Faculty Bibliography

Background: KAT P channels couple cellular metabolism and electrophysiology. Their molecular composition varies in different tissues and species. Rodent atrial KAT P channels have the SUR1 regulatory subunit, are activated by diazoxide and have been implicated in arrhythmogenesis in hypertension and excess beta-adrenergic tone. In contrast, human atrial KATP channels are insensitive to diazoxide and modulate APD only during extreme metabolic stress, where the SUR2A regulatory subunit is thought to be predominant. Objective: We hypothesized that changes in the human atrial action potential associated with beta-agonism are mediated by recruitment of SUR1-containing KATP channels. Methods: We used human induced pluripotent stem cell (hiPSC)-derived atrial cardiomyocytes where expression of a fuorescent reporter is driven by the atrial-specifc gene sarcolipin. Atrial specifcation was induced with retinoic acid. Di-4-ANBDQBS was used to perform optical action potential measurements on days 65-80 of differentiation. Excised patch clamping was used to evaluate KAT P channel density. Heterozygous ABCC8 (SUR1+/-) cells were generated using CRISPR/CAS9. Results: Optical mapping data are for APD90 with stimulation at 1.25 Hz The combination of isoproterenol (ISO, 10mu M) and rolipram (ROL, 10mu M) abbreviated APD compared to control (247.4+/-12.5ms, n=16 vs 344.2+/-22.9ms, n=22; p=0.002). This was ameliorated by 10mu M glibenclamide (312.0+/-18.9ms, n=23 vs 247.4+/-12.5ms, n=16; p=0.01). More patches from cells exposed to ISO and ROL had functional KATP channels (4/22 vs 0/24, p=0.045). Diazoxide shortened APD (267.3+/-21.7ms, n=20 vs 344.2+/-22.9ms, n=22; p=0.02). This was potentiated by prior beta-agonism (179.7+/-14.3ms, n=18 vs 267.3+/-21.7ms, n=20; p=0.002). Deletion of one ABCC8 allele ameliorated APD shortening with exposure to ISO, ROL, and diazoxide (240.9+/-18.2ms, n=14 vs 179.7+/-14.3ms, n=18; p=0.012). Functional KATP channel density after exposure to beta-agonists was reduced in SUR1+/-cells (1/40 vs 4/22, p=0.049). Conclusion: SUR1-containing KATP channels partially mediate beta-adrenergic APD shortening in human atrial cells and may represent a therapeutic target for atrial arrhythmia prevention

Background: Genetic/genomic research has greatly advanced our understanding of heart disease. Yet a comprehensive map of the protein landscape of living human hearts is still lacking. Since cardiac protein degradation progresses rapidly post-mortem, it is essential to limit studies to samples collected from live individuals and immediately process these for data acquisition. Objective: To defne the human cardiac chamber-specifc proteome from samples collected in vivo. Methods: Cardiac biopsies of right atria (RA), left atria (LA) and left ventricle (LV) were collected from seven humans undergoing open chest surgery and analyzed by high-resolution mass spectrometry. Results: We identifed 7314 proteins across cardiac chambers. Proteome-wide analysis revealed hundreds of proteins differentially expressed between RA and LA, and between atria and LV with high statistical signifcance. We found over-representation of: a) fbrosis-related proteins in LA, b) autonomic regulation-related proteins in RA, and c) sarcomeric and intercalated disc proteins in LV. We used our data to advance disease-related leads obtained via genetics/genomics. Specifcally, previous GWA studies identifed six loci associated with mitral valve prolapse (MVP). We queried our dataset for abundance of proteins coded by all genes within those 6 loci, recovering gene candidates for fve of these: two genes previously associated with MVP and four new ones. Separately, we found signifcant chamber-specifc over-representation of disease-causing dilated cardiomyopathy variants in LV, with the corresponding observation that protein abundance in LV is signifcantly higher for high-confdence variants. Conclusion: We present the frst comprehensive atlas of chamber-specifc human cardiac protein expression obtained from samples collected in vivo. We identify hundreds of proteins with chamber-specifc expression; some correspond to known functional characteristics and others to novel chamber-specifc identifers, with potential as drug targets. Our studies offer a crucial link between genomic data and the mechanisms of disease

Atherosclerosis, the underlying cause of coronary artery (CAD) and other cardiovascular diseases, is initiated by macrophage-mediated immune responses to lipoprotein and cholesterol accumulation in artery walls, which result in the formation of plaques. Unlike at other sites of inflammation, the immune response becomes maladaptive and inflammation fails to resolve. The most common treatment for reducing the risk from atherosclerosis is low density lipoprotein cholesterol (LDL-C) lowering. Studies have shown, however, that while significant lowering of LDL-C reduces the risk of heart attacks to some degree, there is still residual risk for the majority of the population. We and others have observed "residual inflammatory risk" of atherosclerosis after plasma cholesterol lowering in pre-clinical studies, and that this phenomenon is clinically relevant has been dramatically reinforced by the recent Canakinumab Anti-inflammatory Thrombosis Outcomes Study (CANTOS) trial. This review will summarize the role of the innate immune system, specifically macrophages, in atherosclerosis progression and regression, as well as the pre-clinical and clinical models that have provided significant insights into molecular pathways involved in the resolution of plaque inflammation and plaque regression. Partnered with clinical studies that can be envisioned in the post-CANTOS period, including progress in developing targeted plaque therapies, we expect that pre-clinical studies advancing on the path summarized in this review, already revealing key mechanisms, will continue to be essential contributors to achieve the goals of dampening plaque inflammation and inducing its resolution in order to maximize the therapeutic benefits of conventional risk factor modifications, such as LDL-C lowering.

