Influence of hemoadsorption during cardiopulmonary bypass on blood vesicle count and function

Lukas Wisgrill, Christian Lamm, Lena Hell, Johannes Thaler, Angelika Berger, Rene Weiss, Viktoria Weber, Harald Rinoesl, Michael J Hiesmayr, Andreas Spittler, Martin H Bernardi, Lukas Wisgrill, Christian Lamm, Lena Hell, Johannes Thaler, Angelika Berger, Rene Weiss, Viktoria Weber, Harald Rinoesl, Michael J Hiesmayr, Andreas Spittler, Martin H Bernardi

Abstract

Background: Extracorporeal circulation during major cardiac surgery triggers a systemic inflammatory response affecting the clinical course and outcome. Recently, extracellular vesicle (EV) research has shed light onto a novel cellular communication network during inflammation. Hemoadsorption (HA) systems have shown divergent results in modulating the systemic inflammatory response during cardiopulmonary bypass (CPB) surgery. To date, the effect of HA on circulating microvesicles (MVs) in patients undergoing CPB surgery is unknown.

Methods: Count and function of MVs, as part of the extracellular vesicle fraction, were assessed in a subcohort of a single-center, blinded, controlled study investigating the effect of the CytoSorb device during CPB. A total of 18 patients undergoing elective CPB surgery with (n = 9) and without (n = 9) HA device were included in the study. MV phenotyping and counting was conducted via flow cytometry and procoagulatory potential was measured by tissue factor-dependent MV assays.

Results: Both study groups exhibited comparable counts and post-operative kinetics in MV subsets. Tissue factor-dependent procoagulatory potential was not detectable in plasma at any timepoint. Post-operative course and laboratory parameters showed no correlation with MV counts in patients undergoing CPB surgery.

Conclusion: Additional artificial surfaces to the CPB-circuit introduced by the use of the HA device showed no effect on circulating MV count and function in these patients. Larger studies are needed to assess and clarify the effect of HA on circulating vesicle counts and function. Trial registration ClinicalTrials.Gov Identifier: NCT01879176; registration date: June 17, 2013; https://ichgcp.net/clinical-trials-registry/NCT01879176.

Keywords: Blood vesicle; Cardiopulmonary bypass; Extracellular vesicles; Hemoadsorption; Microvesicles.

Conflict of interest statement

MHB and MJH have received travel funding for a lecture from CytoSorbents Europe, GmbH. All other authors declare that they have no competing interests.

Figures

Fig. 1
Fig. 1
Flow cytometric setup and gating strategy. Green fluorescent silica beads (1000 nm) were used to define the side scatter properties and the microvesicle (MV) gate was set right underneath the silica bead population (left panel). Using an unlabeled plasma sample, the scatter and gate settings were validated as seen in the right panel without any trigger. All events above the MV-gate were assumed to be larger than 1000 nm and were separately analyzed as apoptotic bodies (ABs). a Next, the fluorescence trigger was set for the APC-channel (Annexin V (AnnV)) using an AnnV-stained plasma sample without calcium, preventing calcium-dependent labelling of AnnV (left Panel). The fluorescence trigger was set and validated using an AnnV-labelled plasma sample (right panel; b). The gating strategy of CD41 + MVs (platelet-derived MVs; left panel) and CD235 + (erythrocyte-derived MVs; right panel) (c)
Fig. 2
Fig. 2
Individual count of circulating blood vesicles. Individual microvesicle (MV) and apoptotic body (AB) counts (events/µL) in plasma samples from each patients undergoing major cardiac surgery with cardiopulmonary bypass (CPB) in Control group (n = 9; a) and Cytosorb group (n = 9; b) at different timepoints
Fig. 3
Fig. 3
Count and hemolysis parameter correlation of circulating blood vesicles. Vesicle counts (events/µL) in plasma samples from patients undergoing major cardiac surgery with cardiopulmonary bypass (CPB) with (n = 9; CytoSorb; black triangles) or without (n = 9; control; gray circles) hemoadsorption. Samples were analyzed using flow cytometry. Data are presented as mean ± standard deviation (a). Scanning electron microscopy was performed to analyze a hemoadsorption column directly after use. Respective pictures are shown in ×400 and ×5000 magnification (b). Correlation analysis was performed to assess the link between vesicle counts, hemolysis and infection parameters. The color represents the respective rho-value and the presence of circles indicate a significant p-value < 0.05 (c)
Fig. 4
Fig. 4
Correlation analysis of blood cytokine/alarmin levels and microvesicle count. Correlation analysis of total microvesicle (total MV) counts and HMGB1 plasma levels (a). Correlation plot of investigated cytokines, alarmins and inotropic drugs with MV counts. The color represents the respective rho-value and the presence of circles indicate a significant p-value < 0.05 (b)

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