Endothelial Surface Layer Research Articles

Hemodynamic Characteristics of a Tortuous Microvessel Using High-Fidelity Red Blood Cell Resolved Simulations.

Tortuous microvessels are characteristic of microvascular remodeling associated with numerous physiological and pathological scenarios. Three-dimensional (3D) hemodynamics in tortuous microvessels influenced by red blood cells (RBCs), however, are largely unknown, and important questions remain. Is blood viscosity influenced by vessel tortuosity? How do RBC dynamics affect wall shear stress (WSS) patterns and the near-wall cell-free layer (CFL) over a range of conditions? The objective of this work was to parameterize hemodynamic characteristics unique to a tortuous microvessel. RBC-resolved simulations were performed using an immersed boundary method-based 3D fluid dynamics solver. A representative tortuous microvessel was selected from a stimulated angiogenic network obtained from imaging of the rat mesentery and digitally reconstructed for the simulations. The representative microvessel was a venule with a diameter of approximately 20 μm. The model assumes a constant diameter along the vessel length and does not consider variations due to endothelial cell shapes or the endothelial surface layer. Microvessel tortuosity was observed to increase blood apparent viscosity compared to a straight tube by up to 26%. WSS spatial variations in high curvature regions reached 23.6 dyne/cm2 over the vessel cross-section. The magnitudes of WSS and CFL thickness variations due to tortuosity were strongly influenced by shear rate and negligibly influenced by tube hematocrit levels. New findings from this work reveal unique tortuosity-dependent hemodynamic characteristics over a range of conditions. The results provide new thought-provoking information to better understand the contribution of tortuous vessels in physiological and pathological processes and help improve reduced-order models.

Lack of Laminar Shear Stress Facilitates the Endothelial Uptake of Very Small Superparamagnetic Iron Oxide Nanoparticles by Modulating the Endothelial Surface Layer.

To study whether the absence of laminar shear stress (LSS) enables the uptake of very small superparamagnetic iron oxide nanoparticles (VSOP) in endothelial cells by altering the composition, size, and barrier function of the endothelial surface layer (ESL). A quantitative particle exclusion assay with living human umbilical endothelial cells using spinning disc confocal microscopy revealed that the dimension of the ESL was reduced in cells cultivated in the absence of LSS. By combining gene expression analysis, flow cytometry, high pressure freezing/freeze substitution immuno-transmission electron microscopy, and confocal laser scanning microscopy, we investigated changes in ESL composition. We found that increased expression of the hyaluronan receptor CD44 by absence of shear stress did not affect the uptake rate of VSOPs. We identified collagen as a previously neglected component of ESL that contributes to its barrier function. Experiments with inhibitor halofuginone and small interfering RNA (siRNA) demonstrated that suppression of collagen expression facilitates VSOP uptake in endothelial cells grown under LSS. The absence of laminar shear stress disturbs the barrier function of the ESL, facilitating membrane accessibility and endocytic uptake of VSOP. Collagen, a previously neglected component of ESL, contributes to its barrier function.

The role of glycosaminoglycans in blood pressure regulation.

Essential hypertension (HT) is the global health problem and is a major risk factor for the development of cardiovascular and kidney disease. High salt intake has been associated with HT and impaired kidney sodium excretion is considered to be a major mechanism for the development of HT. Although kidney has a very important role in regulation of BP, this traditional view of BP regulation was challenged by recent findings suggesting that nonosmotic tissue sodium deposition is very important for BP regulation. This new paradigm indicates that sodium can be stored and deposited nonosmotically in the interstitium without water retention and without increased BP. One of the major determinants of this deposition is glycosaminoglycans (GAGs). By binding to GAGs found in the endothelial surface layer (ESL) which contains glycocalyx, sodium is osmotically inactivated and not induce concurrent water retention. Thus, GAGs has important function for homeostatic BP and sodium regulation. In the current review, we summarized the role of GAGs in ESL and BP regulation.

