Increased [18F]FDG uptake in the infarcted myocardial area displayed by combined PET/CMR correlates with snRNA-seq-detected inflammatory cell invasion.

Combined [18F]FDG PET-cardiac MRI imaging (PET/CMR) is a useful tool to assess myocardial viability and cardiac function in patients with acute myocardial infarction (AMI). Here, we evaluated the prognostic value of PET/CMR in a porcine closed-chest reperfused AMI (rAMI) model. Late gadolinium enhancement by PET/CMR imaging displayed tracer uptake defect at the infarction site by 3 days after the rAMI in the majority of the animals (group Match, n = 28). Increased [18F]FDG uptake at the infarcted area (metabolism/contractility mismatch) with reduced tracer uptake in the remote viable myocardium (group Mismatch, n = 12) 3 days after rAMI was observed in the animals with larger infarct size and worse left ventricular ejection fraction (LVEF) (34 ± 8.7 vs 42.0 ± 5.2%), with lower LVEF also at the 1-month follow-up (35.8 ± 9.5 vs 43.0 ± 6.3%). Transcriptome analyses by bulk and single-nuclei RNA sequencing of the infarcted myocardium and border zones (n = 3 of each group, and 3 sham-operated controls) revealed a strong inflammatory response with infiltration of monocytes and macrophages in the infarcted and border areas in Mismatch animals. Our data indicate a high prognostic relevance of combined PET/MRI in the subacute phase of rAMI for subsequent impairment of heart function and underline the adverse effects of an excessive activation of the innate immune system in the initial phase after rAMI.

Plasma Small Extracellular Vesicle Cardiac miRNA Expression in Patients with Ischemic Heart Failure, Randomized to Percutaneous Intramyocardial Treatment of Adipose Derived Stem Cells or Placebo: Subanalysis of the SCIENCE Study.

Small extracellular vesicles (EVs) and their cargo are an important component of cell-to-cell communication in cardiac disease. Allogeneic adipose derived stem cells (ADSCs) are thought to be a potential approach for cardiac regenerative therapy in ischemic heart disease. The SCIENCE study investigated the effect of ADSCs administered via intramyocardial injection on cardiac function in patients with ischemic heart disease. The aim of this substudy, based on samples from 15 patients, was to explore small EV miRNA dynamics after treatment with ADSCs compared to a placebo. Small EVs were isolated at several timepoints after the percutaneous intramyocardial application of ADSCs. No significant effect of ADSC treatment on small EV concentration was detected. After 12 months, the expression of miR-126 decreased significantly in ADSC patients, but not in the placebo-treated group. However, all cardiac miRNAs correlated with plasma cardiac biomarkers. In line with the overall negative results of the SCIENCE study, with the exception of one miR, we did not detect any significant regulation of small EV miRNAs in this patient collective.

Identification of Gene Expression Signatures for Phenotype-Specific Drug Targeting of Cardiac Fibrosis.

We have designed translational animal models to investigate cardiac profibrotic gene signatures. Domestic pigs were treated with cardiotoxic drugs (doxorubicin, DOX, n = 5 or Myocet®, MYO, n = 5) to induce replacement fibrosis via cardiotoxicity. Reactive interstitial fibrosis was triggered by LV pressure overload by artificial isthmus stenosis with stepwise developing myocardial hypertrophy and final fibrosis (Hyper, n = 3) or by LV volume overload in the adverse remodeled LV after myocardial infarction (RemoLV, n = 3). Sham interventions served as controls and healthy animals (Control, n = 3) served as a reference in sequencing study. Myocardial samples from the LV of each group were subjected to RNA sequencing. RNA-seq analysis revealed a clear distinction between the transcriptomes of myocardial fibrosis (MF) models. Cardiotoxic drugs activated the TNF-alpha and adrenergic signaling pathways. Pressure or volume overload led to the activation of FoxO pathway. Significant upregulation of pathway components enabled the identification of potential drug candidates used for the treatment of heart failure, such as ACE inhibitors, ARB, ß-blockers, statins and diuretics specific to the distinct MF models. We identified candidate drugs in the groups of channel blockers, thiostrepton that targets the FOXM1-regulated ACE conversion to ACE2, tyrosine kinases or peroxisome proliferator-activated receptor inhibitors. Our study identified different gene targets involved in the development of distinct preclinical MF protocols enabling tailoring expression signature-based approach for the treatment of MF.

