TY - THES A1 - Eppli, Nenad T1 - Untersuchung des Einflusses der ERK1/2-Autophosphorylierung an Threonin 188 auf Mausherzen mittels transgener Mäuse mit ubiquitärer Überexpression von ERK2\(^{T188D}\) T1 - Study on the effects of autophosphorylation of ERK1/2 at threonine 188 on murine hearts by means of transgenic mice ubiquitiously overexpressing ERK2\(^{T188D}\) N2 - Die ERK2Thr188-Autophosphoylierung stellt einen regulatorischen Signalweg dar, der infolge einer hypertrophen Stimulation die kardiale Hypertrophie begünstigt. Eine Hemmung dieser Phosphorylierung in Kardiomyozyten verhindert die Ausbildung der kardialen Hypertrophie ohne Beeinflussung der kardioprotektiven Funktionen von ERK1/2. Demgegenüber führt die dauerhafte Simulation zu einem gain-of-function-Phänotypen mit ausgeprägter Hypertophie, Fibrose und einer reduzierten Herzfunktion. In dieser Arbeit wurde die dauerhafte Simulation ERK2Thr188-Phosphorylierung (T188D) in einem Mausmodell mit ubiquitärer Expression dieser Mutation untersucht. Dabei konnte gezeigt werden, dass sich nach Stimulation durch TAC in diesen Tieren ein etwas stärkerer hypertropher Phänotyp mit vergrößerten Kardiomyozyten, gesteigerter interstitieller Fibrosierung und reduzierter Herzfunktion ausbildet als in Mäusen mit kardiomyozyten-spezifischer Überexpression diese Mutante. In Fibroblasten- und VSMC-Zelllinien wurde eine gesteigerte Proliferation der T188D-überexprimierenden Zellen im Vergleich zu Kontrollen festgestellt. Somit scheint die ERK2Thr188-Phosphorylierung auch in kardialen Nicht-Myozyten einen maladaptiven Einfluss auf das Herz auszuüben. N2 - Autophosphorylation of ERK2 on threonine 188 is a regulatory signaling pathway facilitating cardiac hypertrophy due to a hypertrophic stimulation. The formation of cardiac hypertrophy can be impaired by inhibition of the phosphorylation without interfering with the cardioprotective functions of ERK1/2. In contrast, permanent stimulation of ERK2Thr188 phosphorylation leads to a gain-of-function phenotype with distinct hypertrophy fibrosis and a reduced cardiac function. Here, we examined a murine model with ubiquitous overexpression of ERK2Thr188 phosphorylation (T188D). We point out that mice showed an increased hypertrophic phenotype with augmented cardiomyocytes, enhanced interstitial fibrosis and reduced cardiac function in response to TAC compared to mice with overexpression of this mutant limited to cardiomyocytes. Fibroblasts and vascular smooth muscle cells overexpressing T188D showed an enhanced proliferation compared to controls. Taken together, the phosphorylation of ERK2 on threonine 188 exerts a maladaptive influence on non-myocytes in the heart as well. KW - Herzhypertrophie KW - cardiac hypertrophy KW - ERK1/2-Autophosphorylierung KW - autophosphorylation of ERK1/2 Y1 - 2023 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-216558 ER - TY - JOUR A1 - Sedaghat-Hamedani, Farbod A1 - Rebs, Sabine A1 - Kayvanpour, Elham A1 - Zhu, Chenchen A1 - Amr, Ali A1 - Müller, Marion A1 - Haas, Jan A1 - Wu, Jingyan A1 - Steinmetz, Lars M. A1 - Ehlermann, Philipp A1 - Streckfuss-Bömeke, Katrin A1 - Frey, Norbert A1 - Meder, Benjamin T1 - Genotype complements the phenotype: identification of the pathogenicity of an LMNA splice variant by nanopore long-read sequencing in a large DCM family JF - International Journal of Molecular Sciences N2 - Dilated cardiomyopathy (DCM) is a common cause of heart failure (HF) and is of familial origin in 20–40% of cases. Genetic testing by next-generation sequencing (NGS) has yielded a definite diagnosis in many cases; however, some remain elusive. In this study, we used a combination of NGS, human-induced pluripotent-stem-cell-derived cardiomyocytes (iPSC-CMs) and nanopore long-read sequencing to