse atria.

Diabetes mellitus (DM) is an independent risk factor for atrial fibrillation, but the underlying ionic mechanism for this association remains unclear. We recently reported that expression of the small-conductance calcium-activated potassium channel 2 (SK2, encoded by KCCN2) in atria from diabetic mice is significantly down-regulated, resulting in reduced SK currents in atrial myocytes from these mice. We also reported that the level of SK2 mRNA expression is not reduced in DM atria but that the ubiquitin-proteasome system (UPS), a major mechanism of intracellular protein degradation, is activated in vascular smooth muscle cells in DM. This suggests a possible role of the UPS in reduced SK currents. To test this possibility, we examined the role of the UPS in atrial SK2 down-regulation in DM. We found that a muscle-specific E3 ligase, F-box protein 32 (FBXO-32, also called atrogin-1), was significantly up-regulated in diabetic mouse atria. Enhanced FBXO-32 expression in atrial cells significantly reduced SK2 protein expression, and siRNA-mediated FBXO-32 knockdown increased SK2 protein expression. Furthermore, co-transfection of SK2 with FBXO-32 complementary DNA in HEK293 cells significantly reduced SK2 expression, whereas co-transfection with atrogin-1ΔF complementary DNA (a nonfunctional FBXO-32 variant in which the F-box domain is deleted) did not have any effects on SK2. These results indicate that FBXO-32 contributes to SK2 down-regulation and that the F-box domain is essential for FBXO-32 function. In conclusion, DM-induced SK2 channel down-regulation appears to be due to an FBXO-32-dependent increase in UPS-mediated SK2 protein degradation.

Electrophysiologic effects and outcomes of sympatholysis in patients with recurrent ventricular arrhythmia and structural heart disease.

INTRODUCTION: Autonomic modulation has been used as a therapy to control recurrent ventricular arrhythmia (VA). This study was to explore stellate ganglion block (SGB) effect on cardiac electrophysiologic properties and evaluate the long-term outcome of cardiac sympathetic denervation (CSD) for patients with recurrent VA and structural heart disease (SHD). MATERIALS AND METHODS: Patients who had recurrent VA due to SHD were enrolled prospectively. Electrophysiologic study and ventricular tachycardia (VT) induction were performed before and after left and right SGB. VA burden and long-term outcomes were assessed for a separate patient group who underwent left or bilateral CSD for drug-refractory VA due to SHD. RESULTS: Electrophysiologic study of nine patients showed that baseline mean (SD) corrected sinus node recovery time (cSNRT) increased from 320.4 (73.3) ms to 402.9 (114.2) ms after left and 482.4 (95.7) ms after bilateral SGB (P = .03). SGB did not significantly change P-R, QRS, and Q-T intervals and ventricular effective refractory period, nor did the inducibility of VA. Nineteen patients underwent left (n = 14) or bilateral (n = 5) CSD. CSD reduced VA burden and appropriate ICD therapies from a median (interquartile range) of 2.5 (0.4-11.6) episodes weekly to 0.1 (0.0-2.4) episodes weekly at 6-month follow-up (P = .002). Three-year freedom from orthotopic heart transplant (OHT) and death was 52.6%. New York Heart Association functional class III/IV and VT rate less than 160 beats per minute were predictors of recurrent VA, OHT, and death. CONCLUSION: SGB increased cSNRT without changing heart rate. CSD was more beneficial for patients with mild-to-moderate heart failure and faster VA.

Stellate ganglion block and cardiac sympathetic denervation in patients with inappropriate sinus tachycardia.