Decreased retinal and choroidal endothelial surface molecules in spontaneously hypertensive rats

Hypertension is associated with changes in the retina and choroid, with resulting consequences of increased vascular permeability and microhemorrhages. To date, very little information is available regarding the changes in the retinal and choroidal endothelial surface layer. In this study, we have examined changes in protein expression of several molecules including platelet endothelial cell adhesion molecule-1 (PECAM-1), vascular endothelial cadherin (VE-cadherin), glypican-1, and syndecan-1, in spontaneously hypertensive rats (SHR) compared to control normotensive Wistar Kyoto (WKY) rats. In male SHR vs WKY rat retinas, decreases were found for VE-cadherin and syndecan-1; whereas in female retinas, decreases were found for PECAM-1, glypican-1, and syndecan-1. In male SHR vs WKY rat choroid, we found an increase in glypican-1, but choroidal syndecan-1 was decreased in SHR in both males and females. Therefore, decreases in SHR of both retinal and choroidal syndecan-1 were found in both males and females. These losses of syndecan-1 were accompanied by an increase in plasma levels of the proteoglycan, indicating possible systemic endothelial shedding. In contrast, plasma levels of glypican-1 decreased. Interestingly, in normotensive WKY rats, retinal levels of all four endothelial surface molecules were higher in females than in males, in some cases, by substantial amounts. In summary, a number of changes occur in endothelial surface molecules in SHR, with some changes being sex-dependent; it is possible that the loss of these molecules contributes to the vascular dysfunction that occurs in hypertensive retina and choroid.

Removal of the endothelial surface layer via hyaluronidase does not modulate monocyte and neutrophil interactions with the glomerular endothelium.

The endothelial surface layer (ESL), a layer of macromolecules on the surface of endothelial cells, can both impede and facilitate leukocyte recruitment. However, its role in monocyte and neutrophil recruitment in glomerular capillaries is unknown. We used multiphoton intravital microscopy to examine monocyte and neutrophil behavior in the glomerulus following ESL disruption with hyaluronidase. Constitutive retention and migration of monocytes and neutrophils within the glomerular microvasculature was unaltered by hyaluronidase. Consistent with this, inhibition of the hyaluronan-binding molecule CD44 also failed to modulate glomerular trafficking of these immune cells. To investigate the contribution of the ESL during acute inflammation, we induced glomerulonephritis via in situ immune complex deposition. This resulted in increases in glomerular retention of monocytes and neutrophils but did not induce marked reduction in the glomerular ESL. Furthermore, hyaluronidase treatment did not modify the prolonged retention of monocytes and neutrophils in the acutely inflamed glomerular microvasculature. These observations indicate that, despite evidence that the ESL has the capacity to inhibit leukocyte-endothelial cell interactions while also containing adhesive ligands for immune cells, neither of these functions modulate trafficking of monocytes and neutrophils in steady-state or acutely-inflamed glomeruli.

Influence of the vessel wall geometry on the wall-induced migration of red blood cells.

The geometry of the blood vessel wall plays a regulatory role on the motion of red blood cells (RBCs). The overall topography of the vessel wall depends on many features, among which the endothelial lining of the endothelial surface layer (ESL) is an important one. The endothelial lining of vessel walls presents a large surface area for exchanging materials between blood and tissues. The ESL plays a critical role in regulating vascular permeability, hindering leukocyte adhesion as well as inhibiting coagulation during inflammation. Changes in the ESL structure are believed to cause vascular hyperpermeability and entrap immune cells during sepsis, which could significantly alter the vessel wall geometry and disturb interactions between RBCs and the vessel wall, including the wall-induced migration of RBCs and the thickening of a cell-free layer. To investigate the influence of the vessel wall geometry particularly changed by the ESL under various pathological conditions, such as sepsis, on the motion of RBCs, we developed two models to represent the ESL using the immersed boundary method in two dimensions. In particular, we used simulations to study how the lift force and drag force on a RBC near the vessel wall vary with different wall thickness, spatial variation, and permeability associated with changes in the vessel wall geometry. We find that the spatial variation of the wall has a significant effect on the wall-induced migration of the RBC for a high permeability, and that the wall-induced migration is significantly inhibited as the vessel diameter is increased.