identify the causal variant in a multi-generational pedigree of DCM. A four-generation family with familial DCM was investigated. Next-generation sequencing (NGS) was performed on 22 family members. Skin biopsies from two affected family members were used to generate iPSCs, which were then differentiated into iPSC-CMs. Short-read RNA sequencing was used for the evaluation of the target gene expression, and long-read RNA nanopore sequencing was used to evaluate the relevance of the splice variants. The pedigree suggested a highly penetrant, autosomal dominant mode of inheritance. The phenotype of the family was suggestive of laminopathy, but previous genetic testing using both Sanger and panel sequencing only yielded conflicting evidence for LMNA p.R644C (rs142000963), which was not fully segregated. By re-sequencing four additional affected family members, further non-coding LMNA variants could be detected: rs149339264, rs199686967, rs201379016, and rs794728589. To explore the roles of these variants, iPSC-CMs were generated. RNA sequencing showed the LMNA expression levels to be significantly lower in the iPSC-CMs of the LMNA variant carriers. We demonstrated a dysregulated sarcomeric structure and altered calcium homeostasis in the iPSC-CMs of the LMNA variant carriers. Using targeted nanopore long-read sequencing, we revealed the biological significance of the variant c.356+1G>A, which generates a novel 5′ splice site in exon 1 of the cardiac isomer of LMNA, causing a nonsense mRNA product with almost complete RNA decay and haploinsufficiency. Using novel molecular analysis and nanopore technology, we demonstrated the pathogenesis of the rs794728589 (c.356+1G>A) splice variant in LMNA. This study highlights the importance of precise diagnostics in the clinical management and workup of cardiomyopathies. KW - familial DCM KW - laminopathy KW - long-read sequencing KW - nanopore KW - induced pluripotent stem cell cardiomyocytes Y1 - 2022 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-290415 SN - 1422-0067 VL - 23 IS - 20 ER - TY - JOUR A1 - Schanbacher, Constanze A1 - Hermanns, Heike M. A1 - Lorenz, Kristina A1 - Wajant, Harald A1 - Lang, Isabell T1 - Complement 1q/tumor necrosis factor-related proteins (CTRPs): structure, receptors and signaling JF - Biomedicines N2 - Adiponectin and the other 15 members of the complement 1q (C1q)/tumor necrosis factor (TNF)-related protein (CTRP) family are secreted proteins composed of an N-terminal variable domain followed by a stalk region and a characteristic C-terminal trimerizing globular C1q (gC1q) domain originally identified in the subunits of the complement protein C1q. We performed a basic PubMed literature search for articles mentioning the various CTRPs or their receptors in the abstract or title. In this narrative review, we briefly summarize the biology of CTRPs and focus then on the structure, receptors and major signaling pathways of CTRPs. Analyses of CTRP knockout mice and CTRP transgenic mice gave overwhelming evidence for the relevance of the anti-inflammatory and insulin-sensitizing effects of CTRPs in autoimmune diseases, obesity, atherosclerosis and cardiac dysfunction. CTRPs form homo- and heterotypic trimers and oligomers which can have different activities. The receptors of some CTRPs are unknown and some receptors are redundantly targeted by several CTRPs. The way in which CTRPs activate their receptors to trigger downstream signaling pathways is largely unknown. CTRPs and their receptors are considered as promising therapeutic targets but their translational usage is still hampered by the limited knowledge of CTRP redundancy and CTRP signal transduction. KW - adiponectin KW - AMPK KW - C1q/TNF related protein (CTRP) KW - inflammation KW - metabolism Y1 - 2023 