BACKGROUND: Inappropriate sinus tachycardia (IST) remains a clinical challenge because patients often are highly symptomatic and not responsive to medical therapy. OBJECTIVE: To study the safety and efficacy of stellate ganglion (SG) block and cardiac sympathetic denervation (CSD) in patients with IST. METHODS: Twelve consecutive patients who had drug-refractory IST (10 women) were studied. According to a prospectively initiated protocol, five patients underwent an electrophysiologic study before and after SG block (electrophysiology study group). The subsequent seven patients had ambulatory Holter monitoring before and after SG block (ambulatory group). All patients underwent SG block on the right side first, and then on the left side. Selected patients who had heart rate reduction ≥15 beats per minute (bpm) were recommended to consider CSD. RESULTS: The mean (SD) baseline heart rate (HR) was 106 (21) bpm. The HR significantly decreased to 93 (20) bpm (P = .02) at 10 minutes after right SG block and remained significantly slower at 97(19) bpm at 60 minutes. Left SG block reduced HR from 99 (21) to 87(16) bpm (P = .02) at 60 minutes. SG block had no significant effect on blood pressure or HR response to isoproterenol or exercise (all P > .05). Five patients underwent right (n = 4) or bilateral (n = 1) CSD. The clinical outcomes were heterogeneous: one patient had complete and two had partial symptomatic relief, and two did not have improvement. CONCLUSION: SG blockade modestly reduces resting HR but has no significant effect on HR during exercise. Permanent CSD may have a modest role in alleviating symptoms in selected patients with IST.

Atrial Fibrillation: Community Screening Events Improve Awareness in Older Adults.

Educating older adults during atrial fibrillation (AF) screening events to recognize signs and symptoms and seek evaluation may promote detection of AF that occurs between screenings. The authors evaluated learning outcomes of AF awareness education provided during AF screening using a single-arm, pre/posttest design. Participants completed the Knowledge, Attitudes, Beliefs about Atrial Fibrillation Self-Monitoring and Treatment-Seeking (KABAF-SMTS) survey, participated in AF awareness education, and completed a KABAF-SMTS survey 2 weeks after education. Paired t tests revealed that knowledge of AF symptoms increased (p = 0.007). Scores for recognizing the seriousness of AF (p = 0.003), benefits of self-monitoring (p < 0.001), perception of barriers to self-monitoring (p = 0.002), and confidence (p < 0.001) to recognize AF and seek treatment improved. AF awareness education strengthened knowledge, beliefs, and attitudes that may be conducive to recognition and treatment-seeking for AF. [Journal of Gerontological Nursing, 45(9), 31-38.].

Role of the endothelial caveolae microdomain in shear stress-mediated coronary vasorelaxation.

In this study, we determined the role of caveolae and the ionic mechanisms that mediate shear stress-mediated vasodilation (SSD). We found that both TRPV4 and SK channels are targeted to caveolae in freshly isolated bovine coronary endothelial cells (BCECs) and that TRPV4 and KCa2.3 (SK3) channels are co-immunoprecipitated by anti-caveolin-1 antibodies. Acute exposure of BCECs seeded in a capillary tube to 10 dynes/cm2 of shear stress (SS) resulted in activation of TRPV4 and SK currents. However, after incubation with HC067047 (TRPV4 inhibitor), SK currents could no longer be activated by SS, suggesting SK channel activation by SS was mediated through TRPV4. SK currents in BCECs were also activated by isoproterenol or by GSK1016790A (TRPV4 activator). In addition, preincubation of isolated coronary arterioles with apamin (SK inhibitor) resulted in a significant diminution of SSD whereas preincubation with HC067047 produced vasoconstriction by SS. Exposure of BCECs to SS (15 dynes/cm2 16 h) enhanced the production of nitric oxide and prostacyclin (PGI2) and facilitated the translocation of TRPV4 to the caveolae. Inhibition of TRPV4 abolished the SS-mediated intracellular Ca2+ ([Ca2+] i ) increase in BCECs. These results indicate a dynamic interaction in the vascular endothelium among caveolae TRPV4 and SK3 channels. This caveolae-TRPV4-SK3 channel complex underlies the molecular and ionic mechanisms that modulate SSD in the coronary circulation.

Impact of Left Atrial Appendage Closure During Cardiac Surgery on the Occurrence of Early Postoperative Atrial Fibrillation, Stroke, and Mortality: A Propensity Score-Matched Analysis of 10 633 Patients.