Oscillatory shear stress-mediated aberrant O-GlcNAc SIRT3 accelerates glycocalyx inflammatory injury via LKB1/p47phox/Hyal2 signaling

Glycocalyx coating on endothelial surface layer helps to sense shear forces and maintain endothelial function. However, the underlying mechanism of endothelial glycocalyx degradation upon disordered shear stress stimulation is not fully understood. SIRT3, a major NAD+-dependent protein deacetylases, is required for protein stability during vascular homeostasis and partly involved in atherosclerotic process. While few studies showed that SIRT3 is responsible for endothelial glycocalyx homeostasis under shear stress, the underlying mechanisms remain largely unknown. Here, we demonstrated that oscillatory shear stress (OSS) induces glycocalyx injury by activating LKB1/p47phox/Hyal2 axis both in vivo and in vitro. And O-GlcNAc modification served to prolong SIRT3 deacetylase activity and stabilized p47/Hyal2 complex. OSS could decrease SIRT3 O-GlcNAcylation to activate LKB1, further accelerated endothelial glycocalyx injury in inflammatory microenvironment. SIRT3Ser329 mutation or inhibition of SIRT3 O-GlcNAcylation strongly promoted glycocalyx degradation. On the contrary, overexpression of SIRT3 reverse glycocalyx damage upon OSS treatment. Together, our findings indicated that targeting O-GlcNAcylation of SIRT3 could prevent and/or treat diseases whereby glycocalyx injured.

Microvascular perfusion, perfused boundary region and glycocalyx shedding in patients with autosomal dominant polycystic kidney disease: results from the GlycoScore III study.

Vascular abnormalities and endothelial dysfunction are part of the spectrum of autosomal dominant polycystic kidney disease (ADPKD). The mechanisms behind these manifestations, including potential effects on the endothelial surface layer (ESL) and glycocalyx integrity, remain unknown. Forty-five ambulatory adult patients with ADPKD were enrolled in this prospective, observational, cross-sectional, single-centre study. Fifty-one healthy volunteers served as a control group. All participants underwent real-time microvascular perfusion measurements of the sublingual microcirculation using sidestream dark field imaging. After image acquisition, the perfused boundary region (PBR), an inverse parameter for red blood cell (RBC) penetration into the ESL, was automatically calculated. Microvascular perfusion was assessed by RBC filling and capillary density. Concentrations of circulating glycocalyx components were determined by enzyme-linked immunosorbent assay. ADPKD patients showed a significantly larger PBR compared with healthy controls (2.09±0.23µm versus 1.79±0.25µm; P<.001). This was accompanied by significantly lower RBC filling (70.4±5.0% versus 77.9±5.4%; P<.001) as well as a higher valid capillary density {318/mm2 [interquartile range (IQR) 269-380] versus 273/mm2 [230-327]; P=.007}. Significantly higher plasma concentrations of heparan sulphate (1625±807ng/ml versus 1329±316ng/ml; P=.034), hyaluronan (111ng/ml [IQR 79-132] versus 92ng/ml [82-98]; P=.042) and syndecan-1 were noted in ADPKD patients compared with healthy controls (35ng/ml [IQR 27-57] versus 29ng/ml [23-42]; P=.035). Dimensions and integrity of the ESL are impaired in ADPKD patients. Increased capillary density may be a compensatory mechanism for vascular dysfunction to ensure sufficient tissue perfusion and oxygenation.

Shedding of the Endothelial Glycocalyx Independent of Systemic Tryptase Release during Oncologic Oral Surgery: An Observational Study.