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-304136 SN - 2227-9059 VL - 11 IS - 2 ER - TY - JOUR A1 - Gmach, Philipp A1 - Bathe-Peters, Marc A1 - Telugu, Narasimha A1 - Miller, Duncan C. A1 - Annibale, Paolo T1 - Fluorescence spectroscopy of low-level endogenous β-adrenergic receptor expression at the plasma membrane of differentiating human iPSC-derived cardiomyocytes JF - International Journal of Molecular Sciences N2 - The potential of human-induced pluripotent stem cells (hiPSCs) to be differentiated into cardiomyocytes (CMs) mimicking adult CMs functional morphology, marker genes and signaling characteristics has been investigated since over a decade. The evolution of the membrane localization of CM-specific G protein-coupled receptors throughout differentiation has received, however, only limited attention to date. We employ here advanced fluorescent spectroscopy, namely linescan Fluorescence Correlation Spectroscopy (FCS), to observe how the plasma membrane abundance of the β\(_1\)- and β\(_2\)-adrenergic receptors (β\(_{1/2}\)-ARs), labelled using a bright and photostable fluorescent antagonist, evolves during the long-term monolayer culture of hiPSC-derived CMs. We compare it to the kinetics of observed mRNA levels in wildtype (WT) hiPSCs and in two CRISPR/Cas9 knock-in clones. We conduct these observations against the backdrop of our recent report of cell-to-cell expression variability, as well as of the subcellular localization heterogeneity of β-ARs in adult CMs. KW - GPCR KW - β-adrenergic receptors KW - hiPSC-CM KW - cardiomyocyte KW - fluorescence correlation spectroscopy KW - FCS KW - fluorescence KW - CRISPR/Cas9 KW - differentiation Y1 - 2022 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-288277 SN - 1422-0067 VL - 23 IS - 18 ER - TY - JOUR A1 - Djelić, Ninoslav A1 - Borozan, Sunčica A1 - Dimitrijević-Srećković, Vesna A1 - Pajović, Nevena A1 - Mirilović, Milorad A1 - Stopper, Helga A1 - Stanimirović, Zoran T1 - Oxidative stress and DNA damage in peripheral blood mononuclear cells from normal, obese, prediabetic and diabetic persons exposed to thyroid hormone in vitro JF - International Journal of Molecular Sciences N2 - Diabetes, a chronic group of medical disorders characterized byhyperglycemia, has become a global pandemic. Some hormones may influence the course and outcome of diabetes, especially if they potentiate the formation of reactive oxygen species (ROS). There is a close relationship between thyroid disorders and diabetes. The main objective of this investigation was to find out whether peripheral blood mononuclear cells (PBMCs) are more prone to DNA damage by triiodothyronine (T\(_3\)) (0.1, 1 and 10 μM) at various stages of progression through diabetes (obese, prediabetics, and type 2 diabetes mellitus—T2DM persons). In addition, some biochemical parameters of oxidative stress (catalase-CAT, thiobarbituric acid reactive substances—TBARS) and lactate dehydrogenase (LDH) were evaluated. PBMCs from prediabetic and diabetic patients exhibited increased sensitivity for T\(_3\) regarding elevated level of DNA damage, inhibition of catalase, and increase of TBARS and LDH. PBMCs from obese patients reacted in the same manner, except for DNA damage. The results of this study should contribute to a better understanding of the role of thyroid hormones in the progression of T2DM. KW - diabetes KW - oxidative stress KW - DNA damage KW - lymphocytes KW - thyroid hormone Y1 - 2022 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-285988 SN - 1422-0067 VL - 23 IS - 16 ER - TY - THES A1 - Nemec, Katarina T1 - Modulation of parathyroid hormone 1 receptor (PTH1R) signaling by receptor activity-modifying proteins (RAMPs) T1 - Regulierung der Signalübertragung des Parathormon 1-Rezeptors (PTH1R) durch