BACKGROUND: Prophylactic exclusion of the left atrial appendage (LAA) is often performed during cardiac surgery ostensibly to reduce the risk of stroke. However, the clinical impact of LAA closure in humans remains inconclusive. METHODS: Of 10 633 adults who underwent coronary artery bypass grafting and valve surgery between January 2000 and December 2005, 9792 patients with complete baseline characteristics, surgery procedure, and follow-up data were included in this analysis. A propensity score-matching analysis based on 28 pretreatment covariates was performed and 461 matching pairs were derived and analyzed to estimate the association of LAA closure with early postoperative atrial fibrillation (POAF) (atrial fibrillation ≤30 days of surgery), ischemic stroke, and mortality. RESULTS: In the propensity-matched cohort, the overall incidence of POAF was 53.9%. In this group, the rate of early POAF among the patients who underwent LAA closure was 68.6% versus 31.9% for those who did not undergo the procedure (P<0.001). LAA closure was independently associated with an increased risk of early POAF (adjusted odds ratio, 3.88; 95% confidence interval, 2.89-5.20), but did not significantly influence the risk of stroke (adjusted hazard ratio, 1.07; 95% confidence interval, 0.72-1.58) or mortality (adjusted hazard ratio, 0.92; 95% confidence interval, 0.75-1.13). CONCLUSIONS: After adjustment for treatment allocation bias, LAA closure during routine cardiac surgery was significantly associated with an increased risk of early POAF, but it did not influence the risk of stroke or mortality. It remains uncertain whether prophylactic exclusion of the LAA is warranted for stroke prevention during non-atrial fibrillation-related cardiac surgery.

Cryoballoon ablation in Chinese patients with paroxysmal atrial fibrillation: 1-year follow-up.

OBJECTIVES: We assessed the effectiveness and safety of cryoballoon ablation (CBA) in the Chinese population with paroxysmal atrial fibrillation (AF) with a 1-year follow-up and determined the association of early recurrence of atrial tachyarrhythmias (ERAT) with late recurrence (LR). METHODS: A total of 114 patients (age 61 ± 10 years, 78 males) with paroxysmal AF who underwent CBA were consecutively enrolled. After procedures, patients were observed for 3 days with continuous electrocardiogram monitoring in the hospital with routine follow-up visits at 3 months, 6 months, and 1 year. Documented atrial tachyarrhythmia >30 seconds was defined as recurrence. ERAT was defined as any recurrence during the first 3 months, and LR was recurrence between 3 and 12 months. RESULTS: With the first 3 months as blanking period, 76% of patients were free of LR at 12 months. Five patients (4%) experienced complications, including phrenic nerve palsy, stroke, and groin complications. Forty-five percent of patients had ERAT in the first 3 months and 31% of patients had ERAT in the first 3 days. Patients with ERAT had higher LR rate (LRR) than those without ERAT (43% vs 8%, P < 0.001). The LRR of patients with ERAT only in the first 3 days was lower than those with ERAT both in the first 3 days and in 4-90 days (29% vs 64%, P = 0.036). CONCLUSIONS: CBA was an effective and safe treatment option for paroxysmal AF. Patients with ERAT had higher LRR after CBA of AF. The time when ERAT occurred had an impact on LRR.

Down-regulation of the small conductance calcium-activated potassium channels in diabetic mouse atria.

The small conductance Ca(2+)-activated K(+) (SK) channels have recently been found to be expressed in the heart, and genome-wide association studies have shown that they are implicated in atrial fibrillation. Diabetes mellitus is an independent risk factor of atrial fibrillation, but the ionic mechanism underlying this relationship remains unclear. We hypothesized that SK channel function is abnormal in diabetes mellitus, leading to altered cardiac electrophysiology. We found that in streptozotocin-induced diabetic mice, the expression of SK2 and SK3 isoforms was down-regulated by 85 and 92%, respectively, whereas that of SK1 was not changed. SK currents from isolated diabetic mouse atrial myocytes were significantly reduced compared with controls. The resting potentials of isolated atrial preparations were similar between control and diabetic mice, but action potential durations were significantly prolonged in the diabetic atria. Exposure to apamin significantly prolonged action potential durations in control but not in diabetic atria. Production of reactive oxygen species was significantly increased in diabetic atria and in high glucose-cultured HL-1 cells, whereas exposure of HL-1 cells in normal glucose culture to H2O2 reduced the expression of SK2 and SK3. Tyrosine nitration in SK2 and SK3 was significantly increased by high glucose culture, leading to accelerated channel turnover. Treatment with Tiron prevented these changes. Our results suggest that increased oxidative stress in diabetes results in SK channel-associated electrical remodeling in diabetic atria and may promote arrhythmogenesis.

