Connexin Remodeling Research Articles

The Synergistic Relationship Between Atrial Fibrillation and Diabetes Mellitus: Implications for Cardiovascular and Metabolic Health.

Type 2 diabetes mellitus (T2DM) and atrial fibrillation (AF) are widespread chronic conditions that profoundly impact public health. While the intricate mechanisms linking these two diseases remain incompletely understood, this review sets out to comprehensively analyze the current evidence about their pathophysiology, epidemiology, diagnosis, prognosis, and treatment. We reveal that T2DM can influence the electrical and structural properties of the atria through multiple pathways, including oxidative stress, inflammation, fibrosis, connexin remodeling, glycemic variability, and autonomic dysfunction. Moreover, it significantly influences AF's clinical course, elevating the risk of heart failure, stroke, and cardiovascular mortality. Our review also explores treatment options for individuals with T2DM and AF, encompassing antidiabetic and antiarrhythmic drugs and non-pharmacological interventions, such as cardioversion catheter ablation and direct current cardioversion. This review depicts an insight into the clinical interplay between T2DM and AF. It deepens our comprehension of the fundamental mechanisms, potential therapeutic interventions, and their implications for patient care. This comprehensive resource benefits researchers seeking to deepen their knowledge in this domain. Ultimately, our findings pave the way for more effective strategies in managing AF within the context of T2DM.

Cannabinoid receptor agonist attenuates angiotensin II-induced enlargement and mitochondrial dysfunction in rat atrial cardiomyocytes.

Pathological remodeling of atrial tissue renders the atria more prone to arrhythmia upon arrival of electrical triggers. Activation of the renin-angiotensin system is an important factor that contributes to atrial remodeling, which may result in atrial hypertrophy and prolongation of P-wave duration. In addition, atrial cardiomyocytes are electrically coupled via gap junctions, and electrical remodeling of connexins may result in dysfunction of coordinated wave propagation within the atria. Currently, there is a lack of effective therapeutic strategies that target atrial remodeling. We previously proposed that cannabinoid receptors (CBR) may have cardioprotective qualities. CB13 is a dual cannabinoid receptor agonist that activates AMPK signaling in ventricular cardiomyocytes. We reported that CB13 attenuates tachypacing-induced shortening of atrial refractoriness and inhibition of AMPK signaling in the rat atria. Here, we evaluated the effects of CB13 on neonatal atrial rat cardiomyocytes (NRAM) stimulated by angiotensin II (AngII) in terms of atrial myocyte enlargement and mitochondrial function. CB13 inhibited AngII-induced enhancement of atrial myocyte surface area in an AMPK-dependent manner. CB13 also inhibited mitochondrial membrane potential deterioration in the same context. However, AngII and CB13 did not affect mitochondrial permeability transition pore opening. We further demonstrate that CB13 increased Cx43 compared to AngII-treated neonatal rat atrial myocytes. Overall, our results support the notion that CBR activation promotes atrial AMPK activation, and prevents myocyte enlargement (an indicator that suggests pathological hypertrophy), mitochondrial depolarization and Cx43 destabilization. Therefore, peripheral CBR activation should be further tested as a novel treatment strategy in the context of atrial remodeling.

Trimethylamine N-oxide facilitates the progression of atrial fibrillation in rats with type 2 diabetes by aggravating cardiac inflammation and connexin remodeling.

Diabetes is an independent risk factor for atrial fibrillation (AF). This study aimed to elucidate the pathophysiology of diabetes-related AF from the perspective of the gut microbial metabolite trimethylamine N-oxide (TMAO). In the present study, male rats received either a normal diet to serve as the control group or a high-fat diet/streptozotocin to induce type 2 diabetes mellitus. Then, diabetic rats were divided into two groups based on the presence or absence of 3,3-dimethyl-1-butanol (DMB, a specific TMAO inhibitor) in drinking water: the diabetic cardiomyopathy (DCM) group and the DCM + DMB group. Eight weeks later, compared with control rats, rats in the DCM group exhibited gut microbiota dysbiosis and systemic TMAO elevation. The inflammatory cytokines IL-1β, IL-6, and TNF-α were markedly increased in the atria of rats in the DCM group. Downregulated expression of connexin 40 and lateralized distribution of connexin 43 were also observed in the atria of DCM rats. AF inducibility was significantly higher in DCM rats than in control rats. Furthermore, DMB treatment effectively ameliorated atrial inflammation and connexin remodeling while markedly reducing plasma TMAO levels. DMB treatment also decreased the vulnerability of diabetic rats to AF. In conclusion, TMAO might promote atrial inflammation and connexin remodeling in the development of diabetes, which may play a key role in mediating diabetes-related AF.

