Epicardial Layer Research Articles

The influence of genetic mutations on multilayer global longitudinal strain values in patients with hypertrophic cardiomyopaty

Abstract Background Hypertrophic cardiomyopathy (HCM) is a hereditary heart muscle disorder characterized by diverse genetic mutations. Multilayer global longitudinal strain (GLS) analysis has emerged as a valuable tool for assessing myocardial function. This study aimed to investigate the influence of genetic mutations on myocardial deformation patterns in HCM patients using multilayer GLS analysis. Methods A total of 83 adult patients diagnosed with HCM were included in the study. Genetic testing was performed to identify specific genetic mutations associated with HCM. Echocardiographic assessments were conducted to measure multilayer GLS values for endocardial, mid-myocardial, and epicardial layers. Patients were divided into gene-positive and gene-negative groups for comparative analysis. Results In the gene-negative group (n=41), the mean age was 57.1 ± 11.0 years, whereas in the gene-positive group (n=42), it was 55.5 ± 11.6 years (p=0.516). Significant differences were observed in endocardial (-16.1 ± 2.3 vs. -13.4 ± 3.2, p < 0.001) and mid-myocardial (-14.2 ± 2.1 vs. -12.3 ± 3.1, p = 0.002) GLS involvement between gene-negative and gene-positive HCM patients. However, no significant disparity in epicardial strain was detected. Conclusion The findings demonstrated a greater involvement of the endocardial and mid-myocardial layers in the group of HCM patients with positive genetic mutations. Multilayer GLS analysis has the potential to aid in the identification of subtle alterations in myocardial function associated with specific genetic mutations, contributing to refined risk assessment and personalized management strategies for HCM patients.

Report of a giant invasive, wall-penetrating cardiac lipoma

BackgroundCardiac lipoma, a seldom-encountered benign tumor positioned beneath the endocardium, has the potential to impair electrophysiological functions. Diagnosis is principally based on imaging modalities. The uniqueness of this case lies in the tumor’s extension both internally and externally within the right atrium, rendering it of special interest. From a clinical standpoint, surgical removal is commonly advocated, wherein early intervention is pivotal in improving patients’ long-term prognoses.Case presentationA 35-year-old male was admitted to the hospital for treatment subsequent to the identification of a cardiac mass two days prior. Initial diagnostic assessments, encompassing CT scans and echocardiography, identified a space-occupying lesion within the heart. The patient underwent surgical excision of the cardiac tumor, utilizing mild hypothermic extracorporeal circulation via femoral vessel access. Intraoperative findings revealed adipose-like tissue of a “dumbbell-shaped” configuration situated both within and external to the right atrium, measuring approximately 8 cm*9 cm internally and 7 cm*6 cm externally, with the extracardiac mass being marginally larger. Postoperative pathological analysis confirmed a cardiac lipoma diagnosis. A follow-up echocardiogram conducted three months post-surgery exhibited no notable abnormalities. The patient is under continuous observation to monitor for any recurrence or potential long-term complications.ConclusionIn this case report, we detail with precision a rare cardiac pathology manifested by an expansive infiltrative lipoma that pervades the endocardial and epicardial layers of the right atrium. After thorough preoperative diagnostic workup and evaluation, we contend that surgical intervention represents the optimal therapeutic approach for managing such conditions, with the goal of preemptively reducing the likelihood of cardiac compression or intracardiac obstruction.

X-ray Radiotherapy Impacts Cardiac Dysfunction by Modulating the Sympathetic Nervous System and Calcium Transients.

Recent epidemiological studies have shown that patients with right-sided breast cancer (RBC) treated with X-ray irradiation (IR) are more susceptible to developing cardiovascular diseases, such as arrhythmias, atrial fibrillation, and conduction disturbances after radiotherapy (RT). Our aim was to investigate the mechanisms induced by low to moderate doses of IR and to evaluate changes in the cardiac sympathetic nervous system (CSNS), atrial remodeling, and calcium homeostasis involved in cardiac rhythm. To mimic the RT of the RBC, female C57Bl/6J mice were exposed to X-ray doses ranging from 0.25 to 2 Gy targeting 40% of the top of the heart. At 60 weeks after RI, Doppler ultrasound showed a significant reduction in myocardial strain, ejection fraction, and atrial function, with a significant accumulation of fibrosis in the epicardial layer and apoptosis at 0.5 mGy. Calcium transient protein expression levels, such as RYR2, NAK, Kir2.1, and SERCA2a, increased in the atrium only at 0.5 Gy and 2 Gy at 24 h, and persisted over time. Interestingly, 3D imaging of the cleaned hearts showed an early reduction of CSNS spines and dendrites in the ventricles and a late reorientation of nerve fibers, combined with a decrease in SEMA3a expression levels. Our results showed that local heart IR from 0.25 Gy induced late cardiac and atrial dysfunction and fibrosis development. After IR, ventricular CSNS and calcium transient protein expression levels were rearranged, which affected cardiac contractility. The results are very promising in terms of identifying pro-arrhythmic mechanisms and preventing arrhythmias during RT treatment in patients with RBC.