Regulation of large conductance Ca2+-activated K+ (BK) channel ß1 subunit expression by muscle RING finger protein 1 in diabetic vessels.

The large conductance Ca(2+)-activated K(+) (BK) channel, expressed abundantly in vascular smooth muscle cells (SMCs), is a key determinant of vascular tone. BK channel activity is tightly regulated by its accessory ß1 subunit (BK-ß1). However, BK channel function is impaired in diabetic vessels by increased ubiquitin/proteasome-dependent BK-ß1 protein degradation. Muscle RING finger protein 1 (MuRF1), a muscle-specific ubiquitin ligase, is implicated in many cardiac and skeletal muscle diseases. However, the role of MuRF1 in the regulation of vascular BK channel and coronary function has not been examined. In this study, we hypothesized that MuRF1 participated in BK-ß1 proteolysis, leading to the down-regulation of BK channel activation and impaired coronary function in diabetes. Combining patch clamp and molecular biological approaches, we found that MuRF1 expression was enhanced, accompanied by reduced BK-ß1 expression, in high glucose-cultured human coronary SMCs and in diabetic vessels. Knockdown of MuRF1 by siRNA in cultured human SMCs attenuated BK-ß1 ubiquitination and increased BK-ß1 expression, whereas adenoviral expression of MuRF1 in mouse coronary arteries reduced BK-ß1 expression and diminished BK channel-mediated vasodilation. Physical interaction between the N terminus of BK-ß1 and the coiled-coil domain of MuRF1 was demonstrated by pulldown assay. Moreover, MuRF1 expression was regulated by NF-κB. Most importantly, pharmacological inhibition of proteasome and NF-κB activities preserved BK-ß1 expression and BK-channel-mediated coronary vasodilation in diabetic mice. Hence, our results provide the first evidence that the up-regulation of NF-κB-dependent MuRF1 expression is a novel mechanism that leads to BK channelopathy and vasculopathy in diabetes.

Hydrogen sulfide impairs shear stress-induced vasodilation in mouse coronary arteries.

Hydrogen sulfide has emerged as an important endothelium-dependent vasodilator, but its role in shear stress-mediated dilation of coronary arteries is unclear. We examined the role of H2S on shear stress-mediated dilation of isolated mouse coronary arteries. In these vessels, Na2S produced concentration-dependent dilation, which was significantly inhibited by iberiotoxin and by 4-aminopyridine. In addition, BK and Kv currents in mouse coronary smooth muscle cells were directly activated by Na2S, suggesting that H2S produced vasodilation through BK and Kv channel activation. Using a pressure servo controller system, freshly isolated mouse coronary arteries were subjected to physiological levels of shear stress (1 to 25 dynes/cm(2)) and produced graded dilatory responses, but such effects were diminished in the presence of 100 µM Na2S. Pre-incubation with the cystathionine γ-lyase inhibitor, D,L-propargylglycine (PPG), resulted in a paradoxical augmentation of shear stress-mediated vasodilation. However, in the presence of L-NAME or in coronary arteries from eNOS knockout mice, PPG inhibited shear stress-mediated vasodilation, suggesting an interaction between NO and H2S signaling. Na2S inhibited eNOS activity in cultured mouse aortic endothelial cells and reduced the level of phospho-eNOS(serine 1177). These results suggest that both NO and H2S are important shear stress-mediated vasodilators in mouse coronary arteries but there is a complex interaction between these two signaling pathways that results in paradoxical vasoconstrictive effects of H2S through inhibition of NO generation.

Real-time physiologic biomarker for prediction of atrial fibrillation recurrence, stroke, and mortality after electrical cardioversion: A prospective observational study.