CDR132L improves systolic and diastolic function in a large animal model of chronic heart failure.

AIMS: Cardiac miR-132 activation leads to adverse remodelling and pathological hypertrophy. CDR132L is a synthetic lead-optimized oligonucleotide inhibitor with proven preclinical efficacy and safety in heart failure (HF) early after myocardial infarction (MI), and recently completed clinical evaluation in a Phase 1b study (NCT04045405). The aim of the current study was to assess safety and efficacy of CDR132L in a clinically relevant large animal (pig) model of chronic heart failure following MI. METHODS AND RESULTS: In a chronic model of post-MI HF, slow-growing pigs underwent 90 min left anterior descending artery occlusion followed by reperfusion. Animals were randomized and treatment started 1-month post-MI. Monthly intravenous (IV) treatments of CDR132L over 3 or 5 months (3× or 5×) were applied in a blinded randomized placebo-controlled fashion. Efficacy was evaluated based on serial magnetic resonance imaging, haemodynamic, and biomarker analyses. The treatment regime provided sufficient tissue exposure and CDR132L was well tolerated. Overall, CDR132L treatment significantly improved cardiac function and reversed cardiac remodelling. In addition to the systolic recovery, diastolic function was also ameliorated in this chronic model of HF. CONCLUSION: Monthly repeated dosing of CDR132L is safe and adequate to provide clinically relevant exposure and therapeutic efficacy in a model of chronic post-MI HF. CDR132L thus should be explored as treatment for the broad area of chronic heart failure.

Secondary mitral regurgitation-Insights from microRNA assessment.

BACKGROUND: While secondary mitral regurgitation (sMR) is associated with adverse outcome in heart failure with reduced ejection fraction (HFrEF), key pathophysiologic mechanisms remain poorly understood and might be elucidated by microRNAs (miRNA/miR), that were recently related to cardiac remodelling. This study sought to assess (i) the differences of miRNA profiles in patients with severe sMR compared to matched disease controls, (ii) the correlation between circulating miRNAs and surrogates of sMR severity as well as (iii) the prognostic implications of miRNA levels in severe sMR. MATERIALS AND METHODS: Sixty-six HFrEF patients were included, of these 44 patients with severe sMR 2:1 matched to HFrEF controls with no/mild sMR. A comprehensive set of miRNAs (miR-21, miR-29a, miR-122, miR-132, miR-133a, miR-let7i) were measured and correlated to echocardiographic sMR severity. RESULTS: miRNA patterns differed distinctly between patients with severe sMR and HFrEF controls (P < .05). Among the panel of assessed miRNAs, miR-133a correlated most strongly with surrogates of sMR severity (r = -0.41, P = .001 with sMR vena contracta width). Interestingly, elevated levels of miR-133 were associated with an increased risk for cardiovascular death and/or HF hospitalizations with and adjusted HR of 1.85 (95% CI 1.24-2.76, P = .003). CONCLUSIONS: This study unveils distinct pathophysiologic maladaptions at a cellular level in patients with severe sMR compared to no/mild sMR by showing significant differences in miRNA profiles and correlations with sMR severity, supporting the concept that sMR drives cardiac remodelling in heart failure. Moreover, the increased risk for adverse outcome in HFrEF patients with severe sMR conveyed by miR-133a might indicate irreversible myocardial damage.

Novel Identified Circular Transcript of RCAN2, circ-RCAN2, Shows Deviated Expression Pattern in Pig Reperfused Infarcted Myocardium and Hypoxic Porcine Cardiac Progenitor Cells In Vitro.

