Subendocardial Region Research Articles

The smooth-walled human RVOT contains trabeculations that cause conduction delay

Abstract Funding Acknowledgements None. Background The right ventricular outflow tract (RVOT) is the outlet from the right ventricle and is the initiating substrate of life-threatening arrhythmias. While the luminal wall of the RVOT is often assumed to be without the complex trabecular meshwork that characterizes the right ventricular free wall, the anatomy of the RVOT is an understudied subject. Aim to investigate whether trabeculations occur in the RVOT and to assess whether this impacts electrical propagation. Methods Human hearts were investigated using high-resolution MRI, histology, RNA-sequencing, and optical and electrical mapping. Results We used high-resolution MRI and serial sectioning to reconstruct the macroscopic details of the human RVOT and identified cases exhibiting much trabeculation. The smooth lumen of the RVOT varied between 9% and 23% of the total RV anterior surface (N=11). Histological analysis on additional six hearts indicated that the RVOT compact layer is thinner when trabeculations are present. RNA sequencing of four human donor hearts revealed enrichment in the subendocardial region of 88 genes associated with cardiac conduction and trabeculations (P adjusted<0.05). Finally, we selected two human donor hearts showing trabeculations in the RVOT from which we generated wedge preparation and performed optical and electrical mapping. The trabecular regions demonstrated high degree of fractionation when compared to non-trabeculated regions, which coincided with delayed activation. Conclusion Trabeculations are found in the RVOT, and their extent varies among individuals. This impacts on the thickness of the compact wall in the RVOT, restricting the depth of tissue at which clinical interventions can be performed, as well as influencing electrical propagation and possible arrhythmogenicity.

Myocardial CD34+ Stromal Cells/Telocytes Demonstrate a Dynamic Pattern of Interactions with CD68+ Macrophages During Post-Myocardial Infarction Healing

Introduction: Myocardial CD34-positive (+) stromal cells/telocytes (SC/TCs) have been recently recognized as a novel type of resident cells involved in post-myocardial infarction (MI) repair. Alternatively, CD68-positive (+) macrophages are well-known constituents in this process. However, the relationship between these two cell populations remains mostly obscure. Therefore, this study was aimed to determine whether resident CD34+ SC/TCs and phagocytic CD68+ macrophages have any evident pattern of interactions during the inflammatory and proliferative phases of post-MI scar formation. Methods: A large transmural MI was induced in middle-aged male Sprague-Dawley rats (n = 15) under ketamine and xylazine anesthesia by permanent ligation of the left anterior descending coronary artery. The post-MI rats were infused with BrdU (5-bromo-2'-deoxyuridine) in a dose of 12.5 mg/kg/day for 72 hours via intraperitoneal osmotic minipumps on day 0, 4, or 11 after surgery. The rats were euthanized on day 3, 7 and 14 after MI, and their hearts were processed for histology and immunostaining. Results: On the third day after MI, CD34+ SC/TCs and CD68+ macrophages were absent within the core of the necrotic tissue, whereas both types of cells were juxtaposed in the space surrounding the area of coagulative necrosis. Furthermore, the presence of BrdU labeling in the nuclei of numerous CD34+ SC/TCs suggests their proliferation in the region of the active macrophage clearing. On the seventh day after MI, CD68+ macrophages were observed to have had advanced toward the center of the necrotic region leaving behind the area occupied primarily with acellular basement membranes from the resorbed CMs, whereas most of CD34+ SC/TCs trailed slightly behind. In part, such delay in centripetal progression of CD34+ SC/TCs through the areas of resorption could be attributed to the fact that these cells, in contrast to macrophages, often migrated along the acellular basement membranes left from the dead CMs that required phagocytic removal of necrotic debris to occur first. On the fourteenth day after MI, the population of CD68+ cells appeared notably reduced and demonstrated a patchy distribution predominantly along the remnants of the necrotic tissue. Meanwhile, the substantially expanded population of CD34+ SC/TCs were observed throughout the entire transmural scar, excluding the areas containing dead “mummified” CMs, aggregates of myofibroblasts and the fibroelastic thickening in subepicardial and subendocardial regions. Conclusion: Our findings, for the first time, demonstrate that populations of resident CD34+ SC/TCs and phagocytic CD68+ macrophages exhibit a dynamic pattern of interactions during early phases of post-MI scar formation, suggesting their importance in the process of post-ischemic myocardial healing. Supported by Camden Health Research Initiative. 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.

Alteration of endothelial permeability ensures cardiomyocyte survival from ischemic insult in the subendocardium of the heart.