Repolarization Heterogeneity in Human Post-Infarct Ventricular Tachycardia.

Slow conduction, caused by fibrosis between surviving myocytes and connexin remodeling, is an important prerequisite for post-infarction ventricular tachycardia (VT); however, slow conduction is present throughout the infarct whereas VT circuits are finite in number and discrete. In a porcine model of VT, re-entrant circuits occur at region of significant repolarization heterogeneity caused by up-regulation of potassium channel β-subunits KCNE3 (increasing repolarization current) and KCNE4 (decreasing repolarization current), causing heterogeneous action potential durations. This study was designed to determine whether re-entrant circuits in human post-infarction VT are associated with repolarization heterogeneity. In 6 patients, left ventricular mapping was performed during induced VT to identify sites within the VT circuit. Subsequently, unipolar mapping (3.5-mm tip ablation catheter) was performed to characterize activation-recovery intervals (ARIs), which are surrogates for local action potential durations, at sites documented within the VT circuit isthmus (IN) compared to sites within the infarct scar but outside of the VT circuit (OUT). ARIs were significantly shorter in the IN compared with the OUT sites (420.2 ± 79.3 ms vs 462 ± 52.8ms; P=0.01). In all patients. sites that were associated with the circuit always had shorter ARI values than did those sampled from OUT regions. VT circuit sites in human post-infarct VT are associated with repolarization heterogeneity, similar to what was previously reported in a porcine model. This suggests the possibility of a common mechanism between humans and the porcine model of post-infarct VT, and that development of ablation strategies or small molecule or genetic therapies to restore normal repolarization kinetics may be antiarrhythmic.

Atrial Myocyte NLRP3/CaMKII Nexus Forms a Substrate for Postoperative Atrial Fibrillation.

Postoperative atrial fibrillation (POAF) is a common and troublesome complication of cardiac surgery. POAF is generally believed to occur when postoperative triggers act on a preexisting vulnerable substrate, but the underlying cellular and molecular mechanisms are largely unknown. To identify cellular POAF mechanisms in right atrial samples from patients without a history of atrial fibrillation undergoing open-heart surgery. Multicellular action potentials, membrane ion-currents (perforated patch-clamp), or simultaneous membrane-current (ruptured patch-clamp) and [Ca2+]i-recordings in atrial cardiomyocytes, along with protein-expression levels in tissue homogenates or cardiomyocytes, were assessed in 265 atrial samples from patients without or with POAF. No indices of electrical, profibrotic, or connexin remodeling were noted in POAF, but Ca2+-transient amplitude was smaller, although spontaneous sarcoplasmic reticulum (SR) Ca2+-release events and L-type Ca2+-current alternans occurred more frequently. CaMKII (Ca2+/calmodulin-dependent protein kinase-II) protein-expression, CaMKII-dependent phosphorylation of the cardiac RyR2 (ryanodine-receptor channel type-2), and RyR2 single-channel open-probability were significantly increased in POAF. SR Ca2+-content was unchanged in POAF despite greater SR Ca2+-leak, with a trend towards increased SR Ca2+-ATPase activity. Patients with POAF also showed stronger expression of activated components of the NLRP3 (NACHT, LRR, and PYD domains-containing protein-3)-inflammasome system in atrial whole-tissue homogenates and cardiomyocytes. Acute application of interleukin-1β caused NLRP3-signaling activation and CaMKII-dependent RyR2/phospholamban hyperphosphorylation in an immortalized mouse atrial cardiomyocyte cell-line (HL-1-cardiomyocytes) and enhanced spontaneous SR Ca2+-release events in both POAF cardiomyocytes and HL-1-cardiomyocytes. Computational modeling showed that RyR2 dysfunction and increased SR Ca2+-uptake are sufficient to reproduce the Ca2+-handling phenotype and indicated an increased risk of proarrhythmic delayed afterdepolarizations in POAF subjects in response to interleukin-1β. Preexisting Ca2+-handling abnormalities and activation of NLRP3-inflammasome/CaMKII signaling are evident in atrial cardiomyocytes from patients who subsequently develop POAF. These molecular substrates sensitize cardiomyocytes to spontaneous Ca2+-releases and arrhythmogenic afterdepolarizations, particularly upon exposure to inflammatory mediators. Our data reveal a potential cellular and molecular substrate for this important clinical problem.