Circular RNAs (circRNAs) are crucial in gene regulatory networks and disease development, yet circRNA expression in myocardial infarction (MI) is poorly understood. Here, we harvested myocardium samples from domestic pigs 3 days after closed-chest reperfused MI or sham surgery. Cardiac circRNAs were identified by RNA-sequencing of rRNA-depleted RNA from infarcted and healthy myocardium tissue samples. Bioinformatics analysis was performed using the CIRIfull and KNIFE algorithms, and circRNAs identified with both algorithms were subjected to differential expression (DE) analysis and validation by qPCR. Circ-RCAN2 and circ-C12orf29 expressions were significantly downregulated in infarcted tissue compared to healthy pig heart. Sanger sequencing was performed to identify the backsplice junctions of circular transcripts. Finally, we compared the expressions of circ-C12orf29 and circ-RCAN2 between porcine cardiac progenitor cells (pCPCs) that were incubated in a hypoxia chamber for different time periods versus normoxic pCPCs. Circ-C12orf29 did not show significant DE in vitro, whereas circ-RCAN2 exhibited significant ischemia-time-dependent upregulation in hypoxic pCPCs. Overall, our results revealed novel cardiac circRNAs with DE patterns in pCPCs, and in infarcted and healthy myocardium. Circ-RCAN2 exhibited differential regulation by myocardial infarction in vivo and by hypoxia in vitro. These results will improve our understanding of circRNA regulation during acute MI.

Early Elevation of Systemic Plasma Clusterin after Reperfused Acute Myocardial Infarction in a Preclinical Porcine Model of Ischemic Heart Disease.

Clusterin exerts anti-inflammatory, cytoprotective and anti-apoptotic effects. Both an increase and decrease of clusterin in acute myocardial infarction (AMI) has been reported. We aimed to clarify the role of clusterin as a systemic biomarker in AMI. AMI was induced by percutaneous left anterior artery (LAD) occlusion for 90 min followed by reperfusion in 24 pigs. Contrast ventriculography was performed after reperfusion to assess left ventricular ejection fraction (LVEF), left ventricular end diastolic volume (LVEDV) and left ventricular end systolic volume (LVESV) and additional cMRI + late enhancement to measure infarct size and LV functions at day 3 and week 6 post-MI. Blood samples were collected at prespecified timepoints. Plasma clusterin and other biomarkers (cTnT, NT-proBNP, neprilysin, NGAL, ET-1, osteopontin, miR21, miR29) were measured by ELISA and qPCR. Gene expression profiles of infarcted and remote region 3 h (n = 5) and 3 days (n = 5) after AMI onset were analysed by RNA-sequencing. AMI led to an increase in LVEDV and LVESV during 6-week, with concomitant elevation of NT-proBNP 3-weeks after AMI. Plasma clusterin levels were increased immediately after AMI and returned to normal levels until 3-weeks. Plasma NGAL, ET-1 and miR29 was significantly elevated at 3 weeks follow-up, miR21 increased after reperfusion and at 3 weeks post-AMI, while circulating neprilysin levels did not change. Elevated plasma clusterin levels 120 min after AMI onset suggest that clusterin might be an additional early biomarker of myocardial ischemia.

Effect of Ischemic Preconditioning and Postconditioning on Exosome-Rich Fraction microRNA Levels, in Relation with Electrophysiological Parameters and Ventricular Arrhythmia in Experimental Closed-Chest Reperfused Myocardial Infarction.

We investigated the antiarrhythmic effects of ischemic preconditioning (IPC) and postconditioning (PostC) by intracardiac electrocardiogram (ECG) and measured circulating microRNAs (miRs) that are related to cardiac conduction. Domestic pigs underwent 90-min. percutaneous occlusion of the mid left anterior coronary artery, followed by reperfusion. The animals were divided into three groups: acute myocardial infarction (AMI, n = 7), ischemic preconditioning-acute myocardial infarction (IPC-AMI) (n = 9), or AMI-PostC (n = 5). IPC was induced by three 5-min. episodes of repetitive ischemia/reperfusion cycles (rI/R) before AMI. PostC was induced by six 30-s rI/R immediately after induction of reperfusion 90 min after occlusion. Before the angiographic procedure, a NOGA endocardial mapping catheter was placed again the distal anterior ventricular endocardium to record the intracardiac electrogram (R-amplitude, ST-Elevation, ST-area under the curve (AUC), QRS width, and corrected QT time (QTc)) during the entire procedure. An arrhythmia score was calculated. Cardiac MRI was performed after one-month. IPC led to significantly lower ST-elevation, heart rate, and arrhythmia score during ischemia. PostC induced a rapid recovery of R-amplitude, decrease in QTc, and lower arrhythmia score during reperfusion. Slightly higher levels of miR-26 and miR-133 were observed in AMI compared to groups IPC-AMI and AMI-PostC. Significantly lower levels of miR-1, miR-208, and miR-328 were measured in the AMI-PostC group as compared to animals in group AMI and IPC-AMI. The arrhythmia score was not significantly associated with miRNA plasma levels. Cardiac MRI showed significantly smaller infarct size in the IPC-AMI group when compared to the AMI and AMI-PostC groups. Thus, IPC led to better left ventricular ejection fraction at one-month and it exerted antiarrhythmic effects during ischemia, whereas PostC exhibited antiarrhythmic properties after reperfusion, with significant downregulaton of ischemia-related miRNAs.

