Rationale
Pre-clinical and autopsy studies have fueled the hypothesis that a dysregulated vascular endothelium might play a central role in the pathogenesis of ARDS and multi-organ failure in ...COVID-19.
Objectives
To comprehensively characterize and quantify microvascular alterations in patients with COVID-19.
Methods
Hospitalized adult patients with moderate-to-severe or critical COVID-19 (
n
= 23) were enrolled non-consecutively in this prospective, observational, cross-sectional, multi-center study. Fifteen healthy volunteers served as controls. All participants underwent intravital microscopy by sidestream dark field imaging to quantify vascular density, red blood cell velocity (
V
RBC
), and glycocalyx dimensions (perfused boundary region, PBR) in sublingual microvessels. Circulating levels of endothelial and glycocalyx-associated markers were measured by multiplex proximity extension assay and enzyme-linked immunosorbent assay.
Measurements and main results
COVID-19 patients showed an up to 90% reduction in vascular density, almost exclusively limited to small capillaries (diameter 4–6 µm), and also significant reductions of
V
RBC
. Especially, patients on mechanical ventilation showed severe glycocalyx damage as indicated by higher PBR values (i.e., thinner glycocalyx) and increased blood levels of shed glycocalyx constituents. Several markers of endothelial dysfunction were increased and correlated with disease severity in COVID-19. PBR (AUC 0.75,
p
= 0.01), ADAMTS13 (von Willebrand factor-cleaving protease; AUC 0.74,
p
= 0.02), and vascular endothelial growth factor A (VEGF-A; AUC 0.73,
p
= 0.04) showed the best discriminatory ability to predict 60-day in-hospital mortality.
Conclusions
Our data clearly show severe alterations of the microcirculation and the endothelial glycocalyx in patients with COVID-19. Future therapeutic approaches should consider the importance of systemic vascular involvement in COVID-19.
The endothelial glycocalyx (eGC) covers the luminal surface of the vascular endothelium and plays an important protective role in systemic inflammatory states and particularly in sepsis. Its ...breakdown leads to capillary leak and organ dysfunction. Moreover, sepsis-induced alterations of sublingual microcirculation are associated with a worse clinical outcome. The present study was performed to investigate the associations between eGC dimensions and established parameters of microcirculation dysfunction in sepsis.
This observational, prospective, cross-sectional study included 40 participants, of which 30 critically ill septic patients were recruited from intensive care units of a university hospital and 10 healthy volunteers served as controls. The established microcirculation parameters were obtained sublingually and analyzed according to the current recommendations. In addition, the perfused boundary region (PBR), an inverse parameter of the eGC dimensions, was measured sublingually, using novel data acquisition and analysis software (GlycoCheck™). Moreover, we exposed living endothelial cells to 5% serum from a subgroup of study participants, and the delta eGC breakdown, measured with atomic force microscopy (AFM), was correlated with the paired PBR values.
In septic patients, sublingual microcirculation was impaired, as indicated by a reduced microvascular flow index (MFI) and a reduced proportion of perfused vessels (PPV) compared to those in healthy controls (MFI, 2.93 vs 2.74, p = 0.002; PPV, 98.53 vs 92.58, p = 0.0004). PBR values were significantly higher in septic patients compared to those in healthy controls, indicating damage of the eGC (2.04 vs 2.34, p < 0.0001). The in vitro AFM data correlated exceptionally well with paired PBR values obtained at the bedside (rs = - 0.94, p = 0.02). Both PBR values and microcirculation parameters correlated well with the markers of critical illness. Interestingly, no association was observed between the PBR values and established microcirculation parameters.
Our findings suggest that eGC damage can occur independently of microcirculatory impairment as measured by classical consensus parameters. Further studies in critically ill patients are needed to unravel the relationship of glycocalyx damage and microvascular impairment, as well as their prognostic and therapeutic importance in sepsis.
Retrospectively registered: Clinicaltrials.gov, NCT03960307.