The endothelial glycocalyx and endothelial surface layer are crucial for several functions of the vasculature. Damage to the glycocalyx (“shedding”) occurs during diverse clinical conditions, including major surgery. Mast cell tryptase has been proposed as one possible “sheddase”. During oncologic oral surgery, glycocalyx shedding could be detrimental due to loss of vascular barrier function and consequent oedema in the musculocutaneous flap graft. Concentrations of the glycocalyx components heparan sulphate and syndecan-1, as well as of tryptase in blood serum before and after surgery, were measured in 16 patients undergoing oncologic oral surgery. Secondary measures were the concentrations of these substances on postoperative days 1 and 2. Heparan sulphate rose from 692 (median, interquartile range: 535–845) to 810 (638–963) ng/mL during surgery. Syndecan-1 increased from 35 (22–77) ng/mL to 138 (71–192) ng/mL. Tryptase remained virtually unchanged with 4.2 (3–5.6) before and 4.2 (2.5–5.5) ng/mL after surgery. Concentrations of heparan sulphate and syndecan-1 in serum increased during surgery, indicating glycocalyx shedding. Tryptase concentration remained equal, suggesting other sheddases than systemic tryptase release to be responsible for damage to the glycocalyx. Investigating strategies to protect the glycocalyx during oncologic oral surgery might hold potential to improve flap viability and patient outcome.

Modeling of the cerebral blood circulation in a capillary network accounting for the influence of the endothelial surface layer

Background and objectiveThe paper describes a mathematical model of blood flow in capillaries with accounting for the endothelial surface layer (ESL). MethodThe influence of ESL is modeled by a boundary layer with zero flow velocity. Finite element modeling and an analytical approach based on the homogenization of the core region of blood flow occupied by erythrocytes are developed to describe the resistance of a capillary. The reliability of the results obtained is verified for different values of the discharge hematocrit and vessel diameter using known in vivo data. ResultsThe proposed approach is applied to the numerical simulation of blood circulation in a capillary network of the germinal matrix of infants born at 25 gestational weeks. The influence of the hematocrit level and effective thickness of ESL on the resistance of the capillary network of the germinal matrix of preterm infants is studied. It was found that a decrease in the effective thickness of ESL in the capillary network (and/or a decrease in the hematocrit) leads to reducing the resistance of the capillary network. ConclusionA decrease in the effective thickness of ESL in the capillary network leads to an increase in the pressure drop in arterioles, which may be considered as an additional risk factor for hemorrhages in fragile blood vessels within the germinal matrix.

The endothelial glycocalyx in critical illness: A pediatric perspective

The vascular endothelium is the interface between circulating blood and end organs and thus has a critical role in preserving organ function. The endothelium is lined by a glycan-rich glycocalyx that uniquely contributes to endothelial function through its regulation of leukocyte and platelet interactions with the vessel wall, vascular permeability, coagulation, and vasoreactivity. Degradation of the endothelial glycocalyx can thus promote vascular dysfunction, inflammation propagation, and organ injury. The endothelial glycocalyx and its role in vascular pathophysiology has gained increasing attention over the last decade. While studies characterizing vascular glycocalyx injury and its downstream consequences in a host of adult human diseases and in animal models has burgeoned, studies evaluating glycocalyx damage in pediatric diseases are relatively few. As children have unique physiology that differs from adults, significant knowledge gaps remain in our understanding of the causes and effects of endothelial glycocalyx disintegrity in pediatric critical illness. In this narrative literature overview, we offer a unique perspective on the role of the endothelial glycocalyx in pediatric critical illness, drawing from adult and preclinical data in addition to pediatric clinical experience to elucidate how marked derangement of the endothelial surface layer may contribute to aberrant vascular biology in children. By calling attention to this nascent field, we hope to increase research efforts to address important knowledge gaps in pediatric vascular biology that may inform the development of novel therapeutic strategies.