Rezeptoraktivitäts-modifizierende Proteine (RAMPs) N2 - The receptor activity-modifying proteins (RAMPs) are ubiquitously expressed membrane proteins that interact with several G protein-coupled receptors (GPCRs), the largest and pharmacologically most important family of cell surface receptors. RAMPs can regulate GPCR function in terms of ligand-binding, G-protein coupling, downstream signaling, trafficking, and recycling. The integrity of their interactions translates to many physiological functions or pathological conditions. Regardless of numerous reports on its essential importance for cell biology and pivotal role in (patho-)physiology, the molecular mechanism of how RAMPs modulate GPCR activation remained largely elusive. This work presents new insights that add to the common understanding of the allosteric regulation of receptor activation and will help interpret how accessory proteins - RAMPs - modulate activation dynamics and how this affects the fundamental aspects of cellular signaling. Using a prototypical class B GPCR, the parathyroid hormone 1 receptor (PTH1R) in the form of advanced genetically encoded optical biosensors, I examined RAMP's impact on the PTH1R activation and signaling in intact cells. A panel of single-cell FRET and confocal microscopy experiments as well canonical and non-canonical functional assays were performed to get a holistic picture of the signaling initiation and transduction of that clinically and therapeutically relevant GPCR. Finally, structural modeling was performed to add molecular mechanistic details to that novel art of modulation. I describe here that RAMP2 acts as a specific allosteric modulator of PTH1R, shifting PTH1R to a unique pre-activated state that permits faster activation in a ligand-specific manner. Moreover, RAMP2 modulates PTH1R downstream signaling in an agonist-dependent manner, most notably increasing the PTH-mediated Gi3 signaling sensitivity and kinetics of cAMP accumulation. Additionally, RAMP2 increases PTH- and PTHrP-triggered β-arrestin2 recruitment to PTH1R and modulates cytosolic ERK1/2 phosphorylation. Structural homology modeling shows that structural motifs governing GPCR-RAMP interaction originate in allosteric hotspots and rationalize functional modulation. Moreover, to interpret the broader role of RAMP's modulation in GPCRs pharmacology, different fluorescent tools to investigate RAMP's spatial organization were developed, and novel conformational biosensors for class B GPCRs were engineered. Lastly, a high throughput assay is proposed and prototyped to expand the repertoire of RAMPs or other membrane protein interactors. These data uncover the critical role of RAMPs in GPCR activation and signaling and set up a novel platform for studying GPCR modulation. Furthermore, these insights may provide a new venue for precise modulation of GPCR function and advanced drug design. N2 - G Protein-gekoppelte Rezeptoren (GPCRs) bilden die größte und pharmakologisch wichtigste Familie von Zelloberflächenrezeptoren, die zahlreiche (patho-)physiologische Prozesse im menschlichen Körper steuern. GPCRs übertragen während des Rezeptoraktivierungsprozesses extrazelluläre Signale in das Zellinnere, wo durch die extrazelluläre Stimulation Konformationsänderungen des Rezeptorkerns auslöst und die Bindung intrazellulärer Bindungspartner – G Proteine, G Protein-gekoppelte Rezeptorkinase und Arrestine - ermöglicht. Es handelt sich also um einen kritischen Prozess in der Signaltransduktion, der durch einige endogene Moleküle wie Ionen, Lipide oder andere Proteine moduliert werden kann und Auswirkungen auf nachgeschaltete Signalkaskaden hat. GPCRs bilden gewebeabhängige Oligomere mit ihren interagierenden Partnern, Rezeptor-Aktivitäts-modifizierende Proteinen (RAMPs), ubiquitär exprimierten Membranproteinen. Bekannt ist, dass sie die Ligandenbindung, die G- Protein-Kopplung, die nachgeschaltete Signalisierung, das Trafficking und das Recycling einiger GPCRs modulieren. Ihre Rolle im kritischsten Prozess der Signaltransduktion - der Rezeptoraktivierung - wurde jedoch nur begrenzt erforscht. Anhand des physiologisch und therapeutisch wichtigen Parathormon-Rezeptors (PTH1R), einem GPCR der Klasse B, wurden die Modulationseffekte von RAMPs auf den Prozess der Rezeptoraktivierung und ihre Folgen für die nachgeschaltete Signalübertragung analysiert. Hierzu wurden verschiedene optische Biosensoren zur Messung der Aktivierung des PTH1R und seiner Signalkaskade entwickelt und in verschiedenen Versuchsanordnungen eingesetzt, mit dem Ziel einen holistischen Blick auf die Interaktion zwischen PTH1R und RAMPs und ihre funktionellen Auswirkungen zu erhalten. Die Interaktion zwischen PTH1R und RAMPs erwies sich als besonders ausgeprägt für RAMP2, und RAMP2 zeigte eine spezifische allosterische Modulation der PTH1R-Konformation, sowohl im basalen als auch im Liganden- aktivierten Zustand. Ein einzigartiger voraktivierter oder (meta-stabiler) Zustand ermöglichte eine schnellere Rezeptoraktivierung auf Liganden-spezifische Weise. Außerdem beeinflusste RAMP2 die G Protein- und Nicht-G Protein-vermittelte Signalübertragung indem es die PTH-vermittelte Gi3-Signalempfindlichkeit und die Kinetik der cAMP-Akkumulation modulierte. Weiterhin erhöhte RAMP2 die Menge der β-Arrestin2-Rekrutierung an PTH1R auf Liganden-spezifische Weise. Dies könnte mit einer erhöhten zytosolischen ERK-Menge zusammenhängen, die hat sich von der nukleären ERK-Phosphorylierung unterscheidet. Um einen molekularen Mechanismus für die vorgestellten Ergebnisse vorzuschlagen, wurden mehrere strukturelle Modelle entwickelt und analysiert. Diese Arbeit liefert den Beweis, dass RAMP die GPCR-Aktivierung mit funktionellen Auswirkungen auf die zelluläre Signalübertragung reguliert. Die Ergebnisse sollten im Zusammenhang mit zellspezifischen Koexpressionsmustern interpretiert werden und können zur Entwicklung von fortschrittlichen Therapeutika positiv beitragen. Da GPCRs praktisch alle Zellfunktionen koordinieren und seit jeher wichtigen Angriffspunkten für Medikamente sind, tragen die vorgestellten Erkenntnisse zum universellen Verständnis der molekularen Mechanismen bei, die den menschlichen Körper orchestrieren. KW - G-Protein gekoppelter Rezeptor KW - GPCR KW - RAMP KW - PTH1R KW - FRET KW - BRET KW - pharmacology KW - Fluoreszenz-Resonanz-Energie-Transfer KW - Förster Resonanz Energie Transfer Y1 - 2023 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-288588 ER - TY - JOUR A1 - Schanbacher, Constanze A1 - Bieber, Michael A1 - Reinders, Yvonne A1 - Cherpokova, Deya A1 - Teichert, Christina A1 - Nieswandt, Bernhard A1 - Sickmann, Albert A1 - Kleinschnitz, Christoph A1 - Langhauser, Friederike A1 - Lorenz, Kristina T1 - ERK1/2 activity is critical for the outcome of ischemic stroke JF - International Journal of Molecular Sciences N2 - Ischemic disorders are the leading cause of death worldwide. The extracellular signal-regulated kinases 1 and 2 (ERK1/2) are thought to affect the outcome of ischemic stroke. However, it is under debate whether activation or inhibition of ERK1/2 is beneficial. In this study, we report that the ubiquitous overexpression of wild-type ERK2 in mice (ERK2\(^{wt}\)) is detrimental after transient occlusion of the middle cerebral artery (tMCAO), as it led to a massive increase in infarct volume and neurological deficits by increasing blood–brain barrier (BBB) leakiness, inflammation, and the number of apoptotic neurons. To compare ERK1/2 activation and inhibition side-by-side, we also used mice with ubiquitous