Advanced myocardial deformation echocardiography for evaluation of the athlete's heart: Functional and mechanistic analysis

BackgroundAssessment of the athlete's heart is challenging because of a phenotypic overlap between reactive physiological adaptation and pathological remodelling. The potential value of myocardial deformation remains controversial in identifying early cardiomyopathy. AimTo identify the echocardiographic phenotype of athletes using advanced two-dimensional speckle tracking imaging, and to define predictive factors of subtle left ventricular systolic dysfunction. MethodsIn total, 191 healthy male athletes who underwent a preparticipation medical evaluation at Nancy University Hospital between 2013 and 2020 were included. Clinical and echocardiographic data were compared with 161 healthy male subjects from the STANISLAS cohort. Borderline global longitudinal strain value was defined as<17.5%. ResultsAthletes demonstrated lower left ventricular ejection fraction (57.9±5.3% vs. 62.6±6.4%; P<0.01) and lower global longitudinal strain (17.5±2.2% vs. 21.1±2.1%; P<0.01). No significant differences were found between athletes with and without a borderline global longitudinal strain value regarding clinical characteristics, structural echocardiographic features and exercise capacity. A borderline global longitudinal strain value was associated with a lower endocardial global longitudinal strain (18.8±1.2% vs. 22.7±1.9%; P=0.02), a lower epicardial global longitudinal strain (14.0±1.1% vs. 16.6±1.2%; P<0.01) and a higher endocardial/epicardial global longitudinal strain ratio (1.36±0.07 vs. 1.32±0.06; P<0.01). No significant difference was found regarding mechanical dispersion (P=0.46). ConclusionsBorderline global longitudinal strain value in athletes does not appear to be related to structural remodelling, mechanical dispersion or exercise capacity. The athlete's heart is characterized by a specific myocardial deformation pattern with a more pronounced epicardial layer strain impairment.

Fatty infiltration and ventricular premature beats originating from right ventricular outflow tract: association or causality?

Anatomical evidence reveals heterogeneous fat distribution in both atrial and ventricular myocardium that are considered normal, but at the same time arrhythmogenic, and numerous cardiac pathophysiological conditions are associated with myocardial fat deposits. The relationship between fatty infiltration, especially in the epicardial layer and its pathophysiological implication is not completely understood. The aim of this study was to establish a positive or negative relationship between the ventricular burden and several parameters related to right ventricle (RV) adipose tissue - the RV thickness, RV indexed mass, body mass index (BMI), age, gender. Twenty-three patients with documented premature ventricular contractions (PVCs) originating from right ventricular outflow tract based on electrocardiography (ECG) evaluation were hospitalized between January 2018-November 2022 for electrophysiological study and PVCs ablation. Data obtained after collecting the clinical characteristics, ECG, RV measurements from transthoracic echocardiography (TTE), cardiac computed tomography (CT) and magnetic resonance imaging (MRI) were analyzed. A weak positive relationship between the ventricular burden and BMI (r=0.14, p=0.49), tricuspid annular plane systolic excursion (TAPSE) (r=0.07, p=0.7), the RV thickness (r=0.03, p=0.8), epicardial adipose tissue (r=0.13, p=0.55), RV mass indexed (r=0.05, p=0.82) was observed. No clear cut-off of the PVCs burden could be established in terms related to the increase in BMI, RV thickness, epicardial adipose tissue, RV mass indexed. No significant positive or negative relationship between the ventricular burden and the RV thickness, RV indexed mass were found in individuals with a high PVCs originating from right ventricular outflow tract (RVOT) burden.

Increasing region of interest width reduces neonatal circumferential strain.

There is growing interest in speckle tracking echocardiography-derived strain as a measure of left ventricular function in neonates. However, knowledge gaps remain regarding the effect of image acquisition and processing parameters on circumferential strain measurements. The aim of this study was to evaluate the effect of using different region of interest (ROI) widths on speckle tracking derived circumferential strain in healthy neonates. Thirty healthy-term-born neonates were examined with speckle-tracking echocardiography in the short-axis view. Circumferential strain values were acquired and compared using two different ROI widths. Furthermore, strain values in the different vendor-defined wall layers were also compared. Increasing ROI width led to a decrease in global circumferential strain (GCS) in the midwall and epicardial layers, the respective decreases in strain being -23.4±.6% to -22.0±1.1%, p<.0001 and 18.5±1.7% to -15.6±2.0%, p<.0001. Segmental analyses were consistent with these results, apart from two segments in the midwall. There was no statistically significant effect on strain for the endocardial layer. A gradient was seen where strain increased from the epicardial to endocardial layers. Increasing ROI width led to a decrease in GCS in the midwall and epicardium. There is an increase in circumferential strain when moving from the epicardial toward the endocardial layer. Clinicians wishing to implement circumferential strain into their practice should consider ROI width variation as a potential confounder in their measurements.