Dependence of red blood cell dynamics in microvessel bifurcations on the endothelial surface layer's resistance to flow and compression.

Red blood cells (RBCs) make up 40-45% of blood and play an important role in oxygen transport. That transport depends on the RBC distribution throughout the body, which is highly heterogeneous. That distribution, in turn, depends on how RBCs are distributed or partitioned at diverging vessel bifurcations where blood flows from one vessel into two. Several studies have used mathematical modeling to consider RBC partitioning at such bifurcations in order to produce useful insights. These studies, however, assume that the vessel wall is a flat impenetrable homogeneous surface. While this is a good first approximation, especially for larger vessels, the vessel wall is typically coated by a flexible, porous endothelial glycocalyx or endothelial surface layer (ESL) that is on the order of 0.5-1 µm thick. To better understand the possible effects of this layer on RBC partitioning, a diverging capillary bifurcation is analyzed using a flexible, two-dimensional model. In addition, the model is also used to investigate RBC deformation and RBC penetration of the ESL region when ESL properties are varied. The RBC is represented using interconnected viscoelastic elements. Stokes flow equations (viscous flow) model the surrounding fluid. The flow in the ESL is modeled using the Brinkman approximation for porous media with a corresponding hydraulic resistivity. The ESL's resistance to compression is modeled using an osmotic pressuredifference. One cell passes through the bifurcation at a time, so there are no cell-cell interactions. A range of physiologically relevant hydraulic resistivities and osmotic pressure differences are explored. Decreasing hydraulic resistivity and/or decreasing osmotic pressure differences (ESL resistance to compression) produced four behaviors: (1) RBC partitioning nonuniformity increased slightly; (2) RBC deformation decreased; (3) RBC velocity decreased relative to blood flow velocity; and (4) RBCs penetrated more deeply into the ESL. Decreasing the ESL's resistance to flow and/or compression to pathological levels could lead to more frequent cell adhesion and clotting as well as impaired vascular regulation due to weaker ATP and nitric oxide release. Potential mechanisms that can contribute to these behaviors are also discussed.

Intravital imaging reveals glomerular capillary distension and endothelial and immune cell activation early in Alport syndrome.

Alport syndrome (AS) is a genetic disorder caused by mutations in type IV collagen that lead to defective glomerular basement membrane, glomerular filtration barrier (GFB) damage, and progressive chronic kidney disease. While the genetic basis of AS is well known, the molecular and cellular mechanistic details of disease pathogenesis have been elusive, hindering the development of mechanism-based therapies. Here, we performed intravital multiphoton imaging of the local kidney tissue microenvironment in a X-linked AS mouse model to directly visualize the major drivers of AS pathology. Severely distended glomerular capillaries and aneurysms were found accompanied by numerous microthrombi, increased glomerular endothelial surface layer (glycocalyx) and immune cell homing, GFB albumin leakage, glomerulosclerosis, and interstitial fibrosis by 5 months of age, with an intermediate phenotype at 2 months. Renal histology in mouse or patient tissues largely failed to detect capillary aberrations. Treatment of AS mice with hyaluronidase or the ACE inhibitor enalapril reduced the excess glomerular endothelial glycocalyx and blocked immune cell homing and GFB albumin leakage. This study identified central roles of glomerular mechanical forces and endothelial and immune cell activation early in AS, which could be therapeutically targeted to reduce mechanical strain and local tissue inflammation and improve kidney function.

Pulmonary surfactant and COVID-19: A new synthesis.