Integrin-Linked Kinase Activation Prevents Ventricular Arrhythmias Induced by Ischemia/Reperfusion Via Inhibition of Connexin 43 Remodeling

Ischemia reperfusion (I/R)-induced arrhythmia is a serious complication in patients with cardiac infarction. Remodeling of connexin (Cx) 43, manifested as phosphorylation, contributes significantly to arrhythmogenesis. Integrin-linked kinase (ILK) attenuated ventricular remodeling and improved cardiac function in rats after myocardial infarction. We hypothesized that ILK, through Cx43 phosphorylation, would be protective against I/R-induced ventricular arrhythmias. Our study showed that I/R-induced ventricular arrhythmias were attenuated by an ILK agonist LPTP and worsened by the ILK inhibitor Cpd22. I/R disrupted Cx43 distribution, but it was partially normalized in the presence of LPTP. Compared with I/R, the phosphorylation of Akt was increased significantly after pretreatment with LPTP. The increase in phosphorylated Akt was physiologically significant because, in the presence of the Akt inhibitor MK2206, the protective effects of LPTP were blocked. This indicated that ILK activation prevented I/R-induced-ventricular arrhythmia, an effect potentially related to inhibition of Cx43 remodeling via Akt activation.

Beneficial and Detrimental Remodeling of Glial Connexin and Pannexin Functions in Rodent Models of Nervous System Diseases.

A variety of glial cell functions are supported by connexin and pannexin proteins. These functions include the modulation of synaptic gain, the control of excitability through regulation of the ion and neurotransmitter composition of the extracellular milieu and the promotion of neuronal survival. Connexins and pannexins support these functions through diverse molecular mechanisms, including channel and non-channel functions. The former comprise the formation of gap junction-mediated networks supported by connexin intercellular channels and the formation of pore-like membrane structures or hemichannels formed by both connexins and pannexins. Non-channel functions involve adhesion properties and the participation in signaling intracellular cascades. Pathological conditions of the nervous system such as ischemia, neurodegeneration, pathogen infection, trauma and tumors are characterized by distinctive remodeling of connexin expression and function. However, whether these changes can be interpreted as part of the pathogenesis, or as beneficial compensatory effects, remains under debate. Here we review the available evidence addressing this matter with a special emphasis in mouse models with selective manipulation of glial connexin and pannexin proteins in vivo. We postulate that the beneficial vs. detrimental effects of glial connexin remodeling in pathological conditions depend on the impact of remodeling on the different connexin and pannexin channel and non-channel functions, on the characteristics of the inflammatory environment and on the type of interaction among glial cells types.

Intermittent hypoxia causes NOX2-dependent remodeling of atrial connexins

BackgroundObstructive sleep apnea has been linked to the development of heart disease and arrhythmias, including atrial fibrillation. Since altered conduction through gap junction channels can contribute to the pathogenesis of such arrhythmias, we examined the abundance and distributions of the major cardiac gap junction proteins, connexin40 (Cx40) and connexin43 (Cx43) in mice treated with sleep fragmentation or intermittent hypoxia (IH) as animal models of the components of obstructive sleep apnea.ResultsWild type C57BL/6 mice or mice lacking NADPH 2 (NOX2) oxidase activity (gp91phox(−/Y)) were exposed to room air or to SF or IH for 6 weeks. Then, the mice were sacrificed, and atria and ventricles were immediately dissected. The abundances of Cx40 or Cx43 in atria and ventricles were unaffected by SF. In contrast, immunoblots showed that the abundance of atrial Cx40 and Cx43 and ventricular Cx43 were reduced in mice exposed to IH. qRT-PCR demonstrated significant reductions of atrial Cx40 and Cx43 mRNAs. Immunofluorescence microscopy revealed that the abundance and size of gap junctions containing Cx40 or Cx43 were reduced in atria by IH treatment of mice. However, no changes of connexin abundance or gap junction size/abundance were observed in IH-treated NOX2-null mice.ConclusionsThese results demonstrate that intermittent hypoxia (but not sleep fragmentation) causes reductions and remodeling of atrial Cx40 and Cx43. These alterations may contribute to the substrate for atrial fibrillation that develops in response to obstructive sleep apnea. Moreover, these connexin changes are likely generated in response to reactive oxygen species generated by NOX2.