Llgl1 mediates timely epicardial emergence and establishment of an apical laminin sheath around the trabeculating cardiac ventricle.

During heart development, the embryonic ventricle becomes enveloped by the epicardium, which adheres to the outer apical surface of the heart. This is concomitant with onset of ventricular trabeculation, where a subset of cardiomyocytes lose apicobasal polarity and delaminate basally from the ventricular wall. Llgl1 regulates the formation of apical cell junctions and apicobasal polarity, and we investigated its role in ventricular wall maturation. We found that llgl1 mutant zebrafish embryos exhibit aberrant apical extrusion of ventricular cardiomyocytes. While investigating apical cardiomyocyte extrusion, we identified a basal-to-apical shift in laminin deposition from the internal to the external ventricular wall. We find that epicardial cells express several laminin subunits as they adhere to the ventricle, and that the epicardium is required for laminin deposition on the ventricular surface. In llgl1 mutants, timely establishment of the epicardial layer is disrupted due to delayed emergence of epicardial cells, resulting in delayed apical deposition of laminin on the ventricular surface. Together, our analyses reveal an unexpected role for Llgl1 in correct timing of epicardial development, supporting integrity of the ventricular myocardial wall.

Early diagnostic value of carotid artery ultrasound parameters combined with epicardial adipose layer thickness in coronary heart disease

BACKGROUND Coronary heart disease is associated with coronary atherosclerosis indicated by carotid intima-media thickness (CIMT) thickening and altered vascular elasticity. The epicardial adipose layer can secrete proinflammatory factors that promote the formation of coronary atherosclerosis. Thus, the epicardial fat layer thickness (EAT) may also predict coronary heart disease. AIM To determine the role of common carotid artery ultrasound parameters and EAT in the early diagnosis of coronary artery disease. METHODS Based on coronary angiography, patients with newly suspected coronary heart disease were divided into case (n = 107) and control (n = 41) groups. The carotid ultrasound parameters, including vascular stiffness (β), elastic coefficient (EP), pulse wave conduction velocity (PWV-β), CIMT, and EAT were compared between the case and control groups and among patients with different lesion numbers in the case group. Pearson correlation was used to evaluate the early diagnostic value of EAT, common carotid artery elasticity, and CIMT for coronary heart disease. RESULTS EP, β, PWV-β, CIMT, and EAT were significantly higher in the case group compared with the levels in the control group (all P &lt; 0.001). In the case group, lesions were detected in one vessel in 34 patients, two vessels in 38 patients, and three vessels in 35 patients. Within the case group, β, EP, PWV-β, CIMT, and EAT levels significantly increased with an increased number of lesions (all P &lt; 0.001). EAT positively correlated with β, EP, PWV-β, and CIMT (all P &lt; 0.01). The area under the curve for diagnosing coronary heart disease using EAT combined with CIMT and carotid elasticity was 0.893, and the sensitivity and specificity were 0.890 and 0.837. CONCLUSION EAT correlated well with changes in carotid artery elasticity and CIMT in patients with coronary heart disease. The combination of EAT, carotid artery elasticity, and CIMT facilitates the early diagnosis of coronary heart disease.

Single-cell RNA sequencing analysis of the human fetal epicardium reveals several new genes involved in regulation of epicardial activation

