Sarcomeric network analysis of ex vivo cultivated human atrial appendage tissue using super-resolution microscopy.

Investigating native human cardiac tissue with preserved 3D macro- and microarchitecture is fundamental for clinical and basic research. Unfortunately, the low accessibility of the human myocardium continues to limit scientific progress. To overcome this issue, utilizing atrial appendages of the human heart may become highly beneficial. Atrial appendages are often removed during open-heart surgery and can be preserved ex vivo as living tissue with varying durability depending on the culture method. In this study, we prepared living thin myocardial slices from left atrial appendages that were cultured using an air-liquid interface system for overall 10 days. Metabolic activity of the cultured slices was assessed using a conventional methyl thiazolyl tetrazolium (MTT) assay. To monitor the structural integrity of cardiomyocytes within the tissue, we implemented our recently described super-resolution microscopy approach that allows both qualitative and quantitative in-depth evaluation of sarcomere network based on parameters such as overall sarcomere content, filament size and orientation. Additionally, expression of mRNAs coding for key structural and functional proteins was analyzed by real-time reverse transcription polymerase chain reaction (qRT-PCR). Our findings demonstrate highly significant disassembly of contractile apparatus represented by degradation of [Formula: see text]-actinin filaments detected after three days in culture, while metabolic activity was constantly rising and remained high for up to seven days. However, gene expression of crucial cardiac markers strongly decreased after the first day in culture indicating an early destructive response to ex vivo conditions. Therefore, we suggest static cultivation of living myocardial slices derived from left atrial appendage and prepared according to our protocol only for short-termed experiments (e.g. medicinal drug testing), while introduction of electro-mechanical stimulation protocols may offer the possibility for long-term integrity of such constructs.

ECG-gated MR angiography at 3T for follow-up after surgery involving the ascending aorta.

We aimed to evaluate electrocardiogram (ECG)-gated MR angiography (MRA) in the follow-up after surgery involving the ascending aorta regarding technical feasibility, image quality, spectrum of findings, and their implications for clinical management. We retrospectively analyzed a cohort of 19 patients (median age 59 years, range 38-79 years), who underwent MRA for follow-up imaging after surgery involving the ascending aorta. Our magnetic resonance imaging protocol consisted of a time-resolved, non-ECG-gated MRA and an ECG-gated MRA performed at 3T. Median examination duration was 25 minutes (range 11-41 minutes). All examinations were assessed by 2 readers in consensus for image quality on a 5-point scale ranging from 1 (non-diagnostic) to 5 (excellent). MRA examinations and patient charts were analyzed for diagnostic findings and their consequences for further management. Subjective image quality was rated as "sufficient" (score 3.1 ±â€…1.1) for the aortic root and as "good" to "excellent" for the ascending aorta (score 4.5 ±â€…0.7), aortic arch (4.5 ±â€…0.7), supra-aortic branches (4.5 ±â€…0.6) and descending aorta (4.6 ±â€…0.7). Abnormal findings were seen in 6 patients (32%) including progressive diameter of remaining aneurysm or dissection (3 patients, 16%) and suture aneurysms (3 patients, 16%). In all 6 of these patients, abnormal findings at MRA had consequences for clinical management. ECG-gated MR angiography at 3T yields good image quality for post-operative surveillance after aortic surgery involving the ascending aorta. This technique may serve as an alternative to computed tomography particularly in younger patients with repeated follow-up.

Surgery for Valvular and Nonvalvular Papillary Fibroelastomas.

Papillary fibroelastomas (PFE) are benign neoplasms, mostly located on valvular surfaces with high embolic potential. This study presents a 27-year single institutional experience on surgical treatment of PFE in an adult patient- cohort with long-term follow-up. This study was approved by the institutional review board. Date and number of IRB approval: 11/23/2017, Institutional Review Board approval number A2014-0149. The need for individual patient consent was waived. We retrospectively evaluated all patients who underwent cardiac surgery for suspected space-occupying lesions in the observation period between June 1991 and June 2018 at our hospital. Clinicopathological features, imaging characteristics, surgical procedures and disease outcome were analyzed. 120 patients were diagnosed with various primary/secondary cardiac tumors and histology confirmed 21 PFEs were found in 16 patients. There was no significant age difference between patients with valvular vs nonvalvular PFEs (P = 0.26). Valvular lesions were found in aortic valve (n = 6), mitral valve (n = 2) and tricuspid valve (n = 1). Nonvalvular PFEs were found in right atrium (n = 2), left ventricle (n = 2), left atrial appendage (n = 2) and aortic wall (n = 1). Valvular lesions were significantly smaller in size compared to non-valvular lesions (P = 0.0013). Left-side PFEs were associated with a high embolization episodes (10/13 patients, 77%) not related to the size. One patient died in-hospital. All other patients were discharged out of the hospital postoperative. Follow-up was performed regularly for a median of 2.8 years (range 0.1-11 years) postoperative. Nonvalvular PFE tended to be larger in size and at least when located on the left sided heart had equally high propensity to embolize compared to valvular PFE. We strongly advocate surgical excision in all left-sided PFE.