Transcriptional Alterations by Ischaemic Postconditioning in a Pig Infarction Model: Impact on Microvascular Protection.

Although the application of cardioprotective ischaemia/reperfusion (I/R) stimuli after myocardial infarction (MI) is a promising concept for salvaging the myocardium, translation to a clinical scenario has not fulfilled expectations. We have previously shown that in pigs, ischaemic postconditioning (IPostC) reduces myocardial oedema and microvascular obstruction (MVO), without influencing myocardial infarct size. In the present study, we analyzed the mechanisms underlying the IPostC-induced microvascular protection by transcriptomic analysis, followed by pathway analysis. Closed-chest reperfused MI was induced by 90 min percutaneous balloon occlusion of the left anterior descending coronary artery, followed by balloon deflation in anaesthetised pigs. Animals were randomised to IPostC (n = 8), MI (non-conditioned, n = 8), or Control (sham-operated, n = 4) groups. After three hours or three days follow-up, myocardial tissue samples were harvested and subjected to RNA-seq analysis. Although the transcriptome analysis revealed similar expression between IPostC and MI in transcripts involved in cardioprotective pathways, we identified gene expression changes responding to IPostC at the three days follow-up. Focal adhesion signaling, downregulated genes participating in cardiomyopathy and activation of blood cells may have critical consequences for microvascular protection. Specific analyses of the gene subsets enriched in the endothelium of the infarcted area, revealed strong deregulation of transcriptional functional clusters, DNA processing, replication and repair, cell proliferation, and focal adhesion, suggesting sustentative function in the endothelial cell layer protection and integrity. The spatial and time-dependent transcriptome analysis of porcine myocardium supports a protective effect of IPostC on coronary microvasculature post-MI.

Porcine model of progressive cardiac hypertrophy and fibrosis with secondary postcapillary pulmonary hypertension.

BACKGROUND: Meaningful translational large animal models for cardiac diseases are indispensable for studying disease mechanisms, development of novel therapeutic strategies, and evaluation of potential drugs. METHODS: For induction of heart failure, cardiac hypertrophy and fibrosis, a bare metal stent was implanted in the descending aorta of growing pigs (n = 7), inducing pressure stress on the left ventricle (group HYPI). The constant stent size in growing pigs resulted in antegrade partial obstruction of the aortic flow with a gradual increase in afterload. Five pigs with sham intervention served as control. Serial haemodynamic, pressure-volume loop measurements and transthoracic echocardiography (TTE) were performed to detect developing pressure overload of the LV and cardiac MRI with late enhancement for measuring LV and RV mass and ejection fraction. RESULTS: At 5-month follow-up, CT and contrast aortography, and intraluminal echocardiography confirmed aortic isthmus stenosis with a mean trans-stenotic gradient of 64 ± 13.9 mmHg. Invasive haemodynamic measurements revealed a secondary increase in pulmonary artery pressure (44.6 ± 5.1 vs 25.9 ± 6.2 mmHg, HYPI vs control, p < 0.05). TTE and ex vivo analyses confirmed severe concentric LV hypertrophy (mean circumferential wall thickness, 19.4 ± 3.1, n = 7 vs 11.4 ± 1.0 mm, n = 5, HYPI vs controls, p < 0.05). The LV and RV mass increased significantly, paralleled by increased isovolumic relaxation constant (tau). Histological analyses confirmed substantial fibrosis and myocyte hypertrophy in both LV and RV. Expressions of ANP, BNP, and miRNA-29a were up-regulated, while SERCA2a and miRNA-1 were down-regulated. Plasma NGAL levels increased gradually, while the elevation of NT-proBNP was detected only at the 5-month FUP. CONCLUSION: These data prove that percutaneous artificial aortic stenosis in pigs is useful for inducing clinically relevant progredient heart failure based on myocardial hypertrophy and fibrosis.

