Lee, Hon-Chi

Background: We have previously reported that Sorbs2 (sorbin and SH3 domain 2) knockout mice have significant cardiac structural and electrical remodeling, including atrial enlargement and fibrosis. The mechanisms of such changes were unknown. Hypothesis: We hypothesized that loss of Sorbs2 in cardiac fibroblasts promotes profibrotic signaling pathways and alters the balance between matrix metalloproteinases (MMPs), and their inhibitors (TIMPs) resulting in excessive collagen deposition in the atria. Methods: We used Sorbs2 global knockout ( Sorbs2 KO) mice and age-matched wild-type (WT) controls at 4 to 5 months of age. Left atrial fibrosis was quantitated by Masson's trichrome staining in serial sections. Scratch wound healing assays and transwell migration assays were performed in isolated Sorbs2 KO and WT atrial fibroblasts. Protein expression was determined by Western blots. Interleukin-6 (IL-6) production in cultured fibroblast supernatants was measured by ELISA. Results: Sorbs2 KO mice showed significant increase in atrial diameter (16.6 ± 1.5 mm vs 12.0 ± 1.1 mm in WT; n=3, p < 0.05) and atrial fibrosis (fraction of fibrotic areas: 18.6± 2.9 % vs. 7.6 ± 0.9 % in WT; p < 0.01). Collagen 1 and collagen 3 protein levels in Sorbs2 KO mouse atria were 1.8-fold and 1.7-fold higher than those of WT respectively. Sorbs2 KO atrial fibroblasts showed impaired function with significantly reduced migration and slower scratch-wound closure (migration area after 24h: 18.5 ± 3.8 % vs 70.1 ± 6.7 % in WT; n=6, p < 0.01).Moreover, transforming growth factor β1 (TGF-β1), α-smooth muscle actin, and periostin expression levels were significantly reduced in isolated fibroblasts and atrial myocardia from Sorbs2 KO mice, suggesting that fibrosis is not caused by TGF-b1-dependent mechanisms or from myofibroblast transformation. Analysis of ECM proteins showed significant downregulation of MMP1 and MMP3 but upregulation of TIMP1, while TIMP3 was unchanged. Consistent with a pro-inflammatory phenotype, Sorbs2 KO atrial fibroblasts secreted higher IL-6 levels at baseline (3303± 463 pg/mL vs. 2075 ± 731 pg/mL in WT; n=6, p < 0.01) and with LPS stimulation (7791 ± 604 pg/mL vs. 5588 ± 763 pg/mL in WT; n=6, p < 0.01). Conclusion: Sorbs2 deficiency is associated with abnormal atrial fibroblast function showing impaired migration and scratch wound healing, as well as increased IL-6 production and imbalanced MMP/TIMP. These changes result in collagen accumulation and enhanced fibrosis.

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.

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.

Objective Previous studies have postulated a causal role of patent foramen ovale (PFO) in the aetiology of embolic stroke in the general population. We hypothesised that the presence of concomitant PFO and atrial fibrillation (AF) will add incremental risk of ischaemic stroke to that linked to AF alone. Methods We analysed data on 3069 consecutive patients (mean age 69.4±12.2 years; 67.1% men) undergoing transoesophageal echocardiography-guided electrical cardioversion (ECV) for AF between May 2000 and March 2012. PFO was identified by colour Doppler and agitated saline contrast study. All patients were followed up after ECV for first documentation of ischaemic stroke. Outcomes were compared using Cox regression models. Results The prevalence of PFO was 20.0% and the shunt direction was left-to-right in the majority of patients (71.4%). Patients with PFO had a higher frequency of obstructive sleep apnoea (21.7% vs 17.1%, p=0.01) and higher mean peak left atrial appendage emptying velocity (38.3±21.8 vs 36.1±20.4 cm/s; p=0.04) compared with those without PFO. Otherwise, baseline characteristics were similar between groups. During a mean follow-up period of 7.3±4.6 years, 214 patients (7.0%) had ischaemic stroke. Multivariable analysis showed no significant association between PFO and ischaemic stroke (HR, 0.82 (95% CI 0.57 to 1.18)). PFO shunt direction was strongly associated with stroke: HR, 1.91 (95% CI 1.16 to 3.16) for right-to-left shunt and HR, 0.58 (95% CI 0.36 to 0.93) for left-to-right shunt. Conclusions The presence of concurrent PFO in this largely anticoagulated group of patients with AF was not associated with increased risk of ischaemic stroke.

Background: Percutaneous stellate ganglion blockade (SGB) has been used for drug-refractory electrical storm due to ventricular arrhythmia (VA); however, the effects and long-term outcomes have not been well studied. Methods: This study included 30 consecutive patients who had drug-refractory electrical storm and underwent percutaneous SGB between October 1, 2013, and March 31, 2018. Bupivacaine, alone or combined with lidocaine, was injected into the neck with good local anesthetic spread in the vicinity of the left stellate ganglion (n=15) or both stellate ganglia (n=15). Data were collected for patient clinical characteristics, immediate and long-term outcomes, and procedure-related complications. Results: Clinical characteristics included age, 58±14 years; men, 73.3%; and left ventricular ejection fraction, 34±16%. At 24 hours, 60% of patients were free of VA. Patients whose VA was controlled had a lower hospital mortality rate than patients whose VA continued (5.6% versus 50.0%; P =0.009). Implantable cardioverter-defibrillator interrogation showed a significant 92% reduction in VA episodes from 26±41 to 2±4 in the 72 hours after SGB ( P <0.001). Patients who died during the same hospitalization (n=7) were more likely to have ischemic cardiomyopathy (100% versus 43.5%; P =0.03) and recurrent VA within 24 hours (85.7% versus 26.1%; P =0.009). There were no procedure-related major complications. Conclusions: SGB effectively attenuated electrical storm in more than half of patients without procedure-related complications. Percutaneous SGB may be considered for stabilizing ventricular rhythm in patients for whom other therapies have failed.