Prolongation of atrio-ventricular node conduction in a rabbit model of ischaemic cardiomyopathy: Role of fibrosis and connexin remodelling

Conduction abnormalities are frequently associated with cardiac disease, though the mechanisms underlying the commonly associated increases in PQ interval are not known. This study uses a chronic left ventricular (LV) apex myocardial infarction (MI) model in the rabbit to create significant left ventricular dysfunction (LVD) 8weeks post-MI. In vivo studies established that the PQ interval increases by approximately 7ms (10%) with no significant change in average heart rate. Optical mapping of isolated Langendorff perfused rabbit hearts recapitulated this result: time to earliest activation of the LV was increased by 14ms (16%) in the LVD group. Intra-atrial and LV transmural conduction times were not altered in the LVD group. Isolated AVN preparations from the LVD group demonstrated a significantly longer conduction time (by approximately 20ms) between atrial and His electrograms than sham controls across a range of pacing cycle lengths. This difference was accompanied by increased effective refractory period and Wenckebach cycle length, suggesting significantly altered AVN electrophysiology post-MI. The AVN origin of abnormality was further highlighted by optical mapping of the isolated AVN. Immunohistochemistry of AVN preparations revealed increased fibrosis and gap junction protein (connexin43 and 40) remodelling in the AVN of LVD animals compared to sham. A significant increase in myocyte–non-myocyte connexin co-localization was also observed after LVD. These changes may increase the electrotonic load experienced by AVN muscle cells and contribute to slowed conduction velocity within the AVN.

Relating specific connexin co-expression ratio to connexon composition and gap junction function

Cardiac connexin 43 (Cx43), Cx40 and Cx45 are co-expressed at distinct ratios in myocytes. This pattern is considered a key factor in regulating the gap junction channels composition, properties and function and remains poorly understood.This work aims to correlate gap junction function with the connexin composition of the channels at accurate ratios Cx43:Cx40 and Cx43:Cx45.Rat liver epithelial cells that endogenously express Cx43 were stably transfected to induce expression of accurate levels of Cx40 or Cx45 that may be present in various areas of the heart (e.g. atria and ventricular conduction system). Induction of Cx40 does not increase the amounts of junctional connexins (Cx43 and Cx40), whereas induction of Cx45 increases the amounts of junctional connexins (Cx43 and Cx45). Interestingly, the non-junctional fraction of Cx43 remains unaffected upon induction of Cx40 and Cx45. Co-immunoprecipitation studies show low level of Cx40/Cx43 heteromerisation and undetectable Cx45/Cx43 heteromerisation. Functional characterisation shows that induction of Cx40 and Cx45 decreases Lucifer Yellow transfer. Electrical coupling is decreased by Cx45 induction, whereas it is decreased at low induction of Cx40 and increased at high induction.These data indicate a fine regulation of the gap junction channel make-up in function of the type and the ratio of co-expressed Cxs that specifically regulates chemical and electrical coupling. This reflects specific gap junction function in regulating impulse propagation in the healthy heart, and a pro-arrhythmic potential of connexin remodelling in the diseased heart.

Atrial overexpression of angiotensin-converting enzyme 2 improves the canine rapid atrial pacing-induced structural and electrical remodeling. Fan, ACE2 improves atrial substrate remodeling.