COVID-19 pathogenesis poses paradoxes difficult to explain with traditional physiology. For instance, since type II pneumocytes are considered the primary cellular target of SARS-CoV-2; as these produce pulmonary surfactant (PS), the possibility that insufficient PS plays a role in COVID-19 pathogenesis has been raised. However, the opposite of predicted high alveolar surface tension is found in many early COVID-19 patients: paradoxically normal lung volumes and high compliance occur, with profound hypoxemia. That 'COVID anomaly' was quickly rationalised by invoking traditional vascular mechanisms-mainly because of surprisingly preserved alveolar surface in early hypoxemic cases. However, that quick rejection of alveolar damage only occurred because the actual mechanism of gas exchange has long been presumed to be non-problematic, due to diffusion through the alveolar surface. On the contrary, we provide physical chemical evidence that gas exchange occurs by an process of expansion and contraction of the three-dimensional structures of PS and its associated proteins. This view explains anomalous observations from the level of cryo-TEM to whole individuals. It encompasses results from premature infants to the deepest diving seals. Once understood, the COVID anomaly dissolves and is straightforwardly explained as covert viral damage to the 3D structure of PS, with direct treatment implications. As a natural experiment, the SARS-CoV-2 virus itself has helped us to simplify and clarify not only the nature of dyspnea and its relationship to pulmonary compliance, but also the fine detail of the PS including such features as water channels which had heretofore been entirely unexpected. For a long time, physical, colloid and surface chemistry have not intersected with physiology and cell biology as much as we might have hoped. The reasons are starting to become clear. The discipline of physical chemistry suffered from serious unrecognised omissions that rendered it ineffective. These foundational defects included omission of specific ion molecular forces and hydration effects. The discipline lacked a predictive theory of self-assembly of lipids and proteins. Worse, theory omitted any role for dissolved gases, O2, N2, CO2, and their existence as stable nanobubbles above physiological salt concentration. Recent developments have gone some way to explaining the foam-like lung surfactant structures and function. It delivers O2/N2 as nanobubbles, and efflux of CO2, and H2O nanobubbles at the alveolar surface. Knowledge of pulmonary surfactant structure allows an explanation of the mechanism of corona virus entry, and differences in infectivity of different variants. CO2 nanobubbles, resulting from metabolism passing through the molecular frit provided by the glycocalyx of venous tissue, forms the previously unexplained foam which is the endothelial surface layer. CO2 nanobubbles turn out to be lethal to viruses, providing a plausible explanation for the origin of 'Long COVID'. Circulating nanobubbles, stable above physiological 0.17 M salt drive various enzyme-like activities and chemical reactions. Awareness of the microstructure of Pulmonary Surfactant and that nanobubbles of (O2/N2) and CO2 are integral to respiratory and circulatory physiology provides new insights to the COVID-19 and other pathogen activity.

Oxygen flux from capillary to mitochondria: integration of contemporary discoveries.

Resting humans transport ~ 100 quintillion (1018) oxygen (O2) molecules every second to tissues for consumption. The final, short distance (< 50µm) from capillary to the most distant mitochondria, in skeletal muscle where exercising O2 demands may increase 100-fold, challenges our understanding of O2 transport. To power cellular energetics O2 reaches its muscle mitochondrial target by dissociating from hemoglobin, crossing the red cell membrane, plasma, endothelial surface layer, endothelial cell, interstitial space, myocyte sarcolemma and a variable expanse of cytoplasm before traversing the mitochondrial outer/inner membranes and reacting with reduced cytochrome c and protons. This past century our understanding of O2's passage across the body's final O2 frontier has been completely revised. This review considers the latest structural and functional data, challenging the following entrenched notions: (1) That O2 moves freely across blood cell membranes. (2) The Krogh-Erlang model whereby O2 pressure decreases systematically from capillary to mitochondria. (3) Whether intramyocyte diffusion distances matter. (4) That mitochondria are separate organelles rather than coordinated and highly plastic syncytia. (5) The roles of free versus myoglobin-facilitated O2 diffusion. (6) That myocytes develop anoxic loci. These questions, and the intriguing notions that (1) cellular membranes, including interconnected mitochondrial membranes, act as low resistance conduits for O2, lipids and H+-electrochemical transport and (2) that myoglobin oxy/deoxygenation state controls mitochondrial oxidative function via nitric oxide, challenge established tenets of muscle metabolic control. These elements redefine muscle O2 transport models essential for the development of effective therapeutic countermeasures to pathological decrements in O2 supply and physical performance.