Coating of intravascular balloon with paclitaxel prevents constrictive remodeling of the dilated porcine femoral artery due to inhibition of intimal and media fibrosis.

Here we investigated how a coating of intravascular balloon with paclitaxel (drug-coated balloon; DCB, Freeway™) impacted porcine peripheral artery vascular function and remodeling. Domestic swine (n = 54) underwent percutaneous overstretch balloon dilation of femoral and iliac arteries, controlled by angiography and optical coherence tomography (OCT). Paclitaxel tissue uptake was measured at 1 h and 1, 3, and 9 days post-dilation. At these time-points and at 32 ± 2 days, vascular function of the dilated arteries was assessed using the organ chamber model. Neointimal growth and remodeling indices were determined using OCT and histology at 32 ± 2 days. Intima and media fibrosis were quantified by picrosirius red staining. Post-inflation femoral artery tissue drug levels were 460 ± 214, 136 ± 123, 14 ± 6, and 0.1 ± 0.1 ng/mg at 1 h and 1, 3, and 9 days, respectively. Compared to plain balloon, Freeway™ resulted in a significantly smaller neointimal area (P < 0.05), less tunica intima (8.0 ± 5.4 vs 14.2 ± 4.7 %) and media fibrosis (15.6 ± 7.7 vs 24.5 ± 5.4 %), and less femoral artery constrictive remodeling (remodeling index: 1.08 ± 0.08 vs 0.94 ± 0.08). The DCB was associated with significantly increased vasoconstrictor tone and endothelium-dependent vasodilation impairment shortly after post-overstretch injury. Overall, DCB dilation of peripheral arteries resulted in high drug uptake into arterial tissue. Compared with the plain balloon, the DCB was associated with decreased vessel wall fibrosis after balloon overstretch injury, and reduced degrees of constrictive remodeling and neointimal hyperplasia.

Human recombinant activated protein C-coated stent for the prevention of restenosis in porcine coronary arteries.

Activated protein C (APC), an endogenous protein, inhibits inflammation and thrombosis and interrupts the coagulation cascade. Here, we investigated the effect of human recombinant APC on the development of neointimal hyperplasia in porcine coronary arteries. Yukon Choice bare metal stents were coated with 2.6 µg APC/mm(2). Under general anesthesia, APC-coated and bare stents were implanted in the left anterior descending and circumflex coronary arteries of 10 domestic pigs. During the 4-week follow-up, animals were treated with dual antiplatelet therapy and neointimal hyperplasia was evaluated via histology. Scanning electron microscopy indicated successful but unequal coating of stents with APC; nearly complete drug release occurred within 4 h. Enzyme-linked immunosorbent assay revealed that intracoronary stent implantation rapidly increased the levels of monocyte chemoattractant protein-1, an effect that was inhibited by APC release from the coated stent. Fibrin deposition and adventitial inflammation were significantly decreased 1 month after implanting APC-coated stents versus bare stents, paralleled by significantly smaller neointimal area (0.98 ± 0.92 vs. 1.44 ± 0.91 mm(2), P = 0.028), higher lumen area (3.47 ± 0.94 vs. 3.06 ± 0.91 mm(2), P = 0.046), and lower stenosis area (22.2 ± 21.2% vs. 32.1 ± 20.1%, P = 0.034). Endothelialization was complete with APC-coated but not bare (90%) stents. P-selectin immunostaining revealed significantly fewer activated endothelial cells in the neointima in the APC group (4.6 ± 1.9 vs. 11.6 ± 4.1%, P < 0.001). Thus, short exposure of coronary arteries to APC reduced inflammatory responses, neointimal proliferation, and in-stent restenosis, offering a promising therapy to improve clinical outcomes of coronary stenting. However, coating stents with APC for prolonged, controlled drug release remains technically challenging.

Peri-Interventional Triple Therapy with Dabigatran Modifies Vasomotion after Bare-Metal Stent Implantation in a Pig Coronary Artery Model.

