The Endothelial Glycocalyx: A Possible Therapeutic Target in Cardiovascular Disorders.

The physiological, anti-inflammatory, and anti-coagulant properties of endothelial cells (ECs) rely on a complex carbohydrate-rich layer covering the luminal surface of ECs, called the glycocalyx. In a range of cardiovascular disorders, glycocalyx shedding causes endothelial dysfunction and inflammation, underscoring the importance of glycocalyx preservation to avoid disease initiation and progression. In this review we discuss the physiological functions of the glycocalyx with particular focus on how loss of endothelial glycocalyx integrity is linked to cardiovascular risk factors, like hypertension, aging, diabetes and obesity, and contributes to the development of thrombo-inflammatory conditions. Finally, we consider the role of glycocalyx components in regulating inflammatory responses and discuss possible therapeutic interventions aiming at preserving or restoring the endothelial glycocalyx and therefore protecting against cardiovascular disease

Inflammatory stimuli induce shedding of heparan sulfate from arterial but not venous porcine endothelial cells leading to differential proinflammatory and procoagulant responses.

Endothelial dysfunction is an early event of vascular injury defined by a proinflammatory and procoagulant endothelial cell (EC) phenotype. Although endothelial glycocalyx disruption is associated with vascular damage, how various inflammatory stimuli affect the glycocalyx and whether arterial and venous cells respond differently is unknown. Using a 3D round-channel microfluidic system we investigated the endothelial glycocalyx, particularly heparan sulfate (HS), on porcine arterial and venous ECs. Heparan sulfate (HS)/glycocalyx expression was observed already under static conditions on venous ECs while it was flow-dependent on arterial cells. Furthermore, analysis of HS/glycocalyx response after stimulation with inflammatory cues revealed that venous, but not arterial ECs, are resistant to HS shedding. This finding was observed also on isolated porcine vessels. Persistence of HS on venous ECs prevented complement deposition and clot formation after stimulation with tumor necrosis factor α or lipopolysaccharide, whereas after xenogeneic activation no glycocalyx-mediated protection was observed. Contrarily, HS shedding on arterial cells, even without an inflammatory insult, was sufficient to induce a proinflammatory and procoagulant phenotype. Our data indicate that the dimorphic response of arterial and venous ECs is partially due to distinct HS/glycocalyx dynamics suggesting that arterial and venous thrombo-inflammatory disorders require targeted therapies

Sequential limiting in continuous and discontinuous Galerkin methods for the Euler equations

We present a new approach to enforcing local maximum principles in finite element schemes for the compressible Euler equations. In contrast to synchronized limiting techniques for systems of conservation laws, the density, momentum, and total energy are constrained in a sequential manner which guarantees positivity preservation for the pressure and internal energy. After the density limiting step, the total energy and momentum are adjusted to incorporate the irreversible effect of density changes. Then the corresponding antidiffusive corrections are limited to satisfy inequality constraints for the total and kinetic energy. The same element-based limiting strategy is employed in the context of continuous and discontinuous Galerkin methods. The sequential nature of the new limiting procedure makes it possible to achieve crisp resolution of contact discontinuities while using sharp local bounds in the energy constraints. A numerical study is performed for piecewise-linear finite element discretizations of 1D and 2D test problems

Autotransfusion system or integrated automatic suction device in minimized extracorporeal circulation: influence on coagulation and inflammatory response

Objective: To measure surrogate markers of coagulation activation as well as of the systemic inflammatory response in patients undergoing primary elective coronary artery bypass grafting (CABG) using either the so-called Smart suction device or a continuous autotransfusion system (C.A.T.S.®). Methods: Fifty-eight patients being operated with a miniaturized circuit (minimal extracorporeal circuit, MECC) were prospectively randomized to using a so-called Smart suction device or a routine continuous autotransfusion system (C.A.T.S.®) for collection of mediastinal shed blood. The coagulation response was measured by thrombin-antithrombin complex (TAT) and D-dimer. The inflammatory response was measured by Interleukin 6 (IL-6) and complement factor 3a (C3a) at three different time points, before surgery, 2h after surgery, as well as 18h after surgery. Results: No serious adverse cardiovascular event was observed. Serum levels of TAT significantly differed between both groups 2h after surgery (Smart suction 16.12±13.51μgl−1 vs C.A.T.S® 9.83±7.81μgl−1, p=0.040) and returned to baseline values after 18h in both groups. Serum levels of D-dimer showed a corresponding pattern with a peak 2h after surgery (Smart suction 1115±1231ngml−1 vs C.A.T.S.® 507±604ngml−1, p=0.025). IL-6 levels also significantly differed between both groups 2h after surgery (Smart suction 186±306pgml−1 vs C.A.T.S.® 82±71pgml−1, p=0.072). No significant changes in serum levels of C3a over time could be observed. Conclusions: Despite no differences in the clinical course of patients with either Smart suction or C.A.T.S.® being observed, surrogate markers of coagulation and inflammation seem to be less pronounced in patients where cardiotomy blood is not being directly reinfused. As such, C.A.T.S.® should be preferred in routine CABG, as long as no extensive volume substitution is anticipate