BACKGROUND: Left atrial appendage emptying flow velocity (LAAEV) depends largely on left atrioventricular compliance and may play a role in mediating the perpetuation of atrial fibrillation (AF) and AF-related outcomes. METHODS: We identified 3,251 consecutive patients with sustained AF undergoing first-time successful transesophageal echocardiography (TEE)-guided electrical cardioversion who were enrolled in a prospective registry between May 2000 and March 2012. Left atrial appendage emptying flow velocity was stratified into quartiles: ≤20.2, 20.3-33.9, 34-49.9, and ≥50 cm/s. Multivariate Cox regression models were used to identify independent predictors of AF recurrence, ischemic stroke, and all-cause mortality. RESULTS: The mean (SD) age was 69 (12.6) years and 67% were men. Compared with the fourth quartile, patients in the first-third quartiles were significantly older, had higher CHA2DS2-VASc (congestive heart failure, hypertension, age ≥75 years, diabetes mellitus, stroke/transient ischemic attack [TIA], vascular disease, age 65-74 years, sex category) scores, greater frequency of atrial spontaneous echo contrast, and AF of longer duration. Kaplan-Meier analysis showed a decreased probability of event-free survival with decreasing quartiles of LAAEV. Five-year cumulative event rates across first-fourth quartiles were 83%, 80%, 73%, and 73% (P < .001) for first AF recurrence; 7.5%, 7.0%, 4.1%, and 4.0%, for stroke (P = .01); and 31.3%, 26.1%, 24.1%, and 19.4%, for mortality (P < .001), respectively. Multivariate Cox regression analysis revealed an independent association of the first and second quartiles with AF recurrence (P < .001 and P < .001, respectively) and stroke (P = .03, and P = .04, respectively), and of the first quartile with mortality (P = .003). CONCLUSIONS: Patients with decreased LAAEV have an increased risk of AF recurrence, stroke, and mortality after successful electrical cardioversion. Real-time measurement of LAAEV by TEE may be a useful physiologic biomarker for individualizing treatment decisions in patients with AF.

Outcomes and Complications of Lead Removal: Can We Establish a Risk Stratification Schema for a Collaborative and Effective Approach?

BACKGROUND: Removal of an entire cardiovascular implantable electronic device is associated with morbidity and mortality. We sought to establish a risk classification scheme according to the outcomes of transvenous lead removal in a single center, with the goal of using that scheme to guide electrophysiology lab versus operating room extraction. METHODS: Consecutive patients undergoing transvenous lead removal from January 2001 to October 2012 at Mayo Clinic were retrospectively reviewed. RESULTS: A total of 1,378 leads were removed from 652 (age 64 ± 17 years, M 68%) patients undergoing 702 procedures. Mean (standard deviation) lead age was 57.6 (58.8) months. Forty-four percent of leads required laser-assisted extraction. Lead duration (P < 0.001) and an implantable cardioverter defibrillator (ICD) lead (P < 0.001) were associated with the need for laser extraction and procedure failure (P < 0.0001 and P = 0.02). The major complication rate was 1.9% and was significantly associated with longer lead duration (odds ratio: 1.2, 95% confidence interval: 1.1-1.3; P < 0.001). High-risk patients (with a >10-year-old pacing or a >5-year-old ICD lead) had significantly higher major events than moderate-risk (with pacing lead 1-10 years old or ICD lead 1-5 years old) and low-risk (any lead ≤1-year-old) patients (5.3%, 1.2%, and 0%, respectively; P < 0.001). CONCLUSIONS: Transvenous lead removal is highly successful, with few serious procedural complications. We propose a risk stratification scheme that may categorize patients as low, moderate, and high risk for lead extraction. Such a strategy may guide which extractions are best performed in the operating room.

Cardiac Resynchronization Therapy for Patients With Mild to Moderately Reduced Ejection Fraction and Left Bundle Branch Block.