(1) Background: Coronary artery stenting leads to local inflammation, disturbs vasomotion, and slows endothelialization, increasing vascular thrombus risk. We used a pig stenting coronary artery model to assess how peri-interventional triple therapy with dabigatran ameliorates these effects. (2) Methods: In a total of 28 pigs bare-metal stents were implanted. Four days before the percutaneous coronary intervention (PCI), we started 16 of the animals on dabigatran, maintained through 4 days after the procedure. As controls, the remaining 12 pigs received no therapy. In both groups, dual antiplatelet therapy (DAPT) (clopidogrel, 75 mg plus aspirin, 100 mg) was administered until animals were euthanized. Just after the PCI and on day 3 after the procedure, we performed optical coherence tomography (OCT) in eight animals in the dabigatran group and four controls and euthanized them. We followed the eight remaining animals in each group with OCT and angiography for one month before euthanizing them and performed in vitro myometry and histology on harvested coronary arteries from all animals. (3) Results: The dabigatran group showed a significantly increased vasoconstriction at 3 days after PCI (10.97 ± 3.85 mN vs. 7.32 ± 5.41 mN, p = 0.03), but we found no differences between endothelium-dependent and -independent vasodilatation. We also found no group differences in OCT, quantitative angiography, or histomorphometry findings. (4) Conclusions: Starting a short course of dabigatran just before PCI and continuing for a 3-day window along with usual post-PCI DAPT is associated with enhanced vasoconstriction after bare-metal stent implantation without reducing neointimal formation at one month.

Improvement of Symptoms and Cardiac Magnetic Resonance Abnormalities in Patients with Post-Acute Sequelae of SARS-CoV-2 Cardiovascular Syndrome (PASC-CVS) after Guideline-Oriented Therapy.

Cardiac magnetic resonance (CMR) studies reported CMR abnormalities in patients with mild-moderate SARS-CoV-2 infection, suggesting ongoing myocardial inflammation. Patients (n = 278, 43 ± 13 years, 70.5% female) with post-acute sequelae of SARS-CoV-2 cardiovascular syndrome (PASC-CVS) were included prospectively into the Vienna POSTCOV Registry between March 2021 and March 2023 (clinicaltrials.gov NCT05398952). Clinical, laboratory, and CMR findings were recorded. Patients with abnormal CMR results were classified into isolated chronic pericardial (with/without pleural) effusion, isolated cardiac function impairment, or both (myopericarditis) groups. Medical treatment included a nonsteroidal anti-inflammatory agent (NSAID) for pericardial effusion and a condition-adapted maximal dose of heart failure (HF) treatment. Three months after medical therapy, clinical assessment and CMR were repeated in 82 patients. Laboratory analyses revealed normal hematological, inflammatory, coagulation, and cardiac biomarkers. CMR abnormalities were found in 155 patients (55.8%). Condition-adapted HF treatment led to a significant increase in the left ventricular ejection fraction (LVEF) in patients with initially reduced LVEF (from 49 ± 5% to 56 ± 4%, p = 0.009, n = 25). Low-moderate doses of NSAIDs for 3 months significantly reduced pericardial effusion (from 4/3;5.75/mm to 2/0;3/mm, median/interquartile ranges/p < 0.001, n = 51). Clinical symptoms improved markedly with a decrease in CMR abnormalities, which might be attributed to the maintenance of NSAID and HF medical treatment for PASC-CVS.

Effect of monovalent COVID-19 vaccines on viral interference between SARS-CoV-2 and several DNA viruses in patients with long-COVID syndrome.

Epstein-Barr virus (EBV) reactivation may be involved in long-COVID symptoms, but reactivation of other viruses as a factor has received less attention. Here we evaluated the reactivation of parvovirus-B19 and several members of the Herpesviridae family (DNA viruses) in patients with long-COVID syndrome. We hypothesized that monovalent COVID-19 vaccines inhibit viral interference between SARS-CoV-2 and several DNA viruses in patients with long-COVID syndrome, thereby reducing clinical symptoms. Clinical and laboratory data for 252 consecutive patients with PCR-verified past SARS-CoV-2 infection and long-COVID syndrome (155 vaccinated and 97 non-vaccinated) were recorded during April 2021-May 2022 (median 243 days post-COVID-19 infection). DNA virus-related IgG and IgM titers were compared between vaccinated and non-vaccinated long-COVID patients and with age- and sex-matched non-infected, unvaccinated (pan-negative for spike-antibody) controls. Vaccination with monovalent COVID-19 vaccines was associated with significantly less frequent fatigue and multiorgan symptoms (p < 0.001), significantly less cumulative DNA virus-related IgM positivity, significantly lower levels of plasma IgG subfractions 2 and 4, and significantly lower quantitative cytomegalovirus IgG and IgM and EBV IgM titers. These results indicate that anti-SARS-CoV-2 vaccination may interrupt viral cross-talk in patients with long-COVID syndrome (ClinicalTrials.gov Identifier: NCT05398952).