Addition of dextran sulfate to blood cardioplegia attenuates reperfusion injury in a porcine model of cardiopulmonary bypass

Objective: Contact of blood with artificial surfaces and air as well as ischemia/reperfusion injury to the heart and lungs mediate systemic and local inflammation during cardiopulmonary bypass (CPB). Activation of complement and coagulation cascades leads to and accompanies endothelial cell damage. Therefore, endothelial-targeted cytoprotection with the complement inhibitor and endothelial protectant dextran sulfate (DXS, MW 5000) may attenuate CBP-associated myocardial and pulmonary injury. Methods: Eighteen pigs (DXS, n=10; phosphate buffered saline [PBS], n=8) underwent standard cardiopulmonary bypass. After aortic cross-clamping, cardiac arrest was initiated with modified Buckberg blood cardioplegia (BCP), repeated after 30 and 60min with BCP containing either DXS (300mg/10ml, equivalent to 5mg/kg) or 10ml of PBS. Following 30min reperfusion, pigs were weaned from CPB. During 2h of observation, cardiac function was monitored by echocardiography and invasive pressure measurements. Inflammatory and coagulation markers were assessed regularly. Animals were then sacrificed and heart and lungs analyzed. Results: DXS significantly reduced CK-MB levels (43.4±14.8ng/ml PBS, 35.9±11.1ng/ml DXS, p=0.042) and significantly diminished cytokine release: TNFalpha (1507.6±269.2pg/ml PBS, 222.1±125.6pg/ml DXS, p=0.0071), IL1beta (1081.8±203.0pg/ml PBS, 110.7±79.4pg/ml DXS, p=0.0071), IL-6 (173.0±91.5pg/ml PBS, 40.8±19.4pg/ml DXS, p=0.002) and IL-8 (304.6±81.3pg/ml PBS, 25.4±14.2pg/ml DXS, p=0.0071). Tissue endothelin-1 levels were significantly reduced (6.29±1.90pg/100mg PBS, 3.55±1.15pg/100mg DXS p=0.030) as well as thrombin-anti-thrombin formation (20.7±1.0μg/ml PBS, 12.8±4.1μg/ml DXS, p=0.043). Also DXS reduced cardiac and pulmonary complement deposition, neutrophil infiltration, hemorrhage and pulmonary edema (measured as lung water content, 81±3% vs 78±3%, p=0.047), indicative of attenuated myocardial and pulmonary CPB-injury. Diastolic left ventricular function (measured as dp/dtmin), pulmonary artery pressure (21±3mmHg PBS, 19±3mmHg DXS, p=0.002) and right ventricular pressure (21±1mmHg PBS, 19±3mmHg DXS p=0.021) were significantly improved with the use of DXS. Conclusions: Addition of DXS to the BCP solution ameliorates post-CPB injury and to a certain extent improves cardiopulmonary function. Endothelial protection in addition to myocyte protection may improve post-CPB outcome and recover

Edema is a sign of early acute myocardial infarction on post-mortem magnetic resonance imaging

The aim of this study was to investigate if acute myocardial infarction can be detected by post-mortem cardiac magnetic resonance (PMMR) at an earlier stage than by traditional autopsy, i.e., within less than 4h after onset of ischemia; and if so, to determine the characteristics of PMMR findings in early acute infarcts. Twenty-one ex vivo porcine hearts with acute myocardial infarction underwent T2-weighted cardiac PMMR imaging within 3 h of onset of iatrogenic ischemia. PMMR imaging findings were compared to macroscopic findings. Myocardial edema induced by ischemia and reperfusion was visible on PMMR in all cases. Typical findings of early acute ischemic injury on PMMR consist of a central zone of intermediate signal intensity bordered by a rim of increased signal intensity. Myocardial edema can be detected on cardiac PMMR within the first 3h after the onset of ischemia in porcine hearts. The size of myocardial edema reflects the area of ischemic injury in early acute (per-acute) myocardial infarction. This study provides evidence that cardiac PMMR is able to detect acute myocardial infarcts at an earlier stage than traditional autopsy and routine histolog

Surface modification of pig endothelial cells with a branched heparin conjugate improves their compatibility with human blood