Abstract Funding Acknowledgements Type of funding sources: Foundation. Main funding source(s): Dutch Heart Foundation Background The outside mesothelial layer of the heart, the epicardium, is essential for cardiac development. Cells from this layer undergo epithelial-to-mesenchymal transition (EMT), migrate into the tissue and partake in cardiogenesis by contributing cells and by secreting growth factors. In the adult heart, the epicardium is quiescent but upon ischemic injury it is activated and conveys a partial regenerative response. This activation step is important for repair, but suboptimal compared to the cardiogenic potential during development. Understanding the processes and cells that underlie developmental epicardial activation could contribute to enhancing the adult post-injury response, but little is known about the human developing epicardium. Purpose to determine the cellular composition of the human fetal epicardial layer and to identify the processes that regulate the activation of the epicardium. Methods An epicardial-cell enriched dataset was obtained by specifically dissecting the epicardial layers from 4 fetal human hearts (14-15 weeks gestation, obtained under informed consent and according to local ethical approval). Single live cells were sorted into 384-wells plates and sequenced. Data analysis was performed on 2073 cells using R-packages RaceID3 and StemID2. Findings were validated using qPCR and immunohistochemistry on tissue sections and cell culture models. Results Sequencing analysis revealed a distinct clustering of epicardial (epithelial, pre-EMT) and epicardial-derived mesenchymal (post-EMT) cell populations. Besides the identification of novel markers for epicardial cell states, the regulation of epicardial cell activation and EMT was investigated by applying pseudo-time analysis. We selected genes that were highly regulated (&amp;gt;2 fold change) in this trajectory for further analysis in vitro. We identified several genes including CRIP1, SULF1 and others that are expressed in epithelial cells in vitro and in vivo and are rapidly downregulated upon EMT. The expression profiles positively correlated with epithelial cell related genes and negatively with mesenchymal cell genes. Based on GO-terms, and protein expression within the epicardial layers in fetal and adult epicardium, we anticipated a role for these genes in maintaining the epithelial-state. Indeed, siRNA experiments in vitro with cultured primary human epicardial cells confirmed an essential role in EMT regulation. We continue to evaluate other genes identified in our sequencing dataset, resulting in several other potent and previously unknown pathways involved in regulation of epicardial activation. Conclusions We shed light on the composition of the human fetal epicardium using a highly enriched scRNA dataset. We identify specific markers for different cell-states, and reveal novel regulators of activation that in the future may be influenced to optimize the post-injury response.

Exercise-induced regional heterogeneity of myocardial left ventricular contraction in hereditary long QT syndrome

Abstract Background Hereditary long QT syndrome (LQTS) is characterized by prolonged ventricular repolarization and an increased risk for ventricular arrhythmias (VA) and sudden cardiac death. Dispersion of ventricular repolarization in LQTS may be more pronounced during exercise and not only impacts the heart’s electrical functions but also the pattern of mechanical contraction. Little is known about the intraventricular heterogeneity of wall motion abnormalities in LQTS patients. Methods In this cross-sectional study we quantify regional, transmural, and longitudinal alterations in left ventricular (LV) contraction at rest, during exercise, and recovery in genetically confirmed LQTS patients and healthy controls to evaluate their value in differentiating high risk patients as defined by a history of VA. ECG and echocardiographic assessments were conducted during rest, peak ergometer activity (105 to 125 bpm), and four minutes into recovery phases. Longitudinal strain (LStr) parameters were measured for each LV segment separately. We differentiated LStr variations as heterogeneity of LStr amplitude (hetLStr; standard deviation [SD] or mean range) and dispersion of time-to-peak LStr (disLStr; SD or mean range). Here, we present the analyses of disLStr. Results At present, 15 patients with genetically confirmed LQTS (LQT1: 8, LQTS 2: 7; mean QTc 475 ± 46 ms) have been included of which 9 had a history of VA. In addition, we collected data from 8 age- and gender-matched control patients (mean QTc 401 ± 18 ms). LV ejection fraction and LStr were not significantly different between the LQTS and control groups. However, LQTS patients exhibited a significantly lower mean LStr in both the endocardial and epicardial layers across all phases compared to control. We observed a significantly more pronounced disLStr across all segments in the LQTS group during the rest and recovery phases in the endocardial layer and during all phases in the epicardial layer. Regional differences (apical vs. basal) were found in all LQTS patients only endocardially at rest. Globally, disLStr showed transmural differences (epicardial vs. endocardial) in both asymptomatic and symptomatic LQTS patients during peak exercise and recovery. Upon segmental analysis, it was observed that symptomatic patients exhibited these transmural differences apically already at rest (p=.004), as well as in the basal region during peak exercise (p=.01), and in the apical region during recovery (p=.02). Conclusion LQTS patients show regional (base to apex) and transmural (endocardial to epicardial) heterogeneities in myocardial contraction at rest, during exercise, and recovery as compared to healthy patients. These regional variations in myocardial contraction patterns may be more pronounced in high risk LQTS patients. Ongoing analyses of hetLStr are in progress to corroborate our findings.

Heart-on-a-Chip Model of Epicardial-Myocardial Interaction in Ischemia Reperfusion Injury.