Electrophysiological studies in patients with paroxysmal supraventricular tachycardias but no electrocardiogram documentation: findings from a prospective registry.

AIMS: A significant proportion of patients presenting with paroxysmal supraventricular tachycardia (PSVT) has no electrocardiogram (ECG) documentation. In these patients an electrophysiological study (EPS) may be performed to facilitate the diagnosis. METHODS AND RESULTS: In a prospective registry we compared the prevalence of inducible arrhythmias and the clinical outcome in 525 patients with and without ECG documentation. Compared with patients with a documented PSVT a smaller but substantial proportion of patients (63.7%) without ECG documentation had inducible supraventricular tachycardias (SVT). Atrio-ventricular nodal reentrant tachycardia was the most common type in both groups. Patients with an inducible SVT and no documentation were significantly younger, had a shorter episode duration and a lower hospitalization rate, which may be the cause for the lacking documentation. Similar to patients with documented PSVTs most of these patients (90.0%) were asymptomatic or clinically improved after the EPS. Even 43% of patients without an inducible tachycardia improved clinically, probably due to a placebo effect of the EPS. In particular, patients between 31 and 60 years of age seemed to benefit from an EPS because they were more likely to have inducible SVTs that could be cured by radiofrequency ablation. CONCLUSION: Our data show that a substantial proportion of patients with suspected paroxysmal tachycardia, but without ECG documentation, have inducible SVTs and obtain a clear clinical benefit from an EPS. Thus, our data provide justification for using EPS for patients in this category. To the best of our knowledge, ours is the first prospective registry that supports this approach.

Prevalence of atrial fibrillation in patients with high CHADS2- and CHA2DS2VASc-scores: anticoagulate or monitor high-risk patients?

BACKGROUND: In patients with known atrial fibrillation (AF) different scores are utilized to estimate the risk of thromboembolic events and guide oral anticoagulation. Diagnosis of AF strongly depends on the duration of electrocardiogram monitoring. The aim of this study was to use established scores to predict the prevalence of AF. METHODS: The CHADS2- (Congestive Heart failure, hypertension, Age >75 years, Diabetes, Stroke [doubled]) and CHA2DS2VASc-score (Congestive Heart failure, hypertension, Age ≥75 years [doubled], Diabetes, Stroke [doubled], Vascular disease, Age 65-74 years, Sex category [female sex]) was calculated in 150,408 consecutive patients, referred to the University Hospital of Rostock between 2007 and 2012. All factors constituting these scores and a history of AF were prospectively documented with the ICD-10 admission codes. RESULTS: Mean age of our study population was 67.6 ± 13.6 years with a mean CHADS2-score of 1.65 ± 0.92 and CHA2DS2VASc-score of 3.04 ± 1.42. AF was prevalent in 15.9% of the participants. The prevalence of AF increased significantly with every CHADS2- and CHA2DS2VASc-score point up to 54.2% in CHADS2-score of 6 and 71.4% in CHA2DS2VASc-score of 9 (P < 0.001). CONCLUSION: The prevalence of AF increases with increasing CHADS2- and CHA2DS2VASc-score. In intermediate scores intensified monitoring may be recommended. In high scores, thromboembolic complications occurred irrespective of the presence of AF and anticoagulant therapy may be initiated irrespective of documented AF.

Polyethylenimine-mediated gene delivery into human bone marrow mesenchymal stem cells from patients.