Corline Heparin Conjugate (CHC), a compound of multiple unfractionated heparin chains, coats cells with a glycocalyx-like layer and may inhibit (xeno) transplant-associated activation of the plasma cascade systems. Here, we investigated the use of CHC to protect WT and genetically modified (GTKO. hCD46. hTBM) pig aortic endothelial cells (PAEC) in two pig-to-human in vitro xenotransplantation settings. Model 1: incubation of untreated or hTNFa-treated PAEC with 10% human plasma induced complement C3b/c and C5b-9 deposition, cellular activation and coagulation activation in WT and GTKO. hCD46. hTBM PAEC. Coating of untreated or hTNFa-treated PAEC with CHC (100 mu g/ml) protected against human plasma-induced endothelial activation and damage. Model 2: PAEC were grown on microcarrier beads, coated with CHC, and incubated with non-anticoagulated whole human blood. Genetically modified PAEC significantly prolonged clotting time of human blood (115.0 +/- 16.1 min, p < 0.001) compared to WT PAEC (34.0 +/- 8.2 min). Surface CHC significantly improved the human blood compatibility of PAEC, as shown by increased clotting time (WT: 84.3 +/- 11.3 min, p < 0.001;GTKO. hCD46. hTBM: 146.2 +/- 20.4 min, p < 0.05) and reduced platelet adhesion, complement activation, coagulation activation and inhibition of fibrinolysis. The combination of CHC coating and genetic modification provided the greatest compatibility with human blood, suggesting that pre-transplant perfusion of genetically modified porcine organs with CHC may benefit post-transplant xenograft function

Attenuation of myocardial reperfusion injury in pigs by Mirococept, a membrane-targeted complement inhibitor derived from human CR1

Objectives Membrane-targeted application of complement inhibitors may ameliorate ischemia/reperfusion (I/R) injury by directly targeting damaged cells. We investigated whether Mirococept, a membrane-targeted, myristoylated peptidyl construct derived from complement receptor 1 (CR1) could attenuate I/R injury following acute myocardial infarction in pigs. Methods In a closed-chest pig model of acute myocardial infarction, Mirococept, the non-tailed derivative APT154, or vehicle was administered intracoronarily into the area at risk 5 min pre-reperfusion. Infarct size, cardiac function and inflammatory status were evaluated. Results Mirococept targeted damaged vasculature and myocardium, significantly decreasing infarct size compared to vehicle, whereas APT154 had no effect. Cardioprotection correlated with reduced serum troponin I and was paralleled by attenuated local myocardial complement deposition and tissue factor expression. Myocardial apoptosis (TUNEL-positivity) was also reduced with the use of Mirococept. Local modulation of the pro-inflammatory and pro-coagulant phenotype translated to improved left ventricular end-diastolic pressure, ejection fraction and regional wall motion post-reperfusion. Conclusions Local modification of a pro-inflammatory and pro-coagulant environment after regional I/R injury by site-specific application of a membrane-targeted complement regulatory protein may offer novel possibilities and insights into potential treatment strategies of reperfusion-induced injur

Locally targeted cytoprotection with dextran sulfate attenuates experimental porcine myocardial ischaemia/reperfusion injury

Aims Intravascular inflammatory events during ischaemia/reperfusion injury following coronary angioplasty alter and denudate the endothelium of its natural anticoagulant heparan sulfate proteoglycan (HSPG) layer, contributing to myocardial tissue damage. We propose that locally targeted cytoprotection of ischaemic myocardium with the glycosaminoglycan analogue dextran sulfate (DXS, MW 5000) may protect damaged tissue from reperfusion injury by functional restoration of HSPG. Methods and results In a closed chest porcine model of acute myocardial ischaemia/reperfusion injury (60 min ischaemia, 120 min reperfusion), DXS was administered intracoronarily into the area at risk 5 min prior to reperfusion. Despite similar areas at risk in both groups (39±8% and 42±9% of left ventricular mass), DXS significantly decreased myocardial infarct size from 61±12% of the area at risk for vehicle controls to 39±14%. Cardioprotection correlated with reduced cardiac enzyme release creatine kinase (CK-MB, troponin-I). DXS abrogated myocardial complement deposition and substantially decreased vascular expression of pro-coagulant tissue factor in ischaemic myocardium. DXS binding, detected using fluorescein-labelled agent, localized to ischaemically damaged blood vessels/myocardium and correlated with reduced vascular staining of HSPG. Conclusion The significant cardioprotection obtained through targeted cytoprotection of ischaemic tissue prior to reperfusion in this model of acute myocardial infarction suggests a possible role for the local modulation of vascular inflammation by glycosaminoglycan analogues as a novel therapy to reduce reperfusion injur