A CircRNA-miRNA-mRNA Network for Exploring Doxorubicin- and Myocet-Induced Cardiotoxicity in a Translational Porcine Model.

Despite the widespread use of doxorubicin (DOX) as a chemotherapeutic agent, its severe cumulative cardiotoxicity represents a significant limitation. While the liposomal encapsulation of doxorubicin (Myocet, MYO) reduces cardiotoxicity, it is crucial to understand the molecular background of doxorubicin-induced cardiotoxicity. Here, we examined circular RNA expression in a translational model of pigs treated with either DOX or MYO and its potential impact on the global gene expression pattern in the myocardium. This study furthers our knowledge about the regulatory network of circRNA/miRNA/mRNA and its interaction with chemotherapeutics. Domestic pigs were treated with three cycles of anthracycline drugs (DOX, n = 5; MYO, n = 5) to induce cardiotoxicity. Untreated animals served as controls (control, n = 3). We applied a bulk mRNA-seq approach and the CIRIquant algorithm to identify circRNAs. The most differentially regulated circRNAs were validated under cell culture conditions, following forecasting of the circRNA-miRNA-mRNA network. We identified eight novel significantly regulated circRNAs from exonic and mitochondrial regions in the porcine myocardium. The forecasted circRNA-miRNA-mRNA network suggested candidate circRNAs that sponge miR-17, miR-15b, miR-130b, the let-7 family, and miR125, together with their mRNA targets. The identified circRNA-miRNA-mRNA network provides an updated, coherent view of the mechanisms involved in anthracycline-induced cardiotoxicity.

Cell-Based HIF1α Gene Therapy Reduces Myocardial Scar and Enhances Angiopoietic Proteome, Transcriptomic and miRNA Expression in Experimental Chronic Left Ventricular Dysfunction.

Recent preclinical investigations and clinical trials with stem cells mostly studied bone-marrow-derived mononuclear cells (BM-MNCs), which so far failed to meet clinically significant functional study endpoints. BM-MNCs containing small proportions of stem cells provide little regenerative potential, while mesenchymal stem cells (MSCs) promise effective therapy via paracrine impact. Genetic engineering for rationally enhancing paracrine effects of implanted stem cells is an attractive option for further development of therapeutic cardiac repair strategies. Non-viral, efficient transfection methods promise improved clinical translation, longevity and a high level of gene delivery. Hypoxia-induced factor 1α is responsible for pro-angiogenic, anti-apoptotic and anti-remodeling mechanisms. Here we aimed to apply a cellular gene therapy model in chronic ischemic heart failure in pigs. A non-viral circular minicircle DNA vector (MiCi) was used for in vitro transfection of porcine MSCs (pMSC) with HIF1α (pMSC-MiCi-HIF-1α). pMSCs-MiCi-HIF-1α were injected endomyocardially into the border zone of an anterior myocardial infarction one month post-reperfused-infarct. Cell injection was guided via 3D-guided NOGA electro-magnetic catheter delivery system. pMSC-MiCi-HIF-1α delivery improved cardiac output and reduced myocardial scar size. Abundances of pro-angiogenic proteins were analyzed 12, 24 h and 1 month after the delivery of the regenerative substances. In a protein array, the significantly increased angiogenesis proteins were Activin A, Angiopoietin, Artemin, Endothelin-1, MCP-1; and remodeling factors ADAMTS1, FGFs, TGFb1, MMPs, and Serpins. In a qPCR analysis, increased levels of angiopeptin, CXCL12, HIF-1α and miR-132 were found 24 h after cell-based gene delivery, compared to those in untreated animals with infarction and in control animals. Expression of angiopeptin increased already 12 h after treatment, and miR-1 expression was reduced at that time point. In total, pMSC overexpressing HIF-1α showed beneficial effects for treatment of ischemic injury, mediated by stimulation of angiogenesis.