Epicardial cells (EPIs) form the outer layer of the heart and play an important role in development and disease. Current heart-on-a-chip platforms still do not fully mimic the native cardiac environment due to the absence of relevant cell types, such as EPIs. Here, using the Biowire II platform, engineered cardiac tissues with an epicardial outer layer and inner myocardial structure are constructed, and an image analysis approach is developed to track the EPI cell migration in a beating myocardial environment. Functional properties of EPI cardiac tissues improve over two weeks in culture. In conditions mimicking ischemia reperfusion injury (IRI), the EPI cardiac tissues experience less cell death and a lower impact on functional properties. EPI cell coverage is significantly reduced and more diffuse under normoxic conditions compared to the post-IRI conditions. Upon IRI, migration of EPI cells into the cardiac tissue interior is observed, with contributions to alpha smooth muscle actin positive cell population. Altogether, a novel heart-on-a-chip model is designed toincorporate EPIs through a formation process that mimics cardiac development, and this work demonstrates that EPI cardiac tissues respond to injury differently than epicardium-free controls, highlighting the importance of including EPIs in heart-on-a-chip constructs that aim to accurately mimic the cardiac environment.

Remodeling of Ventricular Catecholaminergic Axons Following Chronic Intermittent Hypoxia in Mice

Chronic intermittent hypoxia (CIH) is a widely employed model for sleep apnea. The imbalance of autonomic control may contribute to CIH-induced cardiovascular diseases. Previously, we demonstrated that CIH increases sympathetic innervation of the atria. In this study, we determined whether CIH remodels peripheral cardiac tyrosine hydroxylase (TH, a marker for sympathetic efferent axons) innervation in the whole ventricles. The two challenges here were: 1) labeling the entire TH-IR axon innervation in the full thickness (~500 μm) of whole flat-mount ventricles. 2) Quantifying the topographical distribution of TH-IR axons through the thick ventricular walls at the single axon/varicosity scale. In this study, C57BL/6J mice (male, 2 months old, n=7/group) were exposed to room air (RA, 21% O2) or CIH (alternating between 21% and 5.7% O2 every 6 minutes for 10 hours/day) for 8-10 weeks. We prepared flat-mounts of the whole ventricles and septum and processed them with immunohistochemical labeling for TH. Using the Zeiss M2 Imager, confocal microscope, Zeiss Arivis Vision 4D, and Neurolucida system, we traced and digitized the TH-IR axons. We found that: 1) In RA and CIH mice, several large TH-immunoreactive (TH-IR) bundles from the base area entered the ventricles and septum. These large bundles branched into numerous smaller bundles as they traveled downward toward the apex and finally developed into fine varicose axons and terminals in the epicardial and myocardial layers. 2) Using Vision 4D, we were able to accurately detect, trace, and digitize all TH-IR axons in the ventricle walls with high accuracy. 3) CIH increased overall TH-IR axon density and network complexity of the whole ventricles and septum. 3) Abnormal CIH TH-IR axon terminal structures were frequently seen. 4) We geometrically and precisely registered the tracing data into a scaffold map for presentation and comparison in 3D. Altogether, CIH increased sympathetic innervation of the heart which may contribute to the augmented sympathetic drive. Our success in mapping data in the 3D heart scaffold will establish a heart-brain connectome/atlas. Supported by NIH 1 U01 NS113867-01 NIH R15 1R15HL137143-01A1, 2R01HL137832-05 and Carl Zeiss. This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

Ventricular Epicardial Adipose Distribution on Human Hearts: 3-Dimensional Reconstructions and Quantitative Assessments.

Epicardial interventions have forged new frontiers in cardiac ablation and device therapies. Healthy human hearts typically present with significant adipose tissue layers superficial to the ventricular myocardium and may hinder success or increase the complexities of epicardial interventions. We quantitatively evaluated the distribution of epicardial adipose tissue on the surface of human hearts and provided high-fidelity 3-dimensional reconstructions of these epicardial adipose tissue layers. The regional thickness of adipose tissues was analyzed at 51 anatomical reference points surrounding both ventricles and compared to specific patient demographics. Adipose deposits on the human hearts displayed characteristic patterns, with the thickest accumulations along the interventricular septa (anterior, 9.01 ± 0.50mm; posterior, 6.78 ± 0.50mm) and the right ventricular margin (7.44 ± 0.57mm). We provide one of the most complete characterizations of human epicardial adipose location and relative layer thickness. These results are considered fundamental for an underlying anatomic understanding when performing procedures within the pericardial space.

A Novel Approach to Cardiac Magnetic Resonance Scar Characterization in Patients Affected by Cardiac Amyloidosis: A Pilot Study.