The Glomerular Endothelium Restricts Albumin Filtration.

Inflammatory activation and/or dysfunction of the glomerular endothelium triggers proteinuria in many systemic and localized vascular disorders. Among them are the thrombotic microangiopathies, many forms of glomerulonephritis, and acute inflammatory episodes like sepsis and COVID-19 illness. Another example is the chronic endothelial dysfunction that develops in cardiovascular disease and in metabolic disorders like diabetes. While the glomerular endothelium is a porous sieve that filters prodigious amounts of water and small solutes, it also bars the bulk of albumin and large plasma proteins from passing into the glomerular filtrate. This endothelial barrier function is ascribed predominantly to the endothelial glycocalyx with its endothelial surface layer, that together form a relatively thick, mucinous coat composed of glycosaminoglycans, proteoglycans, glycolipids, sialomucins and other glycoproteins, as well as secreted and circulating proteins. The glycocalyx/endothelial surface layer not only covers the glomerular endothelium; it extends into the endothelial fenestrae. Some glycocalyx components span or are attached to the apical endothelial cell plasma membrane and form the formal glycocalyx. Other components, including small proteoglycans and circulating proteins like albumin and orosomucoid, form the endothelial surface layer and are bound to the glycocalyx due to weak intermolecular interactions. Indeed, bound plasma albumin is a major constituent of the endothelial surface layer and contributes to its barrier function. A role for glomerular endothelial cells in the barrier of the glomerular capillary wall to protein filtration has been demonstrated by many elegant studies. However, it can only be fully understood in the context of other components, including the glomerular basement membrane, the podocytes and reabsorption of proteins by tubule epithelial cells. Discovery of the precise mechanisms that lead to glycocalyx/endothelial surface layer disruption within glomerular capillaries will hopefully lead to pharmacological interventions that specifically target this important structure.

New Endothelial Mechanisms in Glomerular (Patho)biology and Proteinuria Development Captured by Intravital Multiphoton Imaging.

In the past two decades, intravital imaging using multiphoton microscopy has provided numerous new visual and mechanistic insights into glomerular biology and disease processes including the function of glomerular endothelial cells (GEnC), podocytes, and the development of proteinuria. Although glomerular endothelial injury is known to precede podocyte damage in several renal diseases, the primary role of GEnCs in proteinuria development received much less attention compared to the vast field of podocyte pathobiology. Consequently, our knowledge of GEnC mechanisms in glomerular diseases is still emerging. This review highlights new visual clues on molecular and cellular mechanisms of GEnCs and their crosstalk with podocytes and immune cells that were acquired recently by the application of multiphoton imaging of the intact glomerular microenvironment in various proteinuric disease models. New mechanisms of glomerular tissue remodeling and regeneration are discussed based on results of tracking the fate and function of individual GEnCs using serial intravital multiphoton imaging over several days and weeks. The three main topics of this review include (i) the role of endothelial injury and microthrombi in podocyte detachment and albumin leakage via hemodynamic and mechanical forces, (ii) the alterations of the endothelial surface layer (glycocalyx) and its interactions with circulating immune cells in lupus nephritis, and (iii) the structural and functional remodeling and regeneration of GEnCs in hypertension, diabetes, and other experimental injury conditions. By the comprehensive visual portrayal of GEnCs and the many other contributing glomerular cell types, this review emphasizes the complexity of pathogenic mechanisms that result in proteinuria development.

The Endothelial Glycocalyx: Physiology and Pathology in Neonates, Infants and Children.