Previous studies have shown that cardiomyocytes in the subendocardial region of myocardium survive from ischemic insult. This study was undertaken to explore possible mechanisms for the survival of these cardiomyocytes, focusing on changes in endothelial cells (ECs) and blood supply. C57/B6 mice were subjected to permanent ligation of left anterior descending (LAD) coronary artery to induce myocardial ischemia (MI). The hearts were harvested at 1, 4, and 7 days post MI and examined for histological changes. It was found that the survival of cardiomyocytes was associated with a preservation of ECs in the subendocardial region, as revealed by EC-specific tdTomato expression transgenic mice (Tie2tdTomato). However, the EC selective proteins, PECAM1 and VEGFR2, were significantly depressed in these ECs. Consequently, the ratio of PECAM1/tdTomato was significantly decreased, indicating a transformation from PECAM1+ ECs to PECAM1- ECs. Furthermore, EC junction protein, VE-cadherin, was not only depressed but also disassociated from PECAM1 in the same region. These changes led to an increase in EC permeability, as evidenced by increased blood infiltration in the subendocardial region. Thus, the increase in the permeability of ECs due to their transformation in the subendocardial region allows blood infiltration, creating a unique microenvironment and ensuring the survival of cardiomyocytes under ischemic conditions.

A unique perfusion pattern with 15O-water PET as a risk marker for arrhythmias in hypertrophic cardiomyopathy

Abstract Funding Acknowledgements Type of funding sources: Foundation. Main funding source(s): Swedish Heart Lung Foundation. Background Myocardial ischemia in hypertrophic cardiomyopathy (HCM) is associated with an elevated risk of ventricular arrhythmia and sudden cardiac death. Fibrosis often occurs in subendocardial regions of the LV and the RV insertion in the interventricular septum. Globally lowered hyperemic myocardial blood flow (MBF) by 15O-water PET is known to indicate poor prognosis, but is mainly linked to late-stage "burn-out". Recent scanner and image analysis developments allow detailed 3D visualisation of quantitative biventricular 15O-water MBF. Purpose To investigate whether there is a visual myocardial perfusion pattern with 15O-water PET in patients with HCM which could indicate an increased risk of arrhythmias. Methods We reanalysed rest/adenosine stress 15O-water PET in 24 patients with high-risk HCM treated with implantable cardioverter-defibrillator from a previous study. Non-sustained ventricular tachycardia (NSVT) was noted in 10 patients during 1 year preceding PET. PET was used to determine the LV MBF subendocardial-subepicardial gradient (ml/g/min per mm). Parametric MBF images from 15O-water PET at rest and stress were visually analysed at a segmental level. The myocardial segment which visually had both the highest endocardial MBF at rest and the lowest endocardial MBF at stress was selected as the most affected segment for each patient. Results Reduced resting MBF with partial normalisation at stress was seen in the RV insertion in 15 patients, including all NSVT subjects. Sixteen patients, including all NSVT subjects, had a visual perfusion pattern in the septal wall or apex with enhanced subendocardial MBF at rest and reduced MBF at stress in the same segment. The MBF gradient at stress was 0.04±0.01 ml/g/min per mm for the patients with NSVT and 0.07±0.03 ml/g/min per mm for the patients without NSVT in the visually most affected segments (p = 0.01). Conclusions Using 3D visualisation of quantitative MBF we noted a distinct pattern with intramural redistribution of perfusion between rest and stress in HCM, significantly associated with NSVT. Reduced resting perfusion at RV insertion points has not been reported in other cardiomyopathies. Enhanced subendocardial resting perfusion in ischemic segments might indicate reactive postischemic hyperemia. Clinical use of 15O-water PET to predict ventricular arrhythmia requires prospective trials.

Computed tomography imaging-identified location and electrocardiographic characteristics of left bundle branch area pacing in bradycardia patients.

Left bundle branch area pacing (LBBAP) is a novel physiological pacing modality. The relationship between the pacing lead tip location and paced electrocardiographic (ECG) characteristics remains unclear. The objectives are to determine the lead tip location within the interventricular septum (IVS) and assess the location-based ECG QRS duration (QRSd) and left ventricular activation time (LVAT). This multicenter study enrolled 50 consecutive bradycardia patients who met pacemaker therapy guidelines and received LBBAP implantation via the trans-ventricular septal approach. After at least 3 months postimplant, 12-lead ECGs and pacing parameters were obtained. Cardiac computed tomography (CT) imaging was performed to assess the LBBAP lead tip distance from the LV blood pool. Among the 50 patients, analyzable CT images were obtained in 42. In 23 of the 42 patients, the lead tips were within 2 mm to the LV blood pool (the LV subendocardial (LVSE) group), 13 between 2and 4 mm (the Near-LVSE group), and the remaining 6 beyond 4 mm (the Mid-LV septal (Mid-LVS) group). No significant differences in paced QRSd were found among the three groups (LVSE, 107 ± 15 ms; Near-LVSE, 106 ± 13 ms; Mid-LVS, 104 ± 15 ms; p = .87). LVAT in the LVSE (64 ± 7 ms) was significantly shorter than in the Mid-LVS (72 ± 8 ms; p < .05), but not significantly different from that in the Near-LVSE (69 ± 8 ms; p > .05). In routine LBBAP practice, paced narrow QRSd and fast LVAT, indicative of physiological pacing, were consistently achieved for lead tip location in the LV subendocardial or near LV subendocardial region.

Transmural Distribution of Coronary Perfusion and Myocardial Work Density Due to Alterations in Ventricular Loading, Geometry and Contractility.

