Faculty Bibliography
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176
Connexin hemichannels as candidate targets for cardioprotective and anti-arrhythmic treatments
Connexins are crucial cardiac proteins that form hemichannels and gap junctions. Gap junctions are responsible for the propagation of electrical and chemical signals between myocardial cells and cells of the specialized conduction system in order to synchronize the cardiac cycle and steer cardiac pump function. Gap junctions are normally open, while hemichannels are closed, but pathological circumstances may close gap junctions and open hemichannels, thereby perturbing cardiac function and homeostasis. Current evidence demonstrates an emerging role of hemichannels in myocardial ischemia and arrhythmia, and tools are now available to selectively inhibit hemichannels without inhibiting gap junctions as well as to stimulate hemichannel incorporation into gap junctions. We review available experimental evidence for hemichannel contributions to cellular pro-arrhythmic events in ventricular and atrial cardiomyocytes, and link these to insights at the level of molecular control of connexin-43-based hemichannel opening. We conclude that a double-edged approach of both preventing hemichannel opening and preserving gap junctional function will be key for further research and development of new connexin-based experimental approaches for treating heart disease.
PMCID:10014111
PMID: 36919695
ISSN: 1558-8238
CID: 5448872
Nanogold based protein localization enables subcellular visualization of cell junction protein by SBF-SEM
Recent advances in volume electron microscopy (vEM) allow unprecedented visualization of the electron-dense structures of cells, tissues and model organisms at nanometric resolution in three dimensions (3D). Light-based microscopy has been widely used for specific localization of proteins; however, it is restricted by the diffraction limit of light, and lacks the ability to identify underlying structures. Here, we describe a protocol for ultrastructural detection, in three dimensions, of a protein (Connexin 43) expressed in the intercalated disc region of adult murine heart. Our protocol does not rest on the expression of genetically encoded proteins and it overcomes hurdles related to pre-embedding and immunolabeling, such as the penetration of the label and the preservation of the tissue. The pre-embedding volumetric immuno-electron microscopy (pre-embedding vIEM) protocol presented here combines several practical strategies to balance sample fixation with antigen and ultrastructural preservation, and penetration of labeling with blocking of non-specific antigen binding sites. The small 1.4 nm gold along with surrounded silver used as a detection marker buried in the sample also serves as a functional conductive resin that significantly reduces the charging of samples. Our protocol also presents strategies for facilitating the successful cutting of the samples during serial block-face scanning electron microscopy (SBF-SEM) imaging. Our results suggest that the small gold-based pre-embedding vIEM is an ideal labeling method for molecular localization throughout the depth of the sample at subcellular compartments and membrane microdomains.
PMID: 37451776
ISSN: 0091-679x
CID: 5535332
Annual review of genomics & human genetics. 2022:23:255-274.DOI: 10.1146/annurev-genom-112921-011200
The Genetics of Brugada Syndrome
Brugada syndrome is a heritable channelopathy characterized by a peculiar electrocardiogram (ECG) pattern and increased risk of cardiac arrhythmias and sudden death. The arrhythmias originate because of an imbalance between the repolarizing and depolarizing currents that modulate the cardiac action potential. Even if an overt structural cardiomyopathy is not typical of Brugada syndrome, fibrosis and structural changes in the right ventricle contribute to a conduction slowing, which ultimately facilitates ventricular arrhythmias. Currently, Mendelian autosomal dominant transmission is detected in less than 25% of all clinical confirmed cases. Although 23 genes have been associated with the condition, only SCN5A, encoding the cardiac sodium channel, is considered clinically actionable and disease causing. The limited monogenic inheritance has pointed toward new perspectives on the possible complex genetic architecture of the disease, involving polygenic inheritance and a polygenic risk score that can influence penetrance and risk stratification. Expected final online publication date for the Annual Review of Genomics and Human Genetics, Volume 23 is October 2022. Please see http://www.annualreviews.org/page/journal/pubdates for revised estimates.
PMID: 35567276
ISSN: 1545-293x
CID: 5215132
Loss of Nuclear Envelope Integrity and Increased Oxidant Production Cause DNA Damage in Adult Hearts Deficient in PKP2: A Molecular Substrate of ARVC
BACKGROUND:gene, which encodes the PKP2 protein (plakophilin-2). METHODS:studied at a time of preserved left ventricular ejection fraction and in human induced pluripotent stem cell-derived PKP2-deficient myocytes. RESULTS: CONCLUSIONS:
PMID: 35959657
ISSN: 1524-4539
CID: 5287322
American journal of physiology. Cell physiology. 2022:322(6):C1230-C1247.DOI: 10.1152/ajpcell.00099.2022
KATP channel trafficking
Sarcolemmal/plasmalemmal ATP-sensitive K+ (KATP) channels have key roles in many cell types and tissues. Hundreds of studies have described how the KATP channel activity and ATP sensitivity can be regulated by changes in the cellular metabolic state, by receptor signaling pathways and by pharmacological interventions. These alterations in channel activity directly translate to alterations in cell or tissue function, that can range from modulating secretory responses, such as insulin release from pancreatic β-cells or neurotransmitters from neurons, to modulating contractile behavior of smooth muscle or cardiac cells to elicit alterations in blood flow or cardiac contractility. It is increasingly becoming apparent, however, that KATP channels are regulated beyond changes in their activity. Recent studies have highlighted that KATP channel surface expression is a tightly regulated process with similar implications in health and disease. The surface expression of KATP channels is finely balanced by several trafficking steps including synthesis, assembly, anterograde trafï¬cking, membrane anchoring, endocytosis, endocytic recycling and degradation. This review aims to summarize the physiological and pathophysiological implications of KATP channel trafficking and mechanisms that regulate KATP channel trafficking. A better understanding of this topic has potential to identify new approaches to develop therapeutically useful drugs to treat KATP channel-related diseases.