Co-administration of sofosbuvir, an anti-hepatitis C virus medication, and antiarrhythmic amiodarone causes symptomatic severe bradycardia in patients and animal models. However, in a few in vitro studies, the combination of sofosbuvir and amiodarone resulted in tachycardiac effects in human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs). This discrepancy may be attributable to the use of immature hiPSC-CMs in the in vitro studies. To address this, we evaluated the ability of our in-house hiPSC-CMs to assess the interactions between sofosbuvir and amiodarone in vitro. We performed whole-cell patch recordings on hiPSC-CMs to examine the cardiac effect of sofosbuvir and amiodarone, alone or in combination. We found that sofosbuvir and amiodarone caused bradycardiac effects (the beating rate decreased to 75% of the vehicle control, P < 0.001) on our hiPSC-CMs when applied in combination, but they had no significant effect when applied alone. Furthermore, the bradycardiac effect was membrane potential dependent: it increased with depolarization. This raised the possibility that the bradycardiac effects in vivo may originate in nodal cells, which have a more depolarized resting membrane potential compared with ventricular cells. The bradycardiac effects of sofosbuvir plus amiodarone in vitro are consistent with the clinical phenotype and suggest that our hiPSC-CMs may serve as a useful tool in assessing cardiac safety during drug discovery and development process.

Smooth muscle cells (SMCs) play a key role in atherogenesis. However, mechanisms regulating expansion and fate of pre-existing SMCs in atherosclerotic plaques remain poorly defined. Here we show that multiple SMC progenitors mix to form the aorta during development. In contrast, during atherogenesis, a single SMC gives rise to the smooth muscle-derived cells that initially coat the cap of atherosclerotic plaques. Subsequently, highly proliferative cap cells invade the plaque core, comprising the majority of plaque cells. Reduction of integrin β3 (Itgb3) levels in SMCs induces toll-like receptor 4 expression and thereby enhances Cd36 levels and cholesterol-induced transdifferentiation to a macrophage-like phenotype. Global Itgb3 deletion or transplantation of Itgb3^(-/-) bone marrow results in recruitment of multiple pre-existing SMCs into plaques. Conditioned medium from Itgb3-silenced macrophages enhances SMC proliferation and migration. Together, our results suggest SMC contribution to atherogenesis is regulated by integrin β3-mediated pathways in both SMCs and bone marrow-derived cells.

BACKGROUND:Atrial standstill is an arrhythmogenic condition characterized by the absence of spontaneous electrical and mechanical atrial activity or in response to stimulation. There are few reported familial cases which have been associated with SCN5A mutations co-segregating with GJA5 or RYR2 however isolated SCN5A mutations are rare. OBJECTIVE:The purpose of this study was to determine the clinical and biophysical consequence of a novel SCN5A mutation identified in a family with progressive atrial standstill and sudden death. METHODS:The family of a sporadic case of congenital atrial standstill underwent genetic screening. Human Embryonic Kidney 293 cells were transfected with wild-type (WT) or mutant SCN5A cDNAs. Biophysical properties were studied using whole-cell using patch clamp methods. RESULTS:A novel homozygous SCN5A mutation, p.V1340L was identified in the proband and her sister. The proband had complete atrial standstill whereas the sister had partial atrial standstill. Heterozygous mutations were identified in the mother, father and brother. All three had normal sinus rhythm and were asymptomatic. The mutant Nav1.5(V1340L) reduced Nav1.5 current density as well as showed a depolarizing shift in the voltage-dependent steady-state activation (WT: -35.3±1.62 mV; V1340L: -22.4±2.59 mV; P = 0.001). CONCLUSIONS:A homozygous loss-of-function SCN5A mutation likely results in atrial standstill and sudden death due to suppression of initiation of action potential.

The most recent financial upheavals have cast doubt on the adequacy of some of the conventional quantitative risk management strategies, such as VaR (Value at Risk), in many common situations. Consequently, there has been an increasing need for verisimilar financial stress testings, namely simulating and analyzing financial portfolios in extreme, albeit rare scenarios. Unlike conventional risk management which exploits statistical correlations among financial instruments, here we focus our analysis on the notion of probabilistic causation, which is embodied by Suppes-Bayes Causal Networks (SBCNs); SBCNs are probabilistic graphical models that have many attractive features in terms of more accurate causal analysis for generating financial stress scenarios. In this paper, we present a novel approach for conducting stress testing of financial portfolios based on SBCNs in combination with classical machine learning classification tools. The resulting method is shown to be capable of correctly discovering the causal relationships among financial factors that affect the portfolios and thus, simulating stress testing scenarios with a higher accuracy and lower computational complexity than conventional Monte Carlo simulations.

