Mechanism of Blood–Heart-Barrier Leakage: Implications for COVID-19 Induced Cardiovascular Injury

Although blood–heart-barrier (BHB) leakage is the hallmark of congestive (cardio-pulmonary) heart failure (CHF), the primary cause of death in elderly, and during viral myocarditis resulting from the novel coronavirus variants such as the severe acute respiratory syndrome novel corona virus 2 (SARS-CoV-2) known as COVID-19, the mechanism is unclear. The goal of this project is to determine the mechanism of the BHB in CHF. Endocardial endothelium (EE) is the BHB against leakage of blood from endocardium to the interstitium; however, this BHB is broken during CHF. Previous studies from our laboratory, and others have shown a robust activation of matrix metalloproteinase-9 (MMP-9) during CHF. MMP-9 degrades the connexins leading to EE dysfunction. We demonstrated juxtacrine coupling of EE with myocyte and mitochondria (Mito) but how it works still remains at large. To test whether activation of MMP-9 causes EE barrier dysfunction, we hypothesized that if that were the case then treatment with hydroxychloroquine (HCQ) could, in fact, inhibit MMP-9, and thus preserve the EE barrier/juxtacrine signaling, and synchronous endothelial-myocyte coupling. To determine this, CHF was created by aorta-vena cava fistula (AVF) employing the mouse as a model system. The sham, and AVF mice were treated with HCQ. Cardiac hypertrophy, tissue remodeling-induced mitochondrial-myocyte, and endothelial-myocyte contractions were measured. Microvascular leakage was measured using FITC-albumin conjugate. The cardiac function was measured by echocardiography (Echo). Results suggest that MMP-9 activation, endocardial endothelial leakage, endothelial-myocyte (E-M) uncoupling, dyssynchronous mitochondrial fusion-fission (Mfn2/Drp1 ratio), and mito-myocyte uncoupling in the AVF heart failure were found to be rampant; however, treatment with HCQ successfully mitigated some of the deleterious cardiac alterations during CHF. The findings have direct relevance to the gamut of cardiac manifestations, and the resultant phenotypes arising from the ongoing complications of COVID-19 in human subjects.

Enterococcal Endocarditis: Hiding in Plain Sight

Enterococcus faecalis is a major opportunistic bacterial pathogen of increasing clinical relevance. A substantial body of experimental evidence suggests that early biofilm formation plays a critical role in these infections, as well as in colonization and persistence in the GI tract as a commensal member of the microbiome in most terrestrial animals. Animal models of experimental endocarditis generally involve inducing mechanical valve damage by cardiac catheterization prior to infection, and it has long been presumed that endocarditis vegetation formation resulting from bacterial attachment to the endocardial endothelium requires some pre-existing tissue damage. Here we review both historical and contemporary animal model studies demonstrating the robust ability of E. faecalis to directly attach and form stable microcolony biofilms encased within a bacterially-derived extracellular matrix on the undamaged endovascular endothelial surface. We also discuss the morphological similarities when these biofilms form on other host tissues, including when E. faecalis colonizes the GI epithelium as a commensal member of the normal vertebrate microbiome - hiding in plain sight where it can serve as a source for systemic infection via translocation. We propose that these phenotypes may allow the organism to persist as an undetected infection in asymptomatic individuals and thus provide an infectious reservoir for later clinical endocarditis.

Proteomics Analysis Reveals Diverse Molecular Characteristics between Endocardial and Aortic-Valvular Endothelium

The variations in the protein profile of aortic-valvular (AVE) and endocardial endothelial (EE) cells are currently unknown. The current study’s objective is to identify differentially expressed proteins and associated pathways in both the endothelial cells. We used endothelial cells isolated from the porcine (Sus scrofa) aortic valve and endocardium for the profiling of proteins. Label-free proteomics was performed by liquid chromatography-tandem mass spectrometry (LC-MS/MS). Our proteomics analysis revealed that 29 proteins were highly expressed, and 25 proteins were less expressed in the valve than the endocardial endothelium. The cell surface markers, such as CD63, ICAM1, PECAM1, PROCR, and TFRC, were highly expressed in EE. In contrast, CD44 was highly expressed in AVE. The pathway analysis showed that metabolic process-related proteins and extracellular matrix-related proteins were enriched in valves. Differential enrichment of signaling pathways was observed in the endocardium. The hemostasis function-related proteins were increased in both endothelial cells. The proteins and pathways enriched in aortic-valvular and endocardial endothelial cells revealed the distinct phenotype of these two closely related cells.