Myocardial supply changes to accommodate the variation of myocardial demand across the heart wall to maintain normal cardiac function. A computational framework that couples the systemic circulation of a left ventricular (LV) finite element model and coronary perfusion in a closed loop is developed to investigate the transmural distribution of the myocardial demand (work density) and supply (perfusion) ratio. Calibrated and validated against measurements of LV mechanics and coronary perfusion, the model is applied to investigate changes in the transmural distribution of passive coronary perfusion, myocardial work density, and their ratio in response to changes in LV contractility, preload, afterload, wall thickness, and cavity volume. The model predicts the following: (1) Total passive coronary flow varies from a minimum value at the endocardium to a maximum value at the epicardium transmurally that is consistent with the transmural distribution of IMP; (2) Total passive coronary flow at different transmural locations is increased with an increase in either contractility, afterload, or preload of the LV, whereas is reduced with an increase in wall thickness or cavity volume; (3) Myocardial work density at different transmural locations is increased transmurally with an increase in either contractility, afterload, preload or cavity volume of the LV, but is reduced with an increase in wall thickness; (4) Myocardial work density-perfusion mismatch ratio at different transmural locations is increased with an increase in contractility, preload, wall thickness or cavity volume of the LV, and the ratio is higher at the endocardium than the epicardium. These results suggest that an increase in either contractility, preload, wall thickness, or cavity volume of the LV can increase the vulnerability of the subendocardial region to ischemia.

Phenotyping hypertrophic cardiomyopathy using cardiac diffusion magnetic resonance imaging: the relationship between microvascular dysfunction and microstructural changes.

AimsMicrovascular dysfunction in hypertrophic cardiomyopathy (HCM) is predictive of clinical decline, however underlying mechanisms remain unclear. Cardiac diffusion tensor imaging (cDTI) allows in vivo characterization of myocardial microstructure by quantifying mean diffusivity (MD), fractional anisotropy (FA) of diffusion, and secondary eigenvector angle (E2A). In this cardiac magnetic resonance (CMR) study, we examine associations between perfusion and cDTI parameters to understand the sequence of pathophysiology and the interrelation between vascular function and underlying microstructure.Methods and resultsTwenty HCM patients underwent 3.0T CMR which included: spin-echo cDTI, adenosine stress and rest perfusion mapping, cine-imaging, and late gadolinium enhancement (LGE). Ten controls underwent cDTI. Myocardial perfusion reserve (MPR), MD, FA, E2A, and wall thickness were calculated per segment and further divided into subendocardial (inner 50%) and subepicardial (outer 50%) regions. Segments with wall thickness ≤11 mm, MPR ≥2.2, and no visual LGE were classified as ‘normal’. Compared to controls, ‘normal’ HCM segments had increased MD (1.61 ± 0.09 vs. 1.46 ± 0.07 × 10−3 mm2/s, P = 0.02), increased E2A (60 ± 9° vs. 38 ± 12°, P < 0.001), and decreased FA (0.29 ± 0.04 vs. 0.35 ± 0.02, P = 0.002). Across all HCM segments, subendocardial regions had higher MD and lower MPR than subepicardial (MDendo 1.61 ± 0.08 × 10−3 mm2/s vs. MDepi 1.56 ± 0.18 × 10−3 mm2/s, P = 0.003, MPRendo 1.85 ± 0.83, MPRepi 2.28 ± 0.87, P < 0.0001).ConclusionIn HCM patients, even in segments with normal wall thickness, normal perfusion, and no scar, diffusion is more isotropic than in controls, suggesting the presence of underlying cardiomyocyte disarray. Increased E2A suggests the myocardial sheetlets adopt hypercontracted angulation in systole. Increased MD, most notably in the subendocardium, is suggestive of regional remodelling which may explain the reduced subendocardial blood flow.

Intraoperative echocardiography for electrode implantation into the left bundle branch region in patients with indications for cardiac resynchronization therapy

Abstract Background Left bundle branch (LBB) pacing recently became a novel pacing modality for cardiac resynchronization therapy (CRT). Purpose We evaluated the feasibility and utility of intraoperative Echocardiography (TTE) for implantation Select Secure pacing lead 3830 through interventricular septum (IVS) into the LBB area. Methods 60 patients with LBB block and indications for CRT in the period 2019–2020 underwent electrode implantation into the LBB area. Immediately before implantation, the IVS thickness and density (color scale ZOOM function, Siemens SC 2000 Prime) in the basal and middle region of IVS was assessed. White color corresponds to areas with a large amount of fibrosis, gray – conditionally viable myocardium. The implantation of the electrode by screwing it clockwise was carried out under constant echocardiographic control of the electrode advance from the right ventricle perpendicular to the IVS to the left ventricular (LV) endocardium. The ultimate target was the LV subendocardial region. After reaching the end position, the duration of mechanical LV systole was assessed (Velocity Vector 2D Speckle Analysis). Opposite wall delay (OWD) &amp;lt;120 ms – considered as a marker of good mechanical resynchronization. Results Technically, the procedure was successful in 54 from 60 patients (90%). The average IVS thickness at the site of successful implantation was 11±3, the maximum thickness was 18 mm. Average number of attempts to screw in the electrode in IVS per patient – 4.2±2.1. Predictors of failure were: 2 – IVS thickness over 15 mm throughout entire length and in 4 cases – total IVS fibrosis, despite the normal thickness (9±3). In two cases IVS perforation was detected, corrected by immediate electrode repositioning. In 4 (6.6%) cases, the initial point of implantation was not effective: OWD &amp;gt;120 ms, which required reimplantation of the lead in a more optimal position. Conclusion The use of intraoperative echocardiography during electrode implantation in the IVS allows choosing an optimal site for successful implantation, reliably monitoring the electrode position during implantation, and assessing LV intraventricular asynchronism and effectiveness of resynchronization Funding Acknowledgement Type of funding sources: None.