PMID: 35508187
ISSN: 1522-1563
CID: 5216232
Exercise Causes Arrhythmogenic Remodeling of Intracellular Calcium Dynamics in Plakophilin-2-Deficient Hearts
BACKGROUND: METHODS:Experiments were performed in myocytes from a cardiomyocyte-specific, tamoxifen-activated, PKP2 knockout murine line (PKP2cKO). For training, mice underwent 75 minutes of treadmill running once per day, 5 days each week for 6 weeks. We used multiple approaches including imaging, high-resolution mass spectrometry, electrocardiography, and pharmacological challenges to study the functional properties of cells/hearts in vitro and in vivo. RESULTS: CONCLUSIONS:
PMCID:9086182
PMID: 35491884
ISSN: 1524-4539
CID: 5235702
Author Correction: Genome-wide association analyses identify new Brugada syndrome risk loci and highlight a new mechanism of sodium channel regulation in disease susceptibility
PMID: 35474365
ISSN: 1546-1718
CID: 5205632
Preserved cardiac performance and adrenergic response in a rabbit model with decreased ryanodine receptor 2 expression
Ryanodine receptor 2 (RyR2) is an ion channel in the heart responsible for releasing into the cytosol most of the Ca2+ required for contraction. Proper regulation of RyR2 is critical, as highlighted by the association between channel dysfunction and cardiac arrhythmia. Lower RyR2 expression is also observed in some forms of heart disease; however, there is limited information on the impact of this change on excitation-contraction (e-c) coupling, Ca2+-dependent arrhythmias, and cardiac performance. We used a constitutive knock-out of RyR2 in rabbits (RyR2-KO) to assess the extent to which a stable decrease in RyR2 expression modulates Ca2+ handling in the heart. We found that homozygous knock-out of RyR2 in rabbits is embryonic lethal. Remarkably, heterozygotes (KO+/-) show ~50% loss of RyR2 protein without developing an overt phenotype at the intact animal and whole heart levels. Instead, we found that KO+/- myocytes show (1) remodeling of RyR2 clusters, favoring smaller groups in which channels are more densely arranged; (2) lower Ca2+ spark frequency and amplitude; (3) slower rate of Ca2+ release and mild but significant desynchronization of the Ca2+ transient; and (4) a significant decrease in the basal phosphorylation of S2031, likely due to increased association between RyR2 and PP2A. Our data show that RyR2 deficiency, although remarkable at the molecular and subcellular level, has only a modest impact on global Ca2+ release and is fully compensated at the whole-heart level. This highlights the redundancy of RyR2 protein expression and the plasticity of the e-c coupling apparatus.
PMID: 35413295
ISSN: 1095-8584
CID: 5201912
Beta-blocker/ACE inhibitor therapy differentially impacts the steady state signaling landscape of failing and non-failing hearts
Heart failure is a multifactorial disease that affects an estimated 38 million people worldwide. Current pharmacotherapy of heart failure with reduced ejection fraction (HFrEF) includes combination therapy with angiotensin-converting enzyme inhibitors (ACEi) and β-adrenergic receptor blockers (β-AR blockers), a therapy also used as treatment for non-cardiac conditions. Our knowledge of the molecular changes accompanying treatment with ACEi and β-AR blockers is limited. Here, we applied proteomics and phosphoproteomics approaches to profile the global changes in protein abundance and phosphorylation state in cardiac left ventricles consequent to combination therapy of β-AR blocker and ACE inhibitor in HFrEF and control hearts. The phosphorylation changes induced by treatment were profoundly different for failing than for non-failing hearts. HFrEF was characterized by profound downregulation of mitochondrial proteins coupled with derangement of β-adrenergic and pyruvate dehydrogenase signaling. Upon treatment, phosphorylation changes consequent to HFrEF were reversed. In control hearts, treatment mainly led to downregulation of canonical PKA signaling. The observation of divergent signaling outcomes depending on disease state underscores the importance of evaluating drug effects within the context of the specific conditions present in the recipient heart.
PMCID:8934364
PMID: 35306519
ISSN: 2045-2322
CID: 5190982
Luminal Oxidative Regulation of the Ryanodine Receptor: More Sides to the Story? [Editorial]
PMID: 35239403
ISSN: 1524-4571
CID: 5174622