Immunohistological Evaluation of Von Willebrand Factor in the Left Atrial Endocardium and Atrial Thrombi from Cats with Cardiomyopathy

Aortic thromboembolism (ATE) occurs in cats with cardiomyopathy and often results in euthanasia due to poor prognosis. However, the underlying predisposing mechanisms leading to left atrial (LA) thrombus formation are not fully characterised. von Willebrand Factor (vWF) is a marker of endothelium and shows increased expression following endothelial injury. In people with poor LA function and LA remodelling, vWF has been implicated in the development of LA thrombosis. In this study we have shown (1) the expression of endocardial vWF protein detected using immunohistofluorescence was elevated in cats with cardiomyopathy, LA enlargement (LAE) and clinical signs compared to cats with subclinical cardiomyopathy and control cats; (2) vWF was present at the periphery of microthrombi and macrothrombi within the LA where they come into contact with the LA endocardium and (3) vWF was integral to the structure of the macrothrombi retrieved from the atria. These results provide evidence for damage of the endocardial endothelium in the remodelled LA and support a role for endocardial vWF as a pro-thrombotic substrate potentially contributing to the development of ATE in cats with underlying cardiomyopathy and LAE. Results from this naturally occurring feline model may inform research into human thrombogenesis.

A Histological Study on Heart of Pelophylax cf. bedriagae (Anura: Ranidae)

A circulatory system become compulsory for the transport of nutrients and gases due to the growth of animals and the development of various organs. The heart of Pelophylax cf. bedriagae consisted of three chambers (two atria and one ventricle). Two atria were completely separated, however the ventricle were undivided. When looking dorsally on the heart, thin-walled sinus venosus was observed. The sinus venosus is responsible for initiating the heart beat and receives venous blood coming from caput, anterior legs, and body that then flows to the right atrium. Pacemaker cells and ganglion cells were main cells of the sinus venosus. The layer of the heart wall consisted of epicardium, myocardium and endocardium. The epicardium was the outer layer of heart. The myocardium was medium layer which was composed of an outer compact layer and inner trabecular layer. Ventricular trabeculae were lined by a thin layer of endocardial endothelium. The inner trabecular layer of ectothermic vertebrates is accepted as homologous to the Purkinje network. The atrioventricular valves were composed of loose connective tissue.

Endocardial endothelium as a blood-heart barrier

Introduction. Endocardial endothelium is formed from a single layer of closely related cells with complex interrelationships and extensive overlap at the junctional edges. Morphological characteristics of blood-heart barrier. Endocardium is composed of three layers: endocardial endothelium, subendothelial loose connective tissue and subendocardium. The fibrous component of the subendothelium consists of small amount of collagen and elastic fibers. Several cell types are present in subendocardium: telocytes, fibroblasts and nerve endings. Intercellular bonds between the endocardial endothelial cells. Endocardial endothelial cells are attached to one another via sets of binding proteins forming solid, adherent and communicating connections. Communicating connections form transmembrane channels between the neighboring cells, while solid and adherent connections form pericellular structures like stitches. The maintenance of the presumed transendocardial electrochemical potential difference provides a high gradient for certain ions as well as a selective boundary barrier, basal lamina, preventing ionic leakage. The negatively charged glycocalyx also modulates endothelial permeability. Electrophysiological characteristics of heart-blood barrier. Electrophysiological studies have shown the existence of a large number of membrane ion channels in the endocardial endothelial cells: inward rectifying K+ channels, Ca2+ dependent K+channels, voltage-dependent Cl-channels, volume-activated Cl-channels, stretch-activated cation channels and one carrier mediated transport mechanism - Na+K+adenosine triphosphatase. Conclusion. Numerous diseases of the cardiovascular system may be a consequence, but also the cause of the endocardial endothelium dysfunction. Selective damage to the endocardial endothelium and subendocardium is found in arrhythmia, atrial fibrillation, ischemia/reperfusion injury and heart failure. Typical lesions of endocardial and microvascular endothelium have also been described in sepsis, myocardial infarction, inflammation and thrombosis. The result of endothelial dysfunction is the weakening of the endothelial barrier regulation and electrolyte imbalance of the subendocardial interstitium.

Some Structural and Chemical Changes in Endocardial Endothelium of Rats in Emotional and Pain Stress Complicated by Hypercholesterolemia

The objective of the research was to study the content of some neutral lipids of endocardial endothelium in rats in relation to structural changes occurring in it, in the co-existence of emotional and pain stress, as well as alimentary hypercholesterolemia. Materials and methods. The electric-impulse model was used for stress modeling. Alimentary hypercholesterolemia was modeled feeding animals an atherogenic diet. The concentration of triacylglycerols, free and esterified cholesterol were examined using the method of thin-layer chromatography performed on silica gel. The concentration of free fatty acids was determined using the radiochemical method. The state of endocardial endothelium was studied with the help of light microscopy; the impression smears obtained from macro preparations of ventricle were analyzed.Results. In co-existence of stress and hypercholesterolemia, significant increase in free cholesterol as well as free fatty acid concentration was noticed. This essentially exceeded the analogical indices under the action of stress only. Structural changes in the endocardium followed by desquamation of separate endotheliocytes were the result of stress reaction. In the action of both pathogenic factors, this process was intensified; layer-by-layer exfoliation of endotheliocytes was observed. Conclusions. In acute emotional and pain stress, changes in lipid spectrum of membrane structures of endocardial endotheliocytes the main manifestation of which is the accumulation of free cholesterol in cells and increase in the levels of free fatty acids take place. The increase in the number of desquamated endothelial cells is the result of stress action as well. Alimentary hypercholesterolemia significantly increases such pathological changes.