overexpression of the Raf-kinase inhibitor protein (RKIP\(^{wt}\)) and its phosphorylation-deficient mutant RKIP\(^{S153A}\), known inhibitors of the ERK1/2 signaling cascade. RKIP\(^{wt}\) and RKIP\(^{S153A}\) attenuated ischemia-induced damages, in particular via anti-inflammatory signaling. Taken together, our data suggest that stimulation of the Raf/MEK/ERK1/2-cascade is severely detrimental and its inhibition is rather protective. Thus, a tight control of the ERK1/2 signaling is essential for the outcome in response to ischemic stroke. KW - ERK1/2 KW - tMCAO KW - ischemic stroke KW - RKIP Y1 - 2022 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-283991 SN - 1422-0067 VL - 23 IS - 2 ER - TY - JOUR A1 - Bauer, Benedikt A1 - Mally, Angela A1 - Liedtke, Daniel T1 - Zebrafish embryos and larvae as alternative animal models for toxicity testing JF - International Journal of Molecular Sciences N2 - Prerequisite to any biological laboratory assay employing living animals is consideration about its necessity, feasibility, ethics and the potential harm caused during an experiment. The imperative of these thoughts has led to the formulation of the 3R-principle, which today is a pivotal scientific standard of animal experimentation worldwide. The rising amount of laboratory investigations utilizing living animals throughout the last decades, either for regulatory concerns or for basic science, demands the development of alternative methods in accordance with 3R to help reduce experiments in mammals. This demand has resulted in investigation of additional vertebrate species displaying favourable biological properties. One prominent species among these is the zebrafish (Danio rerio), as these small laboratory ray-finned fish are well established in science today and feature outstanding biological characteristics. In this review, we highlight the advantages and general prerequisites of zebrafish embryos and larvae before free-feeding stages for toxicological testing, with a particular focus on cardio-, neuro, hepato- and nephrotoxicity. Furthermore, we discuss toxicokinetics, current advances in utilizing zebrafish for organ toxicity testing and highlight how advanced laboratory methods (such as automation, advanced imaging and genetic techniques) can refine future toxicological studies in this species. KW - danio rerio KW - alternative methods KW - organ toxicity KW - 3R KW - transgenic animals Y1 - 2021 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-284225 SN - 1422-0067 VL - 22 IS - 24 ER - TY - JOUR A1 - Sedaghat-Hamedani, Farbod A1 - Rebs, Sabine A1 - El-Battrawy, Ibrahim A1 - Chasan, Safak A1 - Krause, Tobias A1 - Haas, Jan A1 - Zhong, Rujia A1 - Liao, Zhenxing A1 - Xu, Qiang A1 - Zhou, Xiaobo A1 - Akin, Ibrahim A1 - Zitron, Edgar A1 - Frey, Norbert A1 - Streckfuss-Bömeke, Katrin A1 - Kayvanpour, Elham T1 - Identification of SCN5a p.C335R variant in a large family with dilated cardiomyopathy and conduction disease JF - International Journal of Molecular Sciences N2 - Introduction: Familial dilated cardiomyopathy (DCM) is clinically variable and has been associated with mutations in more than 50 genes. Rapid improvements in DNA sequencing have led to the identification of diverse rare variants with unknown significance (VUS), which underlines the importance of functional analyses. In this study, by investigating human-induced pluripotent stem cell-derived cardiomyocytes (iPSC-CMs), we evaluated the pathogenicity of the p.C335R sodium voltage-gated channel alpha subunit 5 (SCN5a) variant in a large family with familial DCM and conduction disease. Methods: A four-generation family with autosomal dominant familial DCM was investigated. Next-generation sequencing (NGS) was performed in all 16 family members. Clinical deep