The purpose of this study was to investigate whether atrial overexpression of angiotensin-converting enzyme 2 (ACE2) by homogeneous transmural atrial gene transfer can reverse atrial remodeling and its mechanisms in a canine atrial-pacing model. Twenty-eight mongrel dogs were randomly divided into four groups: Sham-operated, AF-control, gene therapy with adenovirus-enhanced green fluorescent protein (Ad-EGFP) and gene therapy with Ad-ACE2 (Ad-ACE2) (n = 7 per subgroup). AF was induced in all dogs except the Sham-operated group by rapid atrial pacing at 450 beats/min for 2 weeks. Ad-EGFP and Ad-ACE2 group then received epicardial gene painting. Three weeks after gene transfer, all animals except the Sham group underwent rapid atrial pacing for another 3 weeks and then invasive electrophysiological, histological and molecular studies. The Ad-ACE2 group showed an increased ACE2 and Angiotensin-(1–7) expression, and decreased Angiotensin II expression in comparison with Ad-EGFP and AF-control group. ACE2 overexpression attenuated rapid atrial pacing-induced increase in activated extracellular signal-regulated kinases and mitogen-activated protein kinases (MAPKs) levels, and decrease in MAPK phosphatase 1(MKP-1) level, resulting in attenuation of atrial fibrosis collagen protein markers and transforming growth factor-β1. Additionally, ACE2 overexpression also modulated the tachypacing-induced up-regulation of connexin 40, down-regulation of connexin 43 and Kv4.2, and significantly decreased the inducibility and duration of AF. ACE2 overexpression could shift the renin–angiotensin system balance towards the protective axis, attenuate cardiac fibrosis remodeling associated with up-regulation of MKP-1 and reduction of MAPKs activities, modulate tachypacing-induced ion channels and connexin remodeling, and subsequently reduce the inducibility and duration of AF.Electronic supplementary materialThe online version of this article (doi:10.1007/s00395-015-0499-0) contains supplementary material, which is available to authorized users.

Connexin Remodeling Contributes to Atrial Fibrillation.

Atrial fibrillation significantly contributes to mortality and morbidity through increased risk of stroke, heart failure and myocardial infarction. Investigations of mechanisms responsible for the development and maintenance of atrial fibrillation have highlighted the importance of gap junctional remodeling. Connexins 40 and 43, the major atrial gap junctional proteins, undergo considerable alterations in expression and localization in atrial fibrillation, creating an environment conducive to sustained reentry. Atrial fibrillation is initiated and/or maintained in this reentrant substrate. This review will focus on connexin remodeling in the context of underlying mechanism and possible therapeutic target for atrial fibrillation.

GW24-e3174 A trial overexpression of ACE2 improves the canine rapid atrial pacing-induced structural remodelling

ObjectivesAtrial fibrillation (AF) is associated with significant morbidity and mortality. Substantial evidence suggests that the renin angiotensin system (RAS) has a role in the pathogenesis of AF. Angiotensin-converting enzyme 2...

Functional Characterization of Atrial Electrograms in a Pacing‐Induced Heart Failure Model of Atrial Fibrillation: Importance of Regional Atrial Connexin40 Remodeling

Heart failure (HF) increases the susceptibility to atrial fibrillation (AF) and is associated with altered cardiomyocyte connexin. The regional remodeling of connexin(s) may contribute to the spatiotemporal organization of AF. This study sought to investigate the regional differences in connexin(s) and fibrosis in specific atrial regions and correlate that with the electrogram properties. Biatrial electroanatomic mapping during sinus rhythm (electrogram voltage and velocity) and AF (dominant frequency, DF) was performed in 6 ventricular pacing-induced HF dogs (at 252 beats/minute for 6 weeks) and 6 controls. Atrial tissues were sampled from 7 specific sites for analysis of the connexin and fibrosis. HF caused marked atrial dilatation, and increased the induced AF duration (P < 0.001). Remodeled connexins, including a lower expression and more lateralization of both connexin40 (Cx40) and Cx43 as well as increased regional dispersion of Cx40, in the presence of diffuse enhanced atrial fibrosis, characterized the atrial substrate of the HF dogs (P < 0.01). Regional analysis showed abnormal velocity and low electrogram voltage in the areas with downregulated Cx40 and Cx43 was enhanced in the presence of marked atrial fibrosis (>30% of area, P < 0.01). During AF, lower expression of the Cx40 was associated with higher DF in areas of less and more fibrosis, respectively (R = 0.67 and 0.58, P < 0.01). An altered expression of connexins correlated with the electrogram properties in the existence of diffuse enhanced atrial fibrosis associated with HF. The regional remodeling of Cx40 is likely an important factor in the maintenance of AF in HF.

