Prolonged Cardiopulmonary Bypass is Associated With Endothelial Glycocalyx Degradation

Abstract

Background: The endothelial glycocalyx (EG) is involved in critical regulatory mechanisms that maintain endothelial vascular integrity. We hypothesized that prolonged cardiopulmonary bypass (CPB) may be associated with EG degradation. We performed an analysis of soluble syndecan-1 levels in relation to duration of CPB, as well as factors associated with cell stress and damage, such as mitochondrial DNA (mtDNA) and inflammation.

Methods: Blood samples from subjects undergoing cardiac surgery with CPB (n = 54) were obtained before and during surgery, 4-8 h and 24 h after completion of CPB, and on postoperative day 4. Flow cytometry was used to determine subpopulations of white blood cells. Plasma levels of mtDNA were determined using quantitative polymerase chain reaction and plasma content of shed syndecan-1 was measured. To determine whether syndecan-1 was signaling white blood cells, the effect of recombinant syndecan-1 on mobilization of neutrophils from bone marrow was tested in mice.

Results: CPB is associated with increased mtDNA during surgery, increased syndecan-1 blood levels at 4-8 h, and increased white blood cell count at 4-8 h and 24 h. Correlation analysis revealed significant positive associations between time on CPB and syndecan-1 (rs = 0.488, P < 0.001) and level of syndecan-1 and neutrophil count (rs = 0.351, P = 0.038) at 4-8 h. Intravenous administration of recombinant syndecan-1 in mice resulted in a 2.5-fold increase in the number of circulating neutrophils, concurrent with decreased bone marrow neutrophil number.

Conclusions: Longer duration of CPB is associated with increased plasma levels of soluble syndecan-1, a signal for EG degradation, which can induce neutrophil egress from the bone marrow. Development of therapy targeting EG shedding may be beneficial in patients with prolonged CPB.

Keywords: Cardiac surgery; Cardiopulmonary bypass; Damage associated molecular patterns; Endothelial glycocalyx; Neutrophils.

Copyright © 2020 Elsevier Inc. All rights reserved.

Figures

Figure 1.. Time-dependent changes in levels of mtDNA, soluble syndecan-1, WBC and IL-6.

A-D. Levels of mtDNA (A, n=50), syndecan-1 (B, n=44), WBC (C, n=44), and IL-6 (D, n=44) were measured in blood plasma obtained from cardiac surgery patients before (pre-op), during surgery and at 4–8 hours, 24 hours and 96 hours (day 4) after surgery. Friedman test with Dunn’s multiple comparisons test. P values are indicated.

Figure 2.. Associations between the duration of CPB and levels of mtDNA, soluble syndecan-1, WBC, and IL-6.

The correlation between CPB time and mtDNA (A,n=50), syndecan-1 (C, n=44), WBC (E-F, n=44), and IL-6 (G-H, n=44). The correlation analysis was performed at 4–8 hours (A, C, E and G) and 24 hours (F and H) after cardiac surgery. Spearman’s rank correlation coefficient and p values are indicated. The comparisons of mtDNA (B) and syndecan-1 (D) levels between groups of patients with shorter (grey-shaded circles) and longer (gray-shaded quadrants) CPB time. The statistical significance was calculated using repeated measures two-way ANOVA with Sidak’s multiple comparisons test. P values are indicated.

Figure 3.. Level of circulating syndecan-1 correlates with the number of neutrophils.

A. The correlation between level of soluble syndecan-1 and WBC at 4–8 hours after the surgery (n=44). B. Flow cytometry showing subpopulations of circulating WBC, stained with isotype-matched control (left) or antibody against CD45 (right). C. Number of neutrophils (left), monocytes (middle) and lymphocytes (right). Friedman test with Dunn’s multiple comparisons test. D. The relationships between levels of syndecan-1 and neutrophils (left), monocytes (middle), and lymphocytes (right). Spearman’s rank correlation coefficient and p values are indicated.

Figure 4.. Syndecan-1 induces mobilization of murine bone marrow myeloid cells.

A. Flow cytometry plots showing percentage of CD11bposLy6Gpos circulating neutrophils (upper right quadrant), CD11bposLy6Gneg monocytes (upper left quadrant) and lymphocytes (lower left quadrant) 6 hours later after injection of PBS (control, left) or 120 μg/kg human recombinant syndecan-1 (right). B. Graphical representation of data from flow cytometry showing number of (left to right): WBC, neutrophils, monocytes, and lymphocytes after administration of PBS (n=6) or syndecan-1 (SDC1, n=7). Unpaired t test. C. Flow cytometry plots showing percentage of CD11bposLy6Gpos neutrophils (black gate) in bone marrow 6 hours later after injection of PBS (control, left) or 120 μg/kg human recombinant syndecan-1 (right). D. Number of CD45pos cells (left) and neutrophils (right) in murine bone marrow after administration of PBS (n=6) or syndecan-1 (SDC1, n=7). Unpaired t test.