After activation, cells of the myeloid lineage undergo robust metabolic transitions, as well as discrete epigenetic changes, that can dictate both ongoing and future inflammatory responses. In atherosclerosis, in which macrophages play central roles in the initiation, growth, and ultimately rupture of arterial plaques, altered metabolism is a key feature that dictates macrophage function and subsequent disease progression. This Review explores how factors central to the plaque microenvironment (for example, altered cholesterol metabolism, oxidative stress, hypoxia, apoptotic and necrotic cells, and hyperglycemia) shape the metabolic rewiring of macrophages in atherosclerosis as well as how these metabolic shifts in turn alter macrophage immune-effector and tissue-reparative functions. Finally, this overview offers insight into the challenges and opportunities of harnessing metabolism to modulate aberrant macrophage responses in disease.

Annexin A6 (AnxA6) is expressed in articular chondrocytes at levels higher than in other mesenchymal cell types. However, the role of AnxA6 in articular chondrocytes is not known. Here we show that complete lack of AnxA6 functions resulted in increased ß-catenin activation in Wnt3a-treated murine articular chondrocytes, whereas AnxA6 expressing articular chondrocytes showed decreased ß-catenin activation. High expression of AnxA6 in human articular chondrocytes showed the highest inhibition of Wnt/ß-catenin signaling. Inhibition of Wnt/ß-catenin signaling activity by AnxA6 together with cytosolic Ca2+ was achieved by interfering with the plasma membrane association of the Wnt signaling complex. AnxA6 also affected the cross-talk between Wnt/ß-catenin signaling and NF-κB signaling by decreasing ß-catenin activity and increasing NF-κB activity in Wnt3a-, interleukin-1beta (IL-1ß)-, and combined Wnt3a/IL-1ß-treated cells. Wnt3a treatment increased the mRNA levels of catabolic markers (cyclooxygenase-2, interleukin-6, inducible nitric oxide synthase) to a much lesser degree than IL-1ß treatment in human articular chondrocytes, and decreased the mRNA levels of matrix metalloproteinase-13 (MMP-13) and articular cartilage markers (aggrecan, type II collagen). Furthermore, Wnt3a decreased the mRNA levels of catabolic markers and MMP-13 in IL-1ß-treated human articular chondrocytes. High expression of AnxA6 resulted in decreased mRNA levels of catabolic markers, and increased MMP-13 and articular cartilage marker mRNA levels in Wnt3a-treated human articular chondrocytes, whereas leading to increased mRNA levels of catabolic markers and MMP-13 in human articular chondrocytes treated with IL-1ß, or combined Wnt3a and IL-1ß. Our findings define a novel role for AnxA6 in articular chondrocytes via its modulation of Wnt/ß-catenin and NF-κB signaling activities and the cross-talk between these two signaling pathways.

BACKGROUND:Patients undergoing cardiovascular (CV) procedures often have suboptimal CV risk factor control and may benefit from strategies targeting healthy lifestyle behaviors and education. Implementation of prevention strategies may be particularly effective at this point of heightened motivation. METHODS:A prospective, randomized, pilot study was conducted in 400 patients undergoing a nonurgent CV procedure (cardiac catheterization ± revascularization) to evaluate the impact of different prevention strategies. Patients were randomized in a 1:1:1 fashion to usual care (UC; group A, n = 134), in-hospital CV prevention consult (PC; group B, n = 130), or PC plus behavioral intervention program (telephone-based motivational interviewing and optional tailored text messages) (group C, n = 133). The primary end point was the Δ change in non-high-density lipoprotein cholesterol (non-HDL-C) from baseline to 6 month. RESULTS:The mean age was 64.6 ± 10.8 years, 23.7% were female, and 31.5% were nonwhite. After 6 months, the absolute difference in non-HDL-C for all participants was -19.8 mg/dL (95% CI -24.1 to -15.6, P < .001). There were no between-group differences in the primary end point for the combined PC groups (B and C) versus UC, with a Δ adjusted between group difference of -5.5 mg/dL (95% CI -13.1 to 2.1, P = .16). Patients in the PC groups were more likely to be on high-intensity statins at 6 months (52.9% vs 38.1%, P = .01). After excluding participants with baseline non-HDL-C <100 mg/dL (initial exclusion criterion), Δ non-HDL-C and Δ low-density lipoprotein cholesterol were improved in the PC groups compared to UC (non-HDL-C -8.13 mg/dL [-16.00 to -0.27], P = .04; low-density lipoprotein cholesterol -7.87mg/dL [-15.10 to -0.64], P = .03). CONCLUSIONS:Although non-HDL-C reduction at 6 months following a nonurgent CV procedure was not significant in the overall cohort, an increased uptake in high-potency statins may translate into improved long-term health outcomes and cost reductions.