The efficacy of MSC-HGF in treating pulmonary arterial hypertension (PAH) and connexin remodelling

Abstract

Abstract 19408: Noninvasive Electrocardiographic Imaging Demonstrates Early Structural and Functional Changes Associated with Paroxysmal Atrial Fibrillation in Humans

BACKGROUND: Atrial fibrillation (AF) causes and perpetuates with a remodeling process that involves both functional (ion channel, connexin) and structural (fibrosis) changes to atrial tissue. Presently, there is no noninvasive method to study this process in humans. Noninvasive electrocardiographic imaging (ECGI) could provide insight into remodeling processes associated with early AF. METHODS: Noninvasive ECGI was performed during sinus rhythm on 20 patients with a history of paroxysmal AF undergoing pulmonary vein isolation procedure after antiarrhythmic drug washout period. Eight patients with no history of AF and normal ventricular function were used for controls. For functional analysis, total atrial conduction time (TACT) and left atrial conduction time (LACT) were measured. For structural analysis, left atrial posterior wall (LAPW) voltage was measured. To account for patient-specific torso variability, this value was indexed to the maximal left atrial appendage voltage (indexed LAPW). RESULTS: Compared to controls, patients with AF had longer TACT (100 vs. 79 ms control, p&lt;0.005) and longer LACT (55 vs. 39 ms control, p = 0.034, top left figure). There were no differences in raw LAPW voltage (0.40 vs. 0.47 mV control, p = 0.53), but significant differences existed after adjusting for patient-specific torso variability (0.36 vs. 0.64 mV control, p &lt; 0.020, top right figure). These differences were strongly associated with duration of PAF diagnosis. Subtle functional differences were observed early in AF, while significant functional and structural changes were seen with AF diagnosis of longer duration (5+ years, see bottom figure). CONCLUSIONS: Very early in human AF, functional effects of ion channel and connexin remodeling are evident with delayed global and regional electrical activation. With longer AF, more pronounced delay and measurable decrease in electrogram voltage are likely a combined effect of ion channel adaptation and early fibrosis.

Suppression of Atrial Fibrillation by Over-Expression of Connexin 43 in a Porcine Model

Atrial fibrillation (AF) is associated with atrial conduction disturbances caused by electrical and structural remodeling. We hypothesized that expression of the gap junction protein connexin 43 is reduced during AF, and that atrial Cx43 gene transfer would prevent persistent atrial fibrillation. The first aim of this study was to assess whether atrial fibrillation (AF) is associated with connexin remodeling in a porcine model. Second, a strategy to suppress persistent AF by gene therapy was developed and evaluated in vivo.AF was inducedin domestic pigs via atrial burst pacing, causing a 62.4% reduction of atrial Cx43 protein. Adenoviruses encoding for connexin 43 (Ad-Cx43) or green fluorescent protein (Ad-GFP) were injected into both atria, followed by epicardial electroporation to enhance transgene expression. Ad-Cx43 treated animals did not exhibit persistent AF during the observation period of 14 days. In contrast, control animals developed persistent AF within 7.4 ± 0.5 days. Rapid ventricular heart rates during AF led to deterioration of cardiac function in control pigs but not in animals treated with Ad-Cx43.In conclusion, our results highlight the contribution of connexin 43 to atrial fibrillation and demonstrate the viability of electrophysiological gene therapy for prevention of atrial arrhythmias.