Peri-interventional Triple Therapy With Dabigatran Improves Vasomotion and Promotes Endothelialization in Porcine Coronary Stenting Model.

Objective: We evaluated the short and long-term effect of peri-interventional dabigatran therapy on vasomotion, endothelialization, and neointimal formation in a porcine coronary artery stenting model. Background: Stenting of coronary arteries induces local inflammation, impairs vasomotion and delays endothelialization. Methods: Twenty-eight animals underwent percutaneous coronary intervention (PCI) with drug eluting stents. Sixteen pigs started dabigatran therapy 4 days prior to PCI and continued for 4 days post-stenting, while 12 animals served as controls. Post-stenting dual antiplatelet therapy (75 mg clopidogrel and 100 mg aspirin) was continued in both groups until termination. Immediately post-stenting and at day 3 optical coherence tomography (OCT) was performed in all animals, followed by euthanasia of 8 dabigatran and 4 control animals. The remaining pigs (8 of each group) were followed up for 1 month, with control angiography and OCT. Tissue burden (degree of peri-strut structure-thrombus and/or fibrin) was evaluated. After euthanasia coronary arteries were harvested for in-vitro myometry and histology. Results: Thrombin generation was lower (p < 0.001) and tissue burden (0.83 ± 0.98 vs. 3.0 ± 2.45; p = 0.031) was significantly decreased in dabigatran treated animals. After 3 days post-PCI endothelium-dependent vasodilation was significantly improved (77 ± 40% vs. 41 ± 31%, p = 0.02) in dabigatran animals. Neither quantitative angiography nor histomorphometry showed differences between the groups. Endothelialization was faster in the dabigatran group as compared with controls (p = 0.045). Conclusion: Short-term peri-interventional triple therapy with dabigatran, aspirin, and clopidogrel led to an enhanced endothelium dependent vasodilation and faster endothelialization. However, neointimal formation 1-month after stent implantation was comparable between groups.

New Insights and Current Approaches in Cardiac Hypertrophy Cell Culture, Tissue Engineering Models, and Novel Pathways Involving Non-Coding RNA.

Cardiac hypertrophy is an ongoing clinical challenge, as risk factors such as obesity, smoking and increasing age become more widespread, which lead to an increasing prevalence of developing hypertrophy. Pathological hypertrophy is a maladaptive response to stress conditions, such as pressure overload, and involve a number of changes in cellular mechanisms, gene expression and pathway regulations. Although several important pathways involved in the remodeling and hypertrophy process have been identified, further research is needed to achieve a better understanding and explore new and better treatment options. More recently discovered pathways showed the involvement of several non-coding RNAs, including micro RNAs (miRNAs), long non-coding RNAs (lncRNAs), and circular RNAs (circRNAs), which either promote or inhibit the remodeling process and pose a possible target for novel therapy approaches. In vitro modeling serves as a vital tool for this further pathway analysis and treatment testing and has vastly improved over the recent years, providing a less costly and labor-intensive alternative to in vivo animal models.

Large Animal Models of Cell-Free Cardiac Regeneration.

The adult mammalian heart lacks the ability to sufficiently regenerate itself, leading to the progressive deterioration of function and heart failure after ischemic injuries such as myocardial infarction. Thus far, cell-based therapies have delivered unsatisfactory results, prompting the search for cell-free alternatives that can induce the heart to repair itself through cardiomyocyte proliferation, angiogenesis, and advantageous remodeling. Large animal models are an invaluable step toward translating basic research into clinical applications. In this review, we give an overview of the state-of-the-art in cell-free cardiac regeneration therapies that have been tested in large animal models, mainly pigs. Cell-free cardiac regeneration therapies involve stem cell secretome- and extracellular vesicles (including exosomes)-induced cardiac repair, RNA-based therapies, mainly regarding microRNAs, but also modified mRNA (modRNA) as well as other molecules including growth factors and extracellular matrix components. Various methods for the delivery of regenerative substances are used, including adenoviral vectors (AAVs), microencapsulation, and microparticles. Physical stimulation methods and direct cardiac reprogramming approaches are also discussed.