BACKGROUND: It is unknown whether cardiac resynchronization therapy (CRT) would improve or halt the progression of heart failure (HF) in patients with mild to moderately reduced ejection fraction (HFmmrEF) and left bundle branch block (LBBB). OBJECTIVE: This study aimed to investigate the outcomes of CRT in patients with HFmmrEF and left ventricular conduction delay. METHODS: A prospective, randomized clinical trial sponsored by the National Heart, Lung, and Blood Institute included 76 patients who met the study inclusion criteria (left ventricular ejection fraction [LVEF] of 36%-50% and LBBB). Patients received CRT-pacemaker and were randomized to CRT-OFF (right ventricular pacing 40 beats/min) or CRT-ON (biventricular pacing 60-150 beats/min). At a 6-month follow-up, pacing programming was changed to the opposite settings. New York Heart Association class, N-terminal pro-brain natriuretic peptide levels, and echocardiographic variables were collected at baseline, 6 months, and 12 months. The primary study end point was the left ventricular end-systolic volume (LVESV) change from baseline, and the primary randomized comparison was the comparison of 6-month to 12-month changes between randomized groups. RESULTS: The mean age of the patients was 68.4 ± 9.8 years (male, 71%). Baseline characteristics were similar between the 2 randomized groups (all P > .05). In patients randomized to CRT-OFF first, then CRT-ON, LVESV was reduced from baseline only after CRT-ON (baseline, 116.1 ± 36.5 mL; CRT-ON, 87.6 ± 26.0 mL; P < .0001). The randomized analysis of LVEF showed a significantly better change from 6 to 12 months in the OFF-ON group (P = .003). LVEF was improved by CRT (baseline, 41.3% ±.7%; CRT-ON, 46.0% ± 8.0%; P = .002). In patients randomized to CRT-ON first, then CRT-OFF, LVESV was reduced after both CRT-ON and CRT-OFF (baseline, 109.8 ± 23.5 mL; CRT-ON, 91.7 ± 30.5 mL [P < .0001]; CRT-OFF, 99.3 ± 28.9 mL [P = .012]). However, the LVESV reduction effect became smaller between CRT-ON and CRT-OFF (P = .027). LVEF improved after both CRT-ON and CRT-OFF (baseline, 42.7% ± 4.3%; CRT-ON, 48.5% ± 8.6% [P < .001]; CRT-OFF, 45.9% ± 7.7% [P = .025]). CONCLUSION: CRT for patients with HFmmrEF significantly improves LVEF and ventricular remodeling after 6 months of CRT. The study provides novel evidence that early CRT benefits patients with HFmmrEF with LBBB.

Muscle-specific f-box only proteins facilitate bk channel ß(1) subunit downregulation in vascular smooth muscle cells of diabetes mellitus.

RATIONALE: activity of the large conductance Ca(2+)-activated K(+) (BK) channels is profoundly modulated by its ß(1) subunit (BK-ß(1)). However, BK-ß(1) expression is downregulated in diabetic vessels. The ubiquitin-proteasome system (UPS) is a major mechanism of intracellular protein degradation. Whether UPS participates in BK-ß(1) downregulation in diabetic vessels is unknown. OBJECTIVE: we hypothesize that UPS facilitates vascular BK-ß(1) degradation in diabetes. METHODS AND RESULTS: using patch clamp and molecular biological approaches, we found that BK-ß(1)-mediated channel activation and BK-ß(1) protein expression were reduced in aortas of streptozotocin-induced diabetic rats and in human coronary arterial smooth muscle cells (CASMCs) cultured in high glucose. This was accompanied by upregulation of F-box only protein (FBXO)-9 and FBXO-32 (atrogin-1), the key components of the Skp1-Cullin-F-box (SCF) type ubiquitin ligase complex. BK-ß(1) expression was suppressed by the FBXO activator doxorubicin but enhanced by FBXO-9 small interfering RNA or by the proteasome inhibitor MG-132. Cotransfection of atrogin-1 in HEK293 cells significantly reduced Flag-hSlo-ß(1) expression by 2.16-fold, compared with expression of Flag-hSlo-ß(1)V146A (a mutant without the PDZ-binding motif). After cotransfection with atrogin-1, the ubiquitination of Flag-hSlo-ß(1) was increased by 1.91-fold, compared with that of hSlo-ß(1)V146A, whereas cotransfection with atrogin-1ΔF (a nonfunctional mutant without the F-box motif) had no effect. Moreover, inhibition of Akt signaling attenuated the phosphorylation of forkhead box O transcription factor (FOXO)-3a and enhanced atrogin-1 expression, which in turn suppressed BK-ß(1) protein levels in human CASMCs. CONCLUSIONS: downregulation of vascular BK-ß(1) expression in diabetes and in high-glucose culture conditions was associated with FOXO-3a/FBXO-dependent increase in BK-ß(1) degradation.