Extracellular matrix remodeling is associated with the survival of cardiomyocytes in the subendocardial region of the ischemic myocardium.

A significant amount of cardiomyocytes in subendocardial region survive from ischemic insults. In order to understand the mechanism by which these cardiomyocytes survive, the present study was undertaken to examine changes in these surviving cardiomyocytes and their extracellular matrix. Male C57BL/6 mice aged 8-12 weeks old were subjected to a permanent left anterior descending coronary artery ligation to induce ischemic injury. The hearts were collected at 1, 4, 7, or 28 days after the surgery and examined by histology. At day 1 after left anterior descending ligation, there was a significant loss of cardiomyocytes through apoptosis, but a proportion of cardiomyocytes were surviving in the subendocardial region. The surviving cardiomyocytes were gradually changed from rod-shaped to round-shaped, and appeared disconnected. Connexin 43, an important gap junction protein, was significantly decreased, and collagen I and III deposition was significantly increased in the extracellular matrix. Furthermore, lysyl oxidase, a copper-dependent amine oxidase catalyzing the cross-linking of collagens, was significantly increased in the extracellular matrix, paralleled with the surviving cardiomyocytes. Inhibition of lysyl oxidase activity reduced the number of surviving cardiomyocytes. Thus, the extracellular matrix remodeling is correlated with the deformation of cardiomyocytes, and the electrical disconnection between the surviving cardiomyocytes due to connexin 43 depletion and the increase in lysyl oxidase would help these deformed cardiomyocytes survive under ischemic conditions.

Post myocardial infarction ionic remodelling promotes repolarisation dispersion and electrocardiogram abnormalities in acute and chronic stages

Abstract Funding Acknowledgements Type of funding sources: Public grant(s) – National budget only. Main funding source(s): Wellcome Trust Background Sudden cardiac death (SCD) occurs in both acute and chronic stages post myocardial infarction (MI), and the left ventricular ejection fraction (LVEF) and electrocardiogram (ECG) characteristics are important biomarkers for clinical decision making. While various ionic remodelling has been reported in literature at various stages post-MI, the effect of them on clinical biomarkers have not been fully explored. Purpose Evaluate the effects of cellular ionic remodelling in acute and chronic post myocardial infarction on ECG and LVEF biomarkers through computational modelling and simulations of human biventricular electromechanics. Methods An electromechanical coupled biventricular model with ToR-ORd (Tomek 2019) human electrophysiology coupled with human excitation-contraction machinery (Land 2017) and orthotropic passive mechanics (Holzapfel 2009) was used as baseline model. The 12-lead ECG were simulated at standard body-surface electrode locations, and the biventricular pressure volume loop was simulated to quantify the simulated LVEF. A cardiac magnetic resonance (CMR) derived human biventricular geometry (Strocchi 2020) was used in combination with rigidly aligned torso geometry (Minchole 2019). An anterior sub-endocardial infarction was delineated with infarct, border zone (BZ) and remote zone (RZ) regions. For acute infarction, three types of BZ ionic remodelling from literature, and their effects on simulated LVEF and ECG characteristics were quantified. For chronic infarction, a single chronic BZ ionic remodelling was combine with two different models of RZ ionic remodelling from literature, and the effects of these remodelling on the simulated LVEF and ECG were quantified. The electromechanical properties of the sub-endocardial infarct region did not significantly affect simulated ECG or LVEF biomarkers. Results For the acute phase, simulations showed pre-cordial ECG abnormalities for all three BZ models, with T-wave inversion and QT prolongation (80 ms) in BZ1, ST-segment elevation with T-wave inversion in BZ2, and decreased T-wave amplitude in BZ3. Activation maps showed conduction block in BZ2 simulations, leading to ST-segment elevation. Repolarisation maps showed high dispersion in BZ1 simulations, leading to T-wave inversion. LVEF for BZ2 decreased by 4% (from 57% in control) due to conduction block in BZ and lack of contraction in that region, LVEF remained constant for BZ1 and BZ3 simulations. For the chronic phase, there was QT prolongation in both RZ1 (80 ms) and RZ2 (170 ms) as well as decreased T-wave amplitude, reflected by elevated repolarisation dispersion. LVEF was not significantly altered for either simulations. Conclusions Post-MI ionic remodelling at the acute and chronic stages cause varying degrees of dispersion of repolarisation and T-wave and ST-segment abnormalities, with minimal effect on mechanical function, except in the case of severe conduction abnormality. Abstract Figure.