phenotyping, including endomyocardial biopsy, was performed. Skin biopsies from two patients and one healthy family member were used to generate human-induced pluripotent stem cells (iPSCs), which were then differentiated into cardiomyocytes. Patch-clamp analysis with Xenopus oocytes and iPSC-CMs were performed. Results: A SCN5a variant (c.1003T>C; p.C335R) could be detected in all family members with DCM or conduction disease. A novel truncating TTN variant (p.Ser24998LysfsTer28) could also be identified in two family members with DCM. Family members with the SCN5a variant (p.C335R) showed significantly longer PQ and QRS intervals and lower left ventricular ejection fractions (LV-EF). All four patients who received CRT-D were non-responders. Electrophysiological analysis with Xenopus oocytes showed a loss of function in SCN5a p.C335R. Na\(^+\) channel currents were also reduced in iPSC-CMs from DCM patients. Furthermore, iPSC-CM with compound heterozygosity (SCN5a p.C335R and TTNtv) showed significant dysregulation of sarcomere structures, which may be contributed to the severity of the disease and earlier onset of DCM. Conclusion: The SCN5a p.C335R variant is causing a loss of function of peak INa in patients with DCM and cardiac conduction disease. The co-existence of genetic variants in channels and structural genes (e.g., SCN5a p.C335R and TTNtv) increases the severity of the DCM phenotype. KW - familial DCM KW - conduction disease KW - SCN5a Y1 - 2021 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-284442 SN - 1422-0067 VL - 22 IS - 23 ER - TY - JOUR A1 - Wagner, Michael A1 - Sadek, Mirna S. A1 - Dybkova, Nataliya A1 - Mason, Fleur E. A1 - Klehr, Johann A1 - Firneburg, Rebecca A1 - Cachorro, Eleder A1 - Richter, Kurt A1 - Klapproth, Erik A1 - Kuenzel, Stephan R. A1 - Lorenz, Kristina A1 - Heijman, Jordi A1 - Dobrev, Dobromir A1 - El-Armouche, Ali A1 - Sossalla, Samuel A1 - Kämmerer, Susanne T1 - Cellular mechanisms of the anti-arrhythmic effect of cardiac PDE2 overexpression JF - International Journal of Molecular Sciences N2 - Background: Phosphodiesterases (PDE) critically regulate myocardial cAMP and cGMP levels. PDE2 is stimulated by cGMP to hydrolyze cAMP, mediating a negative crosstalk between both pathways. PDE2 upregulation in heart failure contributes to desensitization to β-adrenergic overstimulation. After isoprenaline (ISO) injections, PDE2 overexpressing mice (PDE2 OE) were protected against ventricular arrhythmia. Here, we investigate the mechanisms underlying the effects of PDE2 OE on susceptibility to arrhythmias. Methods: Cellular arrhythmia, ion currents, and Ca\(^{2+}\)-sparks were assessed in ventricular cardiomyocytes from PDE2 OE and WT littermates. Results: Under basal conditions, action potential (AP) morphology were similar in PDE2 OE and WT. ISO stimulation significantly increased the incidence of afterdepolarizations and spontaneous APs in WT, which was markedly reduced in PDE2 OE. The ISO-induced increase in I\(_{CaL}\) seen in WT was prevented in PDE2 OE. Moreover, the ISO-induced, Epac- and CaMKII-dependent increase in I\(_{NaL}\) and Ca\(^{2+}\)-spark frequency was blunted in PDE2 OE, while the effect of direct Epac activation was similar in both groups. Finally, PDE2 inhibition facilitated arrhythmic events in ex vivo perfused WT hearts after reperfusion injury. Conclusion: Higher PDE2 abundance protects against ISO-induced cardiac arrhythmia by preventing the Epac- and CaMKII-mediated increases of cellular triggers. Thus, activating myocardial PDE2 may represent a novel intracellular anti-arrhythmic therapeutic strategy in HF. KW - PDE2 KW - arrhythmia KW - CaMKII KW - heart failure Y1 - 2021 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-285888 SN - 1422-0067 VL - 22 IS - 9 ER -