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 Ca 2+ 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.

Abstract Aims Type 2 diabetes (T2D) is strongly associated with cardiovascular morbidity and mortality in patients. Vascular large conductance Ca2+-activated potassium (BK) channels, composed of four pore-forming α subunits (BK-α), and four regulatory β1 subunits (BK-β1), are densely expressed in coronary arterial smooth muscle cells (SMCs) and play an important role in regulating vascular tone and myocardial perfusion. However, the role of BK channels in coronary microvascular dysfunction of human subjects with diabetes is unclear. In this study, we examined BK channel function and protein expression, and BK channel-mediated vasodilation in freshly isolated coronary arterioles from T2D patients. Methods and results Atrial tissues were obtained from 16 patients with T2D and 25 matched non-diabetic subjects during cardiopulmonary bypass procedure. Microvessel videomicroscopy and immunoblot analysis were performed in freshly dissected coronary arterioles and inside-out single BK channel currents was recorded in enzymatically isolated coronary arteriolar SMCs. We found that BK channel sensitivity to physiological Ca2+ concentration and voltage was downregulated in the coronary arteriolar SMCs of diabetic patients, compared with non-diabetic controls. BK channel kinetics analysis revealed that there was significant shortening of the mean open time and prolongation of the mean closed time in diabetic patients, resulting in a remarkable reduction of the channel open probability. Functional studies showed that BK channel activation by dehydrosoyasaponin-1 was diminished and that BK channel-mediated vasodilation in response to shear stress was impaired in diabetic coronary arterioles. Immunoblot experiments confirmed that the protein expressions of BK-α and BK-β1 subunits were significantly downregulated, but the ratio of BK-α/BK-β1 was unchanged in the coronary arterioles of T2D patients. Conclusions Our results demonstrated for the first time that BK channel function and BK channel-mediated vasodilation were abnormal in the coronary microvasculature of diabetic patients, due to decreased protein expression and altered intrinsic properties of BK channels.

The large conductance Ca2+-activated K+ (BK) channel β1-subunit (BK-β1) is a key modulator of BK channel electrophysiology and the downregulation of BK-β1 protein expression in vascular smooth muscle cells (SMCs) underlies diabetic vascular dysfunction. In this study, we hypothesized that the nuclear factor erythroid-2–related factor 2 (Nrf2) signaling pathway plays a significant role in the regulation of coronary BK channel function and vasodilation in high-fat diet (HFD)–induced obese/diabetic mice. We found that the protein expressions of BK-β1 and Nrf2 were markedly downregulated, whereas those of the nuclear factor-κB (NF-κB) and the muscle ring finger protein 1 (MuRF1 [a ubiquitin E3 ligase for BK-β1]) were significantly upregulated in HFD mouse arteries. Adenoviral expression of Nrf2 suppressed the protein expressions of NF-κB and MuRF1 but enhanced BK-β1 mRNA and protein expressions in cultured coronary SMCs. Knockdown of Nrf2 resulted in reciprocal changes of these proteins. Patch-clamp studies showed that coronary BK-β1–mediated channel activation was diminished in HFD mice. Importantly, the activation of Nrf2 by dimethyl fumarate significantly reduced the body weight and blood glucose levels of HFD mice, enhanced BK-β1 transcription, and attenuated MuRF1-dependent BK-β1 protein degradation, which in turn restored coronary BK channel function and BK channel–mediated coronary vasodilation in HFD mice. Hence, Nrf2 is a novel regulator of BK channel function with therapeutic implications in diabetic vasculopathy.

BK channels are major ionic determinants of vasodilation. BK channel function is impaired in diabetic vessels due to accelerated proteolysis of its beta-1 (BK-β1) subunits in response to increased oxidative stress. The nuclear factor E2-related factor-2 (Nrf2) signalling pathway has emerged as a master regulator of cellular redox status, and we hypothesized that it plays a central role in regulating BK channel function in diabetic vessels. We found that Nrf2 expression was markedly reduced in db/db diabetic mouse aortas, and this was associated with significant downregulation of BK-β1. In addition, the muscle ring finger protein 1 (MuRF1), a known E-3 ligase targeting BK-β1 ubiquitination and proteasomal degradation, was significantly augmented. These findings were reproduced by knockdown of Nrf2 by siRNA in cultured human coronary artery smooth muscle cells. In contrast, adenoviral transfer of Nrf2 gene in these cells downregulated MuRF1 and upregulated BK-β1 expression. Activation of Nrf2 by dimethyl fumarate preserved BK-β1 expression and protected BK channel and vascular function in db/db coronary arteries. These results indicate that expression of BK-β1 is closely regulated by Nrf2 and vascular BK channel function can be restored by Nrf2 activation. Nrf2 should be considered a novel therapeutic target in the treatment of diabetic vasculopathy.

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.