Transplantation of mesenchymal stem cells (MSCs) derived from adult bone marrow has been proposed as a potential therapeutic approach for post-infarction left ventricular (LV) dysfunction. However, age-related functional decline of stem cells has restricted their clinical benefits after transplantation into the infarcted myocardium. The limitations imposed on patient cells could be addressed by genetic modification of stem cells. This study was designed to improve our understanding of genetic modification of human bone marrow derived mesenchymal stem cells (hMSCs) by polyethylenimine (PEI, branched with Mw 25 kD), one of non-viral vectors that show promise in stem cell genetic modification, in the context of cardiac regeneration for patients. We optimized the PEI-mediated reporter gene transfection into hMSCs, evaluated whether transfection efficiency is associated with gender or age of the cell donors, analysed the influence of cell cycle on transfection and investigated the transfer of therapeutic vascular endothelial growth factor gene (VEGF). hMSCs were isolated from patients with cardiovascular disease aged from 41 to 85 years. Optimization of gene delivery to hMSCs was carried out based on the particle size of the PEI/DNA complexes, N/P ratio of complexes, DNA dosage and cell viability. The highest efficiency with the cell viability near 60% was achieved at N/P ratio 2 and 6.0 µg DNA/cm(2) . The average transfection efficiency for all tested samples, middle-age group (<65 years), old-age group (>65 years), female group and male group was 4.32%, 3.85%, 4.52%, 4.14% and 4.38%, respectively. The transfection efficiency did not show any correlation either with the age or the gender of the donors. Statistically, there were two subpopulations in the donors; and transfection efficiency in each subpopulation was linearly related to the cell percentage in S phase. No significant phenotypic differences were observed between these two subpopulations. Furthermore, PEI-mediated therapeutic gene VEGF transfer could significantly enhance the expression level.

Stem cell-mediated neovascularization in heart repair.

Accumulating clinical and experimental evidence indicates that stem cells from various sources are promising in the treatment of cardiac dysfunction. They may be incorporated into neovascular foci and thus contribute to postnatal physiological and pathological vasculogenesis and/or produce a variety of growth factors for angiogenesis and cytokines that home other stem cells from other organs for cardiac regeneration. This review focuses on the neovascularization of stem cells from different sources in cardiac repair, with emphasis on adult stem cells.

Mesenchymal stem cells and cardiac repair.

Accumulating clinical and experimental evidence indicates that mesenchymal stem cells (MSCs) are promising cell types in the treatment of cardiac dysfunction. They may trigger production of reparative growth factors, replace damaged cells and create an environment that favours endogenous cardiac repair. However, identifying mechanisms which regulate the role of MSCs in cardiac repair is still at work. To achieve the maximal clinical benefits, ex vivo manipulation can further enhance MSC therapeutic potential. This review focuses on the mechanism of MSCs in cardiac repair, with emphasis on ex vivo manipulation.

Bcl-2 engineered MSCs inhibited apoptosis and improved heart function.

Engraftment of mesenchymal stem cells (MSCs) derived from adult bone marrow has been proposed as a potential therapeutic approach for postinfarction left ventricular dysfunction. However, limited cell viability after transplantation into the myocardium has restricted its regenerative capacity. In this study, we genetically modified MSCs with an antiapoptotic Bcl-2 gene and evaluated cell survival, engraftment, revascularization, and functional improvement in a rat left anterior descending ligation model via intracardiac injection. Rat MSCs were manipulated to overexpress the Bcl-2 gene. In vitro, the antiapoptotic and paracrine effects were assessed under hypoxic conditions. In vivo, the Bcl-2 gene-modified MSCs (Bcl-2-MSCs) were injected after myocardial infarction. The surviving cells were tracked after transplantation. Capillary density was quantified after 3 weeks. The left ventricular function was evaluated by pressure-volume loops. The Bcl-2 gene protected MSCs against apoptosis. In vitro, Bcl-2 overexpression reduced MSC apoptosis by 32% and enhanced vascular endothelial growth factor secretion by more than 60% under hypoxic conditions. Transplantation with Bcl-2-MSCs increased 2.2-fold, 1.9-fold, and 1.2-fold of the cellular survival at 4 days, 3 weeks, and 6 weeks, respectively, compared with the vector-MSC group. Capillary density in the infarct border zone was 15% higher in Bcl-2-MSC transplanted animals than in vector-MSC treated animals. Furthermore, Bcl-2-MSC transplanted animals had 17% smaller infarct size than vector-MSC treated animals and exhibited functional recovery remarkably. Our current findings support the premise that transplantation of antiapoptotic gene-modified MSCs may have values for mediating substantial functional recovery after acute myocardial infarction.