The availability of handheld, noninvasive sublingual video-microscopes allows for visualization of the microcirculation in critically ill patients. Recent studies demonstrate that reduced numbers of ...blood-perfused microvessels and increased penetration of erythrocytes into the endothelial glycocalyx are essential components of microvascular dysfunction. The aim of this study was to identify novel microvascular variables to determine the level of microvascular dysfunction in sepsis and its relationship with clinical variables.
This observational, prospective, cross-sectional study included 51 participants, of which 34 critically ill sepsis patients were recruited from intensive care units of a university hospital. Seventeen healthy volunteers served as controls. All participants underwent sublingual videomicroscopy by sidestream darkfield imaging. A new developed version of the Glycocheck™ software was used to quantify vascular density, perfused boundary region (PBR-an inverse variable of endothelial glycocalyx dimensions), red blood cell (RBC) velocity, RBC content, and blood flow in sublingual microvessels with diameters between 4 and 25 µm.
A detailed analysis of adjacent diameter classes (1 µm each) of vessels between 4 and 25 µm revealed a severe reduction of vascular density in very small capillaries (5-7 µm), which correlated with markers of sepsis severity. Analysis of RBC velocity (V
) revealed a strong dependency between capillary and feed vessel V
in sepsis patients (R
= 0.63, p < 0.0001) but not in healthy controls (R
= 0.04, p = 0.43), indicating impaired capillary (de-)recruitment in sepsis. This finding enabled the calculation of capillary recruitment and dynamic capillary blood volume (CBV
). Moreover, adjustment of PBR to feed vessel V
further improved discrimination between sepsis patients and controls by about 50%. By combining these dynamic microvascular and glycocalyx variables, we developed the microvascular health score (MVHS
™), which decreased from 7.4 4.6-8.7 in controls to 1.8 1.4-2.7 in sepsis patients (p < 0.0001) and correlated with sepsis severity.
We introduce new important diameter-specific quantification and differentiated analysis of RBC kinetics, a key to understand microvascular dysfunction in sepsis. MVHS
, which has a broad bandwidth to detect microvascular (dys-) function, might serve as a valuable tool to detect microvascular impairment in critically ill patients.
Abstract
Background
Bacterial burden as well as duration of bacteremia influence the outcome of patients with bloodstream infections. Promptly decreasing bacterial load in the blood by using ...extracorporeal devices in addition to anti-infective therapy has recently been explored. Preclinical studies with the Seraph® 100 Microbind® Affinity Blood Filter (Seraph® 100), which consists of heparin that is covalently bound to polymer beads, have demonstrated an effective binding of bacteria and viruses. Pathogens adhere to the heparin coated polymer beads in the adsorber as they would normally do to heparan sulfate on cell surfaces. Using this biomimetic principle, the Seraph® 100 could help to decrease bacterial burden in vivo.
Methods
This first in human, prospective, multicenter, non-randomized interventional study included patients with blood culture positive bloodstream infection and the need for kidney replacement therapy as an adjunctive treatment for bloodstream infections. We performed a single four-hour hemoperfusion treatment with the Seraph® 100 in conjunction with a dialysis procedure. Post procedure follow up was 14 days.
Results
Fifteen hemodialysis patients (3F/12 M, age 74.0 68.0–78.5 years, dialysis vintage 28.0 11.0–45.0 months) were enrolled. Seraph® 100 treatment started 66.4 45.7–80.6 hours after the initial positive blood culture was drawn. During the treatment with the Seraph® 100 with a median blood flow of 285 225–300 ml/min no device or treatment related adverse events were reported. Blood pressure and heart rate remained stable while peripheral oxygen saturation improved during the treatment from 98.0 92.5–98.0 to 99.0 98.0–99.5 %;
p
= 0.0184. Four patients still had positive blood culture at the start of Seraph® 100 treatment. In one patient blood cultures turned negative during treatment. The time to positivity (TTP) was increased between inflow and outflow blood cultures by 36 − 7.2 to 96.3 minutes. However, overall TTP increase was not statistical significant.