Approaches for determining cardiac bidomain conductivity values: progress and challenges.

Modelling the electrical activity of the heart is an important tool for understanding electrical function in various diseases and conduction disorders. Clearly, for model results to be useful, it is necessary to have accurate inputs for the models, in particular the commonly used bidomain model. However, there are only three sets of four experimentally determined conductivity values for cardiac ventricular tissue and these are inconsistent, were measured around 40 years ago, often produce different results in simulations and do not fully represent the three-dimensional anisotropic nature of cardiac tissue. Despite efforts in the intervening years, difficulties associated with making the measurements and also determining the conductivities from the experimental data have not yet been overcome. In this review, we summarise what is known about the conductivity values, as well as progress to date in meeting the challenges associated with both the mathematical modelling and the experimental techniques. Graphical abstract Epicardial potential distributions, arising from a subendocardial ischaemic region, modelled using conductivity data from the indicated studies.

Approaching the forensic significance of possible PMMR correlates in a case of assumed cardiac rigor mortis

Abstract We demonstrate a left ventricular subendocardial ring as low signal intensity (SI) findings in a T2-weighted image (T2WI) sequence in postmortem magnetic resonance (PMMR) imaging. As plausible explanation, we propose rigor mortis of the heart muscle and discuss the implications of that. An 18-year-old man with a previous history of arrhythmia suddenly went into cardiopulmonary arrest while eating. Although cardiopulmonary resuscitation was performed for 120 min, death was confirmed. Autopsy revealed the cause of death to be sudden cardiac death due to arrhythmia induced by arrhythmogenic right ventricular cardiomyopathy. In this case, PMMR imaging performed before autopsy demonstrated a low SI area in the subendocardial region, suggesting a cardiac rigor mortis on the short-axial T2WI, apparent diffusion coefficient (ADC) map, and diffusion-weighted image (DWI). Low SI on T2WI is considered to be lesser interstitial fluids distribution in more-pressured subendocardial region, low SI on ADC map to be higher viscosity in subendocardial region, and low SI on DWI to be T2 blackout effect. PMMR imaging may delineate a correlate for assumed cardiac rigor mortis, which seems to be practically impossible to reliably identify macroscopically.

P1338 Anomalous left coronary artery from pulmonary artery syndrome diagnosed in adulthood: case of a mother of five

Abstract Funding Acknowledgements Project no. NVKP_16-1-2016-0017 has been implemented with the support provided from the National Research, Development and Innovation Fund of Hungary Introduction Bland-White-Garland syndrome or ALCAPA (anomalous left coronary artery from pulmonary artery) syndrome is a rare congenital disorder with a prevalence of 1: 300,000. This condition is one the most common causes of infant myocardial infarction. It can ultimately lead to heart failure before the age of one, however when pronounced left and right coronary artery collateralization is present, patients may remain asymptomatic until older age. Due to the development and availability of non-invasive imaging techniques, these patients diagnosed more frequently in recent years. Case description: We present a 53-year-old patient with treated hypertension and five uncomplicated pregnancies. At the age of 40, the patient was referred to coronary angiography due to her dyspnea on exertion. The examination showed that the left main coronary artery (LMCA) originates from the pulmonary artery 7 mm above the pulmonary valve and the right coronary artery is significantly dilated. Based on the mild complaints and good left ventricle systolic function, conservative therapy was recommended by the cardiac surgery team. Since 2018 the patient has had gradually worsening complaints despite of the therapy adjusted for chronic heart failure. Q-waves were depicted on the 12-lead ECG in aVL and V1-V6 leads. Echocardiography showed dilated left ventricle (LV), LV hypertrophy, diffuse hypokinesia, decreased LV systolic function and diastolic dysfunction. At our clinic, we performed cardiac magnetic resonance imaging (CMR) which showed moderately reduced LV ejection fraction, diffuse LV hypokinesia, increased LV volume, septal LV hypertrophy. We visualized the retrograde flow on the LMCA, the shunt volume at the anomalous artery was 1,27 L/min. The CMR also showed irreversible damage of the subendocardial region of the myocardium via late gadolinium enhancement in the area supplied by the LMCA. Since we planned to reevaluate the surgical possibilities, the collateral network with tortuous and dilated left and right coronary arteries were visualized by coronary CT angiography. The surgical team at our institute decided to perform myocardial revascularization and restoration of dual coronary artery supply. Discussion ALCAPA is usually diagnosed in infancy however, in about 10-15% of cases the disease is only recognized in adulthood. The long-term morphological and functional characteristics of the syndrome may include reversible/irreversible ischemic damage, systolic and diastolic dysfunction, mitral regurgitation, significant collateral circulation, left-to-right shunt, dilated coronary arteries. Surgical myocardial revascularization and restoration of dual coronary artery supply is recommended even in adult patients. Abstract P1338 Figure. CTA reconstruction and LGE CMR

A transmural gradient of myocardial remodeling in early-stage heart failure with preserved ejection fraction in the pig.