Background and Objectives: Cardiac magnetic resonance (CMR) imaging has become an essential instrument in the study of cardiomyopathies; it has recently been integrated into the diagnostic workflow for cardiac amyloidosis (CA) with remarkable results. An additional emerging role is the stratification of the arrhythmogenic risk by scar analysis and the possibility of merging these data with electro-anatomical maps. This is made possible by using a software (ADAS 3D, Galgo Medical, Barcelona, Spain) able to provide 3D heart models by detecting fibrosis along the whole thickness of the myocardial walls. Little is known regarding the applications of this software in the wide spectrum of cardiomyopathies and the potential benefits have yet to be discovered. In this study, we tried to apply the ADAS 3D in the context of CA. Materials and Methods: This study was a retrospectively analysis of consecutive CMR imaging of patients affected by CA that were treated in our center (Marche University Hospital). Wherever possible, the data were processed with the ADAS 3D software and analyzed for a correlation between the morphometric parameters and follow-up events. The outcome was a composite of all-cause mortality, unplanned cardiovascular hospitalizations, sustained ventricular arrhythmias (VAs), permanent reduction in left ventricular ejection fraction, and pacemaker implantation. The secondary outcomes were the need for a pacemaker implantation and sustained VAs. Results: A total of 14 patients were deemed eligible for the software analysis: 8 patients with wild type transthyretin CA, 5 with light chain CA, and 1 with transthyretin hereditary CA. The vast majority of imaging features was not related to the composite outcome, but atrial wall thickening displayed a significant association with both the primary (p = 0.003) and the secondary outcome of pacemaker implantation (p = 0.003). The software was able to differentiate between core zones and border zones of scars, with the latter being the most extensively represented in all patients. Interestingly, in a huge percentage of CMR images, the software identified the highest degree of core zone fibrosis among the epicardial layers and, in those patients, we found a higher incidence of the primary outcome, without reaching statistical significance (p = 0.18). Channels were found in the scar zones in a substantial percentage of patients without a clear correlation with follow-up events. Conclusions: CMR imaging plays a pivotal role in cardiovascular diagnostics. Our analysis shows the feasibility and applicability of such instrument for all types of CA. We could not only differentiate between different layers of scars, but we were also able to identify the presence of fibrosis channels among the different scar zones. None of the data derived from the ADAS 3D software seemed to be related to cardiac events in the follow-up, but this might be imputable to the restricted number of patients enrolled in the study.

Lower Limit of Normality of Segmental Multilayer Longitudinal Strain in Healthy Adult Subjects.

Speckle tracking echocardiography is an advanced imaging technique that allows for a more detailed assessment of cardiac global and regional function. Reference values for segmental longitudinal layered strain (subendocardial, mid-myocardial, and subepicardial) are scarce, limiting the clinical use of these measurements in clinical practice. Two hundred consecutive Caucasian healthy subjects (mean age = 37 ± 11 years) were enrolled in the study. The mean values of global longitudinal strain (GLS) for endocardial (Endo), mid-myocardial (Myo) and epicardial (Epi) layers were -22.9 ± 2.7, -20.0 ± 2.4 and -17.5 ± 2.1, respectively. The GLSEndo/GLSMyo ratio was 1.1 ± 0.05, while the GLSEndo/GLSEpi ratio was 1.3 ± 0.05. The apical strain-sparing ratio was >1 in 10% of the subjects (endocardium) and 7% (mid-myocardium). The lower limits for segmental LS were as follows: for endocardial LS, -10% (basal), -12% (mid), -14% (apical); for mid-myocardial LS, -10% -10% (basal), -10% (mid), -10% (apical); and for epicardial LS, -7% (basal), -8% (mid), -8% (apical). The findings of this study provide data regarding the lower limit of normality of LS for each LV segment and suggest, for practical considerations, that an LS value below 10% should be considered abnormal in any segment. Further larger studies are warranted to confirm these findings.

Autophagy Activity in Epicardial Cells in Acute Pericarditis

Pericarditis is a group of polyetiological diseases often associated with emergence of life–threatening conditions. Poor knowledge of underlying cellular mechanisms and lack of relevant approaches to investigation of pericarditis result in major challenges in diagnosis and treatment.The aim of this work was to identify changes in the activity of autophagy in epicardial cells in acute pericarditis.Materials and methods. Acute pericarditis in mice was induced by intrapericardial injection of Freund's adjuvant in the study group (n=15). The control group included animals receiving either intrapericardial injection of phosphate-buffered saline (PBS) (n=15), or sham surgery without injections (n=7). On Days 3 or 5 after surgery the animals were euthanized under isoflurane anesthesia. Immunofluorescence staining of cardiac tissue cryo-sections and immunoblotting were used to assess the intensity of inflammation and autophagy in the epicardium.Results. Inflammation and other signs of acute pericarditis resulting in thickening of some epicardial areas were found: 68+9% in the control (after PBS injection) and 124+22% after Freund's adjuvant injection (p=0.009); other signs included cellular infiltration of epicardium and multiple adhesions. The epicardial layer exhibited signs of mesothelial cells reorganization with 11-fold increase of autophagy markers LC3 II/LC3 I ratio: 0.07+0.02 in the control group (after PBS injection) and 0.84+0.07 - in acute pericarditis (p=0.04), and accumulation of collagen fibers.Conclusion. Development of acute pericarditis is accompanied by activation of epicardial mesothelial cells, intensified autophagy and development of fibrous changes in epicacardial/ subepicardial areas.