Conclusions
Seraph® 100 treatment was well tolerated. Adding Seraph® 100 to antibiotics early in the course of bacteremia might result in a faster resolution of bloodstream infections, which has to be evaluated in further studies
.
Trail registration
: ClinicalTrials.gov Identifier:
NCT02914132
, first posted September 26, 2016.
Deterioration of the endothelial glycocalyx (eGC), a protective carbohydrate-rich layer lining the luminal surface of the endothelium, plays a key role in vascular barrier dysfunction and eventually ...organ-failure in systemic inflammatory response syndrome and sepsis. Early detection of glycocalyx damage could thus become an important goal in critical care. This study was designed to determine the feasibility and reproducibility of quantitative, real-time glycocalyx measurements performed at bedside in the emergency room (ER) and intensive care unit (ICU).
The observational study included 70 patients admitted to the ER or ICU of a university hospital. A physician and the nurse in charge of the patient performed sublingual microcirculatory measurements using sidestream dark field (SDF) imaging. A novel data acquisition and analysis software (GlycoCheck™) was used to analyze the perfused boundary region (PBR), an inverse parameter of endothelial glycocalyx dimensions in vessels with diameters of between 5 and 25 μm.
The method showed a good intra-observer reproducibility. Specifically, intraclass correlation coefficient analysis showed an excellent reproducibility between the physician's measurements (0.77 CI 95%: 0.52-0.89). The bias between the two PBRs was - 0.077 ± 0.24 μm. Moreover, there were no significant differences in the PBR values obtained by the nurses when compared to those reported by the physician (regarded as the "gold standard" measurement). Intraclass correlation coefficient analysis showed excellent reproducibility between the nurses' and physician's PBRs (0.75 95% CI: 0.52-0.87). The mean difference between the two PBRs (i.e., the bias) was 0.007 ± 0.25 μm. The nurses' PBR assessment had a 90% sensitivity (95% CI: 60-99%) and 90% specificity (95% CI: 80-93%) to identify a severely impaired glycocalyx.
Glycocalyx dimensions can be measured at patients' bedside precisely by non-invasive assessment of the PBR. This assessment could become part of standard monitoring and contribute to clinical decision-making and resuscitation protocols in clinical trials and daily practice.
The COVID-19 pandemic is caused by the SARS CoV-2 virus and can lead to severe lung damage and hyperinflammation. In the context of COVID-19 infection, inflammation-induced degradation of the ...glycocalyx layer in endothelial cells has been demonstrated. Syndecan-1 (SDC-1) is an established parameter for measuring glycocalyx injury. This prospective, multicenter, observational, cross-sectional study analyzed SDC-1 levels in 24 convalescent patients that had been infected with SARS-CoV-2 with mild disease course without need of hospitalization. We included 13 age-matched healthy individuals and 10 age-matched hospitalized COVID-19 patients with acute mild disease course as controls. In convalescent COVID-19 patients, significantly elevated SDC-1 levels were detected after a median of 88 days after symptom onset compared to healthy controls, whereas no difference was found when compared to SDC-1 levels of hospitalized patients undergoing acute disease. This study is the first to demonstrate signs of endothelial damage in non-pre-diseased, convalescent COVID-19 patients after mild disease progression without hospitalization. The data are consistent with studies showing evidence of persistent endothelial damage after severe or critical disease progression. Further work to investigate endothelial damage in convalescent COVID-19 patients should follow.
The endothelial glycocalyx (eGC), a delicate carbohydrate-rich structure lining the luminal surface of the vascular endothelium, is vital for maintenance of microvascular homeostasis. In sepsis, ...damage of the eGC triggers the development of vascular hyperpermeability with consecutive edema formation and organ failure. While there is evidence that protection or rebuilding of the eGC might counteract sepsis-induced vascular leakage and improve outcome, approved therapeutics are not yet available. This narrative review aims to outline possible therapeutic strategies to ameliorate organ dysfunction caused by eGC impairment.