Heart failure with preserved ejection fraction (HFpEF) is characterized by diastolic dysfunction. This study aimed to analyze whether early HFpEF is already associated with ultrastructural alterations and whether they differ quantitatively among the layers of the left ventricular wall. HFpEF was induced in pigs by deoxy‐corticosterone acetate (DOCA) treatment along with a high‐salt/high lipid diet over 3 months and compared with weight‐matched normal pigs (n = 5 each). Samples of the left ventricle were taken and processed for light and electron microscopy. Interstitial fibrosis, subcellular composition of cardiomyocytes and mean cardiomyocyte diameter were evaluated by stereology in subendocardial, midmyocardial and subepicardial regions. DOCA enhanced the mean cardiomyocyte diameter in all locations of the ventricle wall to the same degree. The subcellular composition did not differ between the locations and was not altered by DOCA. The volume fraction of interstitium was smaller in the subendocardium of DOCA group than of control group. Within the interstitium, the volume fraction of collagen fibrils (between cardiomyocytes) was increased in the subendocardial and midmyocardial wall layers of the DOCA group but not in the subepicardial layer. Although the capillary length density and average supply area were not altered in response to DOCA in any of the wall layers, the volume fraction of blood vessels related to the interstitial space was enhanced in the subendocardium of the DOCA group but not in the other wall layers. In conclusion, cardiomyocyte changes due to DOCA were similar in subepicardial, midmyocardial and subendocardial regions but DOCA‐induced changes in the interstitium appeared to be more pronounced in the subendocardial ventricular wall layers. This suggests a pivotal role of the subendocardial interstitium in the pathogenesis of HFpEF.

Range Variability in CMR Feature Tracking Multilayer Strain across Different Stages of Heart Failure

Heart failure (HF) is associated with progressive ventricular remodeling and impaired contraction that affects distinctly various regions of the myocardium. Our study applied cardiac magnetic resonance (CMR) feature tracking (FT) to assess comparatively myocardial strain at 3 distinct levels: subendocardial (Endo-), mid (Myo-) and subepicardial (Epi-) myocardium across an extended spectrum of patients with HF. 59 patients with HF, divided into 3 subgroups as follows: preserved ejection fraction (HFpEF, N = 18), HF with mid-range ejection fraction (HFmrEF, N = 21), HF with reduced ejection fraction (HFrEF, N = 20) and a group of age- gender- matched volunteers (N = 17) were included. Using CMR FT we assessed systolic longitudinal and circumferential strain and strain-rate at Endo-, Myo- and Epi- levels. Strain values were the highest in the Endo- layer and progressively lower in the Myo- and Epi- layers respectively, this gradient was present in all the patients groups analyzed but decreased progressively in HFmrEF and further on in HFrEF groups. GLS decreased with the severity of the disease in all 3 layers: Normal > HFpEF > HFmrEF > HFrEF (Endo-: −23.0 ± 3.5 > −20.0 ± 3.3 > −16.4 ± 2.2 > −11.0 ± 3.2, p < 0.001, Myo-: −20.7 ± 2.4 > −17.5.0 ± 2.6 > −14.5 ± 2.1 > −9.6 ± 2.7, p < 0.001; Epi-: −15.7 ± 1.9 > −12.2 ± 2.1 > −10.6 ± 2.3 > −7.7 ± 2.3, p < 0.001). In contrast, GCS was not different between the Normal and HFpEF (Endo-: −34.5 ± 6.2 vs −33.9 ± 5.7, p = 0.51; Myo-: −21.9 ± 3.8 vs −21.3 ± 2.2, p = 0.39, Epi-: −11.4 ± 2.0 vs −10.9 ± 2.3, p = 0.54) but was, as well, markedly lower in the systolic heart failure groups: Normal > HFmrEF > HFrEF (Endo-: −34.5 ± 6.2 > −20.0 ± 4.2 > 12.3 ± 4.2, p < 0.001; Myo-: −21.9 ± 3.8 > −13.0 ± 3.4 > −8.0 ± 2.7. p < 0.001; Epi-: −11.4 ± 2.0 > −7.9 ± 2.3 > −4.5 ± 1.9. p < 0.001). CMR feature tracking multilayer strain assessment identifies large range differences between distinct myocardial regions. Our data emphasizes the importance of sub-endocardial myocardium for cardiac contraction and thus, its predilect role in imaging detection of functional impairment. CMR feature tracking offers a convenient, readily available, platform to evaluate myocardial contraction with excellent spatial resolution, rendering further details about discrete areas of the myocardium. Using this technique across distinct groups of patients with heart failure (HF), we demonstrate that subendocardial regions of the myocardium exhibit much higher strain values than mid-myocardium or subepicardial and are more sensitive to detect contractile impairment. We also show comparatively higher values of circumferential strain compared with longitudinal and a higher sensitivity to detect contractile impairment. A newly characterized group of patients, HF with mid-range ejection fraction (EF), shows similar traits of decompensation but has relatively higher strain values as patients with HF with reduced EF.

Cardiac sympathetic innervation network shapes the myocardium by locally controlling cardiomyocyte size through the cellular proteolytic machinery.