Single-cell RNA sequencing analysis of the human fetal epicardium results in novel epicardial-specific markers and reveals a role for CRIP1 in epicardial EMT

Abstract Background The outside mesothelial layer of the heart, the epicardium, is essential during cardiac development. Cells from this layer undergo epithelial-to-mesenchymal transition (EMT), migrate into the tissue and partake in cardiogenesis by contributing cells and by secretion of growth factors. In the adult heart, the epicardium is quiescent but upon ischemic injury it is activated and conveys a regenerative response. This activation is important for repair, but suboptimal compared to the cardiogenic potential during development. Understanding the processes and cells that underlie developmental epicardial activation could contribute to enhancing the adult post-injury response, but little is known about the human developing epicardium. The Purpose of this study is to unravel the cellular composition of the human fetal epicardial layer and to identify novel processes that regulate the activation of the epicardium. Methods An epicardial-cell enriched dataset was obtained by specifically dissecting the epicardial layers from 4 fetal human hearts (14-15 weeks gestation, obtained under informed consent and according to local ethical approval). Single live cells were sorted into 384-wells plates and sequenced. Data analysis was performed using R-packages RaceID3 and StemID2. Findings were validated using qPCR and immunohistochemistry on tissue sections and cell culture models. Results Analysis of 2073 cells revealed a clustering of epicardial (epithelial, pre-EMT) and epicardial-derived (mesenchymal, post-EMT) cell populations. Since classical markers for the epicardium such as Wilms’ Tumor 1 and T-box transcription factor 18 were unable to specifically demarcate the epithelial state, we identified and validated several new markers (e.g. C3, NRK) that allow robust classification of epicardium-associated populations. To establish the regulation of epicardial activation and EMT, we applied pseudo-time analysis. We selected genes that were highly regulated (&amp;gt;2 fold change) in this trajectory for further analysis in vitro. qPCR indicated that CRIP1 is expressed in epithelial cells and rapidly downregulated upon EMT (3x reduction, n=3, *p=0.0104). The expression profile of CRIP1 positively correlates with UPK3B (R2:0.98) and negatively with mesenchymal TCF21 (R2: 0.99), together with GO-terms a role for CRIP in maintaining the epithelial-state is likely. Indeed, siRNA experiments in vitro with cultured epicardial cells confirmed an essential role for CRIP1 in EMT regulation. Together with its confirmed protein expression within the epicardial layer in fetal and adult tissue, this protein is of interest for regulating activation of the epicardium. Conclusions We shed light on the composition of the human fetal epicardium using a highly enriched scRNA dataset. We identify specific markers for different cell-states, and reveal novel regulators of activation that in the future may be influenced to optimise the post-injury response.

Synchrotron-based X-ray phase contrast imaging of transmural cardiac tissue in patients treated for advanced heart failure

Abstract Background Cardiac imaging is essential for identifying structural changes in advanced heart failure (HF) and understanding underlying pathophysiology. Synchrotron radiation-based X-ray phase contrast imaging (X-PCI) is a non-destructive imaging modality that can provide high resolution three-dimensional (3D) visualisation of cardiac tissue on the microstructural level. Thus far, such analyses including the quantification of myocyte aggregates ranging from the epicardium to the endocardium, have been confined to animal models and human fetal tissue. We aimed to explore the feasibility of using X-PCI to quantify and compare structural organisation and orientation of myocyte aggregates in the myocardium across different advanced HF aetiologies. Methods and design Four adult patients were included - two receiving a left ventricular assist device (LVAD), and two undergoing heart transplantation (HTx). Aetiology of advanced HF in both the LVAD and HTx group was ischaemic heart disease and dilated cardiomyopathy (DCM), respectively. Transmural tissue samples were obtained by left ventricular apical coring (LVAD) and from the explanted hearts (HTx). The tissue specimens were imaged by X-PCI at the TOMCAT beamline using an effective pixel size of 5.8 µm. Orientation of aggregates of cardiomyocytes was assessed using a structure tensor-based method and morphological parameters were derived - the helical angle (HA), or the angle between the circumferential left ventricular plane and myocyte aggregates, and fractional anisotropy (FA), the degree of anisotropy or disorganisation of the local myocardium. Results The general patient characteristics are shown in Table 1. X-PCI enabled 3D visualisation of transmural myocardial tissue samples showing differences in organisation of myocardial structure in different HF aetiologies (Figure 1). The epicardial adipose layer was thicker in DCM, paired with a thinner epicardial myocyte layer (yellow arrows). Furthermore, the epi-to-endocardial HA transition was clearly visualized in ICM (double arrows), whereas there was clear disruption of the HA gradient in DCM samples – a finding further quantified via FA (green arrows). Conclusion X-PCI allows non-destructive advanced imaging of ex-vivo tissue. By harnessing the power of digital 3D imaging on a cellular level, it provides a novel insight to differential information on tissue organisation in different aetiologies of advanced heart failure.Table 1Figure 1