Coronavirus disease 2019 (COVID-19) is a systemic disease associated with injury (thinning) of the endothelial glycocalyx (eGC), a protective layer on the vascular endothelium. The aim of this ...translational study was to investigate the role of the eGC-degrading enzyme heparanase (HPSE), which is known to play a central role in the destruction of the eGC in bacterial sepsis. Excess activity of HPSE in plasma from COVID-19 patients correlated with several markers of eGC damage and perfused boundary region (PBR, an inverse estimate of glycocalyx dimensions of vessels with a diameter 4-25 µm). In a series of translational experiments, we demonstrate that the changes in eGC thickness of cultured cells exposed to COVID-19 serum correlated closely with HPSE activity in concordant plasma samples (R = 0.82, P = 0.003). Inhibition of HPSE by a nonanticoagulant heparin fragment prevented eGC injury in response to COVID-19 serum, as shown by atomic force microscopy and immunofluorescence imaging. Our results suggest that the protective effect of heparin in COVID-19 may be due to an eGC-protective off-target effect.
Microcirculatory disorders are crucial in pathophysiology of organ dysfunction in critical illness. Evaluation of sublingual microcirculation is not routinely conducted in daily practice due to ...time-consuming analysis and susceptibility to artifacts. We investigated the suitability of optical coherence tomography angiography (OCTA) for contactless evaluation of sublingual microcirculation. Sublingual microcirculation was imaged in 10 healthy volunteers, using an OCTA device and an incident dark field (IDF) illumination microscopy (current gold standard). OCTA images were analyzed with regard to flow density and perfused vessel density (PVD
). IDF videos were analyzed following current recommendations. Flow density was automatically extracted from OCTA images (whole en face 48.9% 43.2; 54.5; central ring 52.6% 43.6; 60.6). PVD
did not differ from the PVD calculated from IDF videos (PVD
18.6 mm/mm² 18.0; 21.7) vs. PVD
21.0 mm/mm² 17.5; 22.9; p = 0.430). Analysis according to Bland-Altman revealed a mean bias of 0.95 mm/mm² (95% Confidence interval -1.34 to 3.25) between PVD
and PVD
with limits of agreement of -5.34 to 7.24 mm/mm². This study is the first to demonstrate the suitability of OCTA for evaluating sublingual microcirculation. Comparison of the perfused vessel density between methods showed a plausible level of agreement.
(1) Damage to the endothelial glycocalyx (eGC), a protective layer lining the endothelial luminal surface, is associated with chronic kidney disease (CKD), which leads to a worsening of ...cardiovascular outcomes in these patients. Currently, there are no targeted therapeutic approaches. Whether the dietary supplement Endocalyx
(ECX) protects against endothelial damage caused by uremic toxins is unknown. (2) We addressed this question by performing atomic force microscopy measurements on living endothelial cells. We examined the effect of ECX on eGC thickness at baseline and with pooled serum from hemodialysis patients. ECX was also successfully administered in vivo in mice, in which eGC was assessed using perfused boundary region measurements by intravital microscopy of cremasteric vessels. (3) Both ECX and fucoidan significantly improved baseline eGC thickness. Our data indicate that these effects are dependent on ERK/MAPK and PI3K signaling. After incubation with eGC damaging serum from dialysis patients, ECX increased eGC height. Intravital microscopy in mice revealed a relevant increase in baseline eGC dimensions after feeding with ECX. (4) We identified a dietary supplement containing glycocalyx substrates and fucoidan as potential mediators of eGC preservation in vitro and in vivo. Our findings suggest that fucoidan may be an essential component responsible for protecting the eGC in acute settings. Moreover, ECX might contribute to both protection and rebuilding of the eGC in the context of CKD.
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