The endothelial glycocalyx (EG) as part of the endothelial surface layer (ESL) is an important regulator of vascular function and homeostasis, including permeability, vascular tone, leukocyte recruitment and coagulation. Located at the interface between the endothelium and the blood stream, this highly fragile structure is prone to many disruptive factors such as inflammation and oxidative stress. Shedding of the EG has been described in various acute and chronic diseases characterized by endothelial dysfunction and angiopathy, such as sepsis, trauma, diabetes and cardiovascular disease. Circulating EG components including syndecan-1, hyaluronan and heparan sulfate are being evaluated in animal and clinical studies as diagnostic and prognostic markers in several pathologies, and advances in microscopic techniques have enabled in vivo assessment of the EG. While research regarding the EG in adult physiology and pathology has greatly advanced throughout the last decades, our knowledge of the development of the glycocalyx and its involvement in pathological conditions in the pediatric population is limited. Current evidence suggests that the EG is present early during fetal development and plays a critical role in vessel formation and maturation. Like in adults, EG shedding has been demonstrated in acute inflammatory conditions in infants and children and chronic diseases with childhood-onset. However, the underlying mechanisms and their contribution to disease manifestation and progression still need to be established. In the future, the glycocalyx might serve as a marker to identify pediatric patients at risk for vascular sequelae and as a potential target for early interventions.

Links between Endothelial Glycocalyx Changes and Microcirculatory Parameters in Septic Patients.

The glycocalyx is an endothelial surface layer that is essential for maintaining microvascular homeostasis. Impaired integrity of the endothelial glycocalyx may be directly related to the development of microvascular dysfunction. To explore this hypothesis, we conducted a prospective observational study on adult patients diagnosed with sepsis. The study aimed to evaluate the degree of damage to the glycocalyx and to identify correlations between microcirculatory parameters and glycocalyx thickness based on capillary diameter. Sublingual microcirculation was examined using a handheld Cytocam-incident dark field video microscope. A sidestream dark field video microscope attached to a GlycoCheck monitor was used to determine the perfused boundary regions (PBRs) of sublingual blood vessels grouped by diameter (5–9 μm, 10–19 μm, and 20–25 μm). We identified significant damage to the glycocalyx in sublingual blood vessels of all the aforementioned diameters in septic patients compared to healthy age-matched controls. Furthermore, we found that the PBRs of the smallest capillaries (diameter class 5–9µm) correlated moderately and inversely with both total and perfused blood vessel densities. Collectively, our data suggest that there may be a functional relationship between damage to the endothelial glycocalyx of the smallest capillaries and alterations in the microcirculation observed in response to sepsis.

Effects of controlled ovarian stimulation on vascular barrier and endothelial glycocalyx: a pilot study

PurposeControlled ovarian stimulation significantly amplifies the number of maturing and ovulated follicles as well as ovarian steroid production. The ovarian hyperstimulation syndrome (OHSS) increases capillary permeability and fluid extravasation. Vascular integrity intensely is regulated by an endothelial glycocalyx (EGX) and we have shown that ovulatory cycles are associated with shedding of EGX components. This study investigates if controlled ovarian stimulation impacts on the integrity of the endothelial glycocalyx as this might explain key pathomechanisms of the OHSS.MethodsSerum levels of endothelial glycocalyx components of infertility patients (n=18) undergoing controlled ovarian stimulation were compared to a control group of healthy women with regular ovulatory cycles (n=17).ResultsPatients during luteal phases of controlled ovarian stimulation cycles as compared to normal ovulatory cycles showed significantly increased Syndecan-1 serum concentrations (12.6 ng/ml 6.1125th–19.1375th to 13.9 ng/ml 9.625th–28.975th; p=0.026), indicating shedding and degradation of the EGX.ConclusionA shedding of EGX components during ovarian stimulation has not yet been described. Our study suggests that ovarian stimulation may affect the integrity of the endothelial surface layer and increasing vascular permeability. This could explain key features of the OHSS and provide new ways of prevention of this serious condition of assisted reproduction.