Role and prognostic relevance of cardiac magnetic resonance imaging in the assessment of left ventricular non-compaction cardiomyopathy: review and meta-analysis

Abstract Background Left ventricular non-compaction (LVNC) is a heterogeneous myocardial disorder characterized by prominent myocardial trabeculation with deep inter-trabecular recesses, thin epicardial layer of compacted myocardium and extensive non-compacted inner muscular layer. Although several diagnostic criteria have been proposed, prediction of major adverse cardiovascular and cerebrovascular events (MACCE) still remains challenging in patients with LVNC. Objectives Objectives of our study were to perform systemic review and meta-analysis of published literature evaluating role and prognostic significance of the late gadolinium enhancement (LGE) in cardiac magnetic resonance imaging (CMR) study, its association with MACCE and characteristics of CMR derived feature-tracking (FT) strain analysis in patients with LVNC. Methods The objectives, literature search strategies, inclusion and exclusion criteria, outcome measurements, and methods of statistical analysis were analyzed according to an established protocol of the cochrane collaboration steps and meta-analysis of observational studies in epidemiology recommendations (MOOSE). Case control studies with evaluated prevalence of LGE in LVNC cardiomyopathy patients with and without MACCE and studies comparing FT strain: left ventricular global longitudinal (LV GLS), global circumferential (GCS) and global radial (GRS) strain between patients with and without LVNC has been included in the analysis. Results A total of 936 papers were collected according to our searching criteria, of those 9 studies including 416 LVNC cardiomyopathy patients with MACCE and 673 LVNC patients without MACCE and with evaluated LGE (including three studies with LVNC and heart failure), also 6 studies including 267 LVNC patients and 283 controls with evaluated FT LV GLS, GCS and GRS (measured in four studies) were selected. LGE was more frequently observed in LVNC patients with MACCE as compared to those without MACCE [RR (95 %CI) 2.52 (1.63 to 3.88); p&amp;lt;0.00001] (Fig.1). FT LV GLS was significantly more affected in LVNC patients compared to controls (St. mean difference (95 %CI): 1.40 (0.69 to 2.11); p&amp;lt;0.00001], as well as GCS and GRS (Fig. 2). Conclusions In patients with LVNC evaluated with CMR, LGE shows association with adverse cardiovascular and cerebrovascular events. CMR deformation indices: FT LV GLS, GCS, and GRS are significantly affected in patients with LVNC as compared to controls. This new CMR technique may be a complementary tool for timely detection and differentiation of LVNC cardiomyopathy and to avoid life threatening adverse events.Figure 1Figure 2

How intramyocardial fat can alter the electric field distribution during Pulsed Field Ablation (PFA): Qualitative findings from computer modeling.

Even though the preliminary experimental data suggests that cardiac Pulsed Field Ablation (PFA) could be superior to radiofrequency ablation (RFA) in terms of being able to ablate the viable myocardium separated from the catheter by collagen and fat, as yet there is no formal physical-based analysis that describes the process by which fat can affect the electric field distribution. Our objective was thus to determine the electrical impact of intramyocardial fat during PFA by means of computer modeling. Computer models were built considering a PFA 3.5-mm blunt-tip catheter in contact with a 7-mm ventricular wall (with and without a scar) and a 2-mm epicardial fat layer. High voltage was set to obtain delivered currents of 19, 22 and 25 A. An electric field value of 1000 V/cm was considered as the lethal threshold. We found that the presence of fibrotic tissue in the scar seems to have a similar impact on the electric field distribution and lesion size to that of healthy myocardium only. However, intramyocardial fat considerably alters the electrical field distribution and the resulting lesion shape. The electric field tends to peak in zones with fat, even away from the ablation electrode, so that 'cold points' (i.e. low electric fields) appear around the fat at the current entry and exit points, while 'hot points' (high electric fields) occur in the lateral areas of the fat zones. The results show that intramyocardial fat can alter the electric field distribution and lesion size during PFA due to its much lower electrical conductivity than that of myocardium and fibrotic tissue.