The heart is innervated by a dense sympathetic neuron network which, in the short term, controls chronotropy and inotropy and, in the long term, regulates cardiomyocyte size. Acute neurogenic control of heart rate is achieved locally through direct neuro-cardiac coupling at specific junctional sites (neuro-cardiac junctions). The ventricular sympathetic network topology is well-defined and characteristic for each mammalian species. In the present study, we used cell size regulation to determine whether long-term modulation of cardiac structure is achieved via direct sympatho-cardiac coupling. Local density of cardiac innervation correlated with cell size throughout the myocardial walls in all mammalian species analysed, including humans. The data obtained suggest that constitutive neurogenic control of cardiomyocyte trophism occurs through direct intercellular signalling at neuro-cardiac junctions. It is widely appreciated that sympathetic stimulation of the heart involves a sharp increase in beating rate and significant enhancement of contractility. We have previously shown that, in addition to these evident functions, sympathetic neurons (SNs) also provide trophic input to cardiomyocytes (CMs), regulating cell and organ size. More recently, we have demonstrated that cardiac neurons establish direct interactions with CMs, allowing neuro-cardiac communication to occur locally, with a 'quasi-synaptic' mechanism. Based on the evidence that cardiac SNs are unevenly distributed throughout the myocardial walls, we investigated the hypothesis that CM size distribution reflects the topology of neuronal density. In vitro analyses of SN/CM co-cultures, ex vivo confocal and multiphoton imaging in clarified hearts, and biochemical and molecular approaches were employed, in both rodent and human heart biopsies. In line with the trophic effect of SNs, and with local neuro-cardiac communication, CMs, directly contacted by SNs in co-cultures, were larger than the non-targeted ones. This property reflects the distribution of CM size throughout the ventricles of intact mouse heart, in which cells in the outer myocardial layers, which were contacted by more neuronal processes, were larger than those in the less innervated subendocardial region. Such differences disappeared upon genetic or pharmacological interference with the trophic SN/CM signalling axis. Remarkably, CM size followed the SN distribution pattern in other mammals, including humans. Our data suggest that both the acute and chronic influence of SNs on cardiac function and structure is enacted as a result of the establishment of specific intercellular neuro-cardiac junctions.

Microstructure-based finite element model of left ventricle passive inflation

Isolating the role(s) of microstructural pathological features in affecting diastolic filling is important in developing targeted treatments for heart diseases. We developed a microstructure-based constitutive model of the myocardium and implemented it in an efficient open-source finite element modeling framework to simulate passive inflation of the left ventricle (LV) in a representative 3D geometry based on experimentally measured muscle fiber architecture. The constitutive model was calibrated using previous tissue-level biaxial mechanical test data derived from the canine heart and validated with independent sets of measurements made at both the isolated constituent and organ level. Using the validated model, we investigated the load taken up by each tissue constituent and their effects on LV passive inflation. The model predicts that the LV compliance is sensitive to the collagen ultrastructure, specifically, the collagen fiber azimuthal angle with respect to the local muscle fiber direction and its waviness. The model also predicts that most of the load in the sub-epicardial and sub-endocardial regions is taken up, respectively, by the muscle fibers and collagen fiber network. This result suggests that normalizing LV passive stiffness by altering the collagen fiber network and myocyte stiffness is most effective when applied to the sub-endocardial and sub-epicardial regions, respectively. This finding may have implication for the development of new pharmaceutical treatments targeting individual cardiac tissue constituents to normalize LV filling function in heart diseases. Statement of SignificanceCurrent constitutive models describing the tissue mechanical behavior of the myocardium are largely phenomenological. While able to represent the bulk tissue mechanical behavior, these models cannot distinguish the contribution of the tissue constituents and their ultrastructure to heart function. Although microstructure-based constitutive models can be used to isolate the role of tissue ultrastructure, they have not been implemented in a computational framework that can accommodate realistic 3D organ geometry. The present study addresses these issues by developing and validating a microstructure-based computational modeling framework, which is used to investigate the role of tissue constituents and their ultrastructure in affecting heart function.

Survival of Cardiomyocytes in the Subendocardial Region in Response to Myocardial Ischemic Injury in Mice