Regulation of coronary arterial BK channels by caveolae-mediated angiotensin II signaling in diabetes mellitus.

RATIONALE: The large conductance Ca(2+)-activated K(+) (BK) channel, a key determinant of vascular tone, is regulated by angiotensin II (Ang II) type 1 receptor signaling. Upregulation of Ang II functions and downregulation of BK channel activities have been reported in diabetic vessels. However, the molecular mechanisms underlying Ang II-mediated BK channel modulation, especially in diabetes mellitus, have not been thoroughly examined. OBJECTIVES: The aim in this study was to determine whether caveolae-targeting facilitates BK channel dysfunction in diabetic vessels. METHODS AND RESULTS: Using patch clamp techniques and molecular biological approaches, we found that BK channels, Ang II type 1 receptor, G(alphaq/11) (G protein q/11 alpha subunit), nonphagocytic NAD(P)H oxidases (NOX-1), and c-Src kinases (c-Src) were colocalized in the caveolae of rat arterial smooth muscle cells and the integrity of caveolae in smooth muscle cells was critical for Ang II-mediated BK channel regulation. Most importantly, membrane microdomain targeting of these proteins was upregulated in the caveolae of streptozotocin-induced rat diabetic vessels, leading to enhanced Ang II-induced redox-mediated BK channel modification and causing BK channel and coronary dysfunction. The absence of caveolae abolished the effects of Ang II on vascular BK channel activity and preserved BK channel function in diabetes. CONCLUSIONS: These results identified a molecular scheme of receptor/enzyme/channel/caveolae microdomain complex that facilitates the development of vascular BK channel dysfunction in diabetes.

TRPC6 N338S is a gain-of-function mutant identified in patient with doxorubicin-induced cardiotoxicity.

The canonical transient receptor potential 6 gene, TRPC6, has been implicated as a putative risk gene for chemotherapy-induced congestive heart failure, but knowledge of specific risk variants is lacking. Following our genome-wide association study and subsequent fine-mapping, a rare missense mutant of TRPC6 N338S, was identified in a breast cancer patient who received anthracycline-containing chemotherapy regiments and developed congestive heart failure. However, the function of N338S mutant has not been examined. Using intracellular Ca2+ imaging, patch clamp recording and molecular docking techniques, we assessed the function of N338S mutant heterologously expressed in HEK293 cells and HL-1 cardiac cells. We found that expression of TRPC6 N338S significantly increased intracellular Ca2+ levels ([Ca2+]i) and current densities in response to 50 µM 1-oleoyl 2-acetyl-sn-glycerol (OAG), an activator of TRPC6 channels, compared to those of TRPC6 WT. A 24-h pretreatment with 0.5 µM doxorubicin (DOX) further potentiated the OAG effects on TRPC6 N338S current densities and [Ca2+]i, and these effects were abolished by 1 µM BI-749327, a highly selective TRPC6 inhibitor. Moreover, DOX treatment significantly upregulated the mRNA and protein expressions of TRPC6 N338S, compared to those of TRPC6 WT. Molecular docking and dynamics simulation showed that OAG binds to the pocket constituted by the pore-helix, S5 and S6 domains of TRPC6. However, the N338S mutation strengthened the interaction with OAG, therefore stabilizing the OAG-TRPC6 N338S complex and enhancing OAG binding affinity. Our results indicate that TRPC6 N338S is a gain-of-function mutant that may contribute to DOX-induced cardiotoxicity by increasing Ca2+ influx and [Ca2+]i in cardiomyocytes.