Residual Cx45 and its relationship to Cx43 in murine ventricular myocardium

Gap junction channels in ventricular myocardium are required for electrical and metabolic coupling between cardiac myocytes and for normal cardiac pump function. Although much is known about expression patterns and remodeling of cardiac connexin(Cx)43, little is known about the less abundant Cx45, which is required for embryonic development and viability, is downregulated in adult hearts, and is pathophysiologically upregulated in human end-stage heart failure. We applied quantitative immunoblotting and immunoprecipitation to native myocardial extracts, immunogold electron microscopy to cardiac tissue and membrane sections, electrophysiological recordings to whole hearts, and high-resolution tandem mass spectrometry to Cx45 fusion protein, and developed two new tools, anti-Cx45 antisera and Cre+;Cx45 floxed mice, to facilitate characterization of Cx45 in adult mammalian hearts. We found that Cx45 represents 0.3% of total Cx protein (predominantly 200 fmol Cx43 protein/μg ventricular protein) and colocalizes with Cx43 in native ventricular gap junctions, particularly in the apex and septum. Cre+;Cx45 floxed mice express 85% less Cx45, but do not exhibit overt electrophysiologic abnormalities. Although the basal phosphorylation status of native Cx45 remains unknown, CaMKII phosphorylates 8 Ser/Thr residues in Cx45 in vitro. Thus, although downregulation of Cx45 does not produce notable deficits in electrical conduction in adult, disease-free hearts, Cx45 is a target of the multifunctional kinase CaMKII, and the phosphorylation status of Cx45 and the role of Cx43/Cx45 heteromeric gap junction channels in both normal and diseased hearts merits further investigation.

Gap junctions and conduction of cardiac excitation

c p t t o i Gap junction remodeling and cardiac disease Changes in function and/or remodeling of connexins occur in almost all major arrhythmogenic cardiac disorders. However, since these diseases are often associated with alterations in all major components of cell and tissue function, potentially involving changes in hundreds of genes and proteins, it is often difficult, if not impossible, to delineate a specific arrhythmogenic role for connexin remodeling. In cardiac failure and ischemic heart disease, remodeling of cell-to-cell coupling is likely to play a significant role in arrhythmogenesis. It is generally agreed that cardiac failure leads to a decrease in Cx43 immunosignals and an increase in Cx45 immunosignals in gap junctions, associated with a decrease in cell-to-cell coupling and propagation velocity. Studies associating the pattern and intensity of Cx43 immunosignals in humans with the occurrence of ventricular arrhythmias have suggested that the inhomogeneity of connexin expression in gap junctions rather than its average decrease may represent an arrhythmogenic mechanism in heart failure. In cardiac ischemia, dephosphorylation of onnexins and the associated cell-to-cell uncoupling occur ithin approximately 15 minutes after acute oxygen withrawal, and tissue surviving from myocardial infarction xhibits marked connexin remodeling. The role of connexin remodeling in atrial fibrillation is more disputed. Experimental studies have shown a relatively large variety of changes without delineating a clear picture. However, somatic mutations in both major atrial onnexins, Cx43 and Cx40, have recently been associated ith lone atrial fibrillation. Although these cases seem to e rare, they may help to explain the role of connexin emodeling in atrial propagation. Whether the association etween mutations in atrial connexins and arrhythmogensis reflects a direct effect of changed cell-to-cell coupling r whether additional changes contribute to the disturbed lectrical function remains to be shown.

Connexin 43 gene therapy prevents persistent atrial fibrillation in a porcine model

Atrial fibrillation (AF) is the most common sustained cardiac arrhythmia, and effective treatment of AF still remains an unmet medical need. AF is associated with atrial conduction disturbances caused by electrical and/or structural remodelling. We hypothesized that AF suppresses expression of the gap junction protein connexin (Cx) 43 and that Cx43 gene transfer to both atria would prevent persistent AF. The first aim of this study was to assess whether AF is associated with connexin remodelling in a porcine model. A strategy to suppress persistent AF by gene therapy was then developed and evaluated in vivo. AF was induced in domestic pigs via atrial burst pacing, causing a 62.4% reduction in atrial Cx43 protein. Adenoviruses encoding for Cx43 (AdCx43) or green fluorescent protein (AdGFP) were injected into both atria, followed by epicardial electroporation to enhance transgene expression. Combining direct injection of adenoviruses with electroporation achieved GFP reporter gene expression in ∼50% of atrial cells in vivo. AdCx43-treated animals exhibited a 2.5-fold increase in atrial Cx43 protein content and did not develop persistent AF during the observation period of 14 days. In contrast, control animals developed persistent AF within 7.4 ± 0.5 days. Rapid ventricular heart rates during AF led to deterioration of cardiac function in control pigs but not in pigs treated with AdCx43. Our results highlight the contribution of Cx43 to the pathophysiology of AF and demonstrate the viability of gene therapy for prevention of atrial arrhythmias.