Ischemic injury results in the death of cardiomyocytes, impairing myocardial function and eventually leading to heart failure. However, it was observed that there are cardiomyocytes, located in subendocardial region, that are often survived from ischemic injury. The present study was to explore possible mechanisms for the survival of the cardiomyocytes in this particular subendocardial location. Male C57 mice (8–12 weeks old) were divided into sham-operated control and myocardial ischemia (MI) groups. Mice in the MI group were subjected to permanent ligation of left anterior descending (LAD) coronary artery. After 1, 4 or 7 days post surgery, mice were injected intravenously with either pimonidazole to define hypoxic regions or FITC labeled wheat germ agglutinins (WGA) to trace blood infiltration. One hour after the injection, the mice were euthanatized. The hearts were collected and then cut into serial frozen sections for further analyses including cell apoptosis, blood supply and hypoxic condition. The results showed that the survived cardiomyocytes located in the subendocardial region were adjacent to the wall of the left ventricle. The TUNEL staining showed that apoptotic cells were significantly increased in the ischemic area on the 1st day after LAD ligation, but a few apoptotic cells were identified in the subendocardial region. The staining of pimonidazole showed that most of the affected myocardium was under hypoxic condition except the subendocardial region, but the CD31 positive capillaries were all disappeared in the ischemic area including the subendocardial region. The WGA staining showed a complete occlusion of blood infiltration, but there was WGA leakage in the subendocardial region. Further analysis revealed that the tight junction of the endocardium defined by VE-cadherin immunostaining was disturbed, likely responsible for WGA leakage. These results thus suggest that the breakage of the tight junction of the endocardium would lead to blood infiltration from left ventricle, which might nurture the cardiomyocytes located in the subendocardial region, leading to their survival. Support or Funding Information Supported by scientific research funding of West China Hospital Sichuan University. Morphalogical changes of cardiomyocytes after ischemic injury Cell apoptosis, hypoxia and capillaries distribution in ischemic area Endothelial junction breakage and blood infiltration from left ventricle. Morphalogical changes of cardiomyocytes after ischemic injury Cell apoptosis, hypoxia and capillaries distribution in ischemic area Endothelial junction breakage and blood infiltration from left ventricle. This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Progressive Structural Remodeling of Residual Coronary Arteries in Post‐Myocardial Infarction Scar

We have reported previously that local, patent microvascular networks support long‐term survival of adult cardiac myocytes in subendocardial and subepicardial regions of large transmural post‐myocardial infarction (MI) scars induced by left coronary artery ligation in middle‐aged rats. According to our findings, such microvascular beds become perfused retrogradely from the left ventricular chamber via Thebesian vessels linked to residual veins. However, considering that surviving cardiac myocytes may serve as an endogenous source of contractile cells in healing scars, it is vital to determine whether the original arterial bed can be salvaged as a conduit for blood reperfusion. Therefore, in the current study, we aimed to investigate structural integrity of residual coronary arteries in healing post‐MI scars. In order to achieve this goal, a large transmural MI was induced in 12‐month‐old male Sprague‐Dawley rats by permanent ligation of the left coronary artery. Post‐MI rats were euthanized 4 and 12 weeks after surgery and their hearts were processed into paraffin for histological evaluations. Eight micron transverse sections of each heart were stained with picrosirius red, Masson's trichrome and Verhoeff's elastic tissue stains. In addition, sections were labeled with TUNEL technique and an anti‐smooth muscle (SM) alpha‐actin antibody. Images of stained arteries were captured using an Olympus BX53 microscope equipped with DP72 digital camera. Furthermore, in each heart, the entire scar, stained for fibrillar collagen and SM alpha‐actin, was digitized and then analyzed with Image‐Pro Analyzer v.7.0 software to determine the fractional volumes of intravascular collagen and vascular SM cells. We found that the profiles of residual coronary arteries could be identified clearly in 4‐ and 12‐week‐old scars. Although some vessels appeared normal, most nonperfused arteries revealed gross structural alterations including accumulation of perivascular collagen and a prominent centripetal neointimal growth narrowing the vessel lumen. In the latter case, vascular SM cell hyperplasia was accompanied by a perecellular accumulation of collagen and elastic fibrils. Consistent with the progressive nature of these changes, in 12‐week‐old scars we often observed a complete obliteration of arterial lumens that occurred concurrently with disappearance of vascular SM cells and their replacement by collagenous tissue. The positive TUNEL stain in the walls of residual arteries indicates that the reduction in vascular SM cells might be related to their death via apoptosis. Importantly, our quantitative assessment of 4‐ and 12‐week‐old scars did confirm that the overall dynamic of structural changes observed in residual coronary arteries was associated with progressive accumulation of intravascular collagen (0.52±0.17% vs. 0.80±0.12%, respectively) and reduction in the content of vascular SM cells (2.27±0.51% vs. 1.53±1.19%, respectively). Therefore, our current findings provide additional support for the necessity of earlier reperfusion of the residual coronary arterial bed in healing scars in order to preserve its capacity to serve as a potential conduit for restored blood supply.This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Probing the Depth of the Myocardium: Vasculature, Transit Time, and Perfusion Within the Left Ventricular Wall

The branching architecture of arterial trees traversing the thickness of the left ventricular wall is studied to determine the way in which adequate blood supply is provided to myocardial tissue at different depths within the wall thickness from arterial trees originating at the epicardial surface. The study is based on micro-CT images of tissue biopsies, coupled with a dedicated vascular tree analysis program. The results show that this combination of methodologies allows a more detailed and much more accurate exploration of the vasculature within the sampled tissue than is possible by histological means. The spatial density of the smallest resolvable "end" arterioles is found to be higher in the sub-endocardial region than in the sub-epicardial region, with vascular branching architecture consistent with a fractal structure. The concept of "transit time" is introduced as an approximate measure of the time it takes bulk flow to reach different regions of the myocardium. Our data suggest that a transit time differential is a major contributor to the equalization of transmural perfusion gradient against unequal distribution of "end' arteriolar density.