Therapeutic Anticoagulation with Argatroban and Heparins Reduces Granulocyte Migration: Possible Impact on ECLS-Therapy?

Abstract

Introduction. Anticoagulants such as argatroban and heparins (low-molecular-weight and unfractionated) play an immense role in preventing thromboembolic complications in clinical practice. Nevertheless, they can also have a negative effect on the immune system. This study is aimed at investigating the influence of these substances on polymorphonuclear neutrophils (PMNs), whose nonspecific defense mechanisms can promote thrombogenesis. Methods. Blood samples from 30 healthy volunteers were investigated, whereby PMNs were isolated by density gradient centrifugation and incubated with 0.8 μg/mL of argatroban, 1.0 U/mL of low-molecular-weight heparin (LMWH), 1.0 U/mL of unfractionated heparin (UFH), or without drug (control). A collagen-cell mixture was prepared and filled into 3D μ-slide chemotaxis chambers (IBIDI® GmbH, Germany). Stimulation was initiated by using a chemokine gradient of n-formyl-methionine-leucyl-phenylalanine (fMLP), and microscopic observation was conducted for 4.5 hours. The cells’ track length and track straightness, as well as the number of attracted granulocytes, level of ROS (reactive oxygen species) production, and NET (neutrophil extracellular traps) formation, were analyzed and categorized into migration distances and time periods. Results. All three anticoagulants led to significantly reduced PMN track lengths, with UFH having the biggest impact. The number of tracks observed in the UFH group were significantly reduced compared to the control group. Additionally, the UFH group demonstrated a significantly lower track straightness compared to the control. ROS production and NET formation were unaffected. Conclusion. Our data provide evidence that anticoagulants have an inhibitory effect on the extent of PMN migration and chemotactic migration efficiency, thus indicating their potential immune-modulatory and prothrombotic effects.

Polymorphonuclear Cell Chemotaxis and Suicidal NETosis: Simultaneous Observation Using fMLP, PMA, H7, and Live Cell Imaging

“…There are also other substances capable of modulating PMN migration. Applying the same method described in this paper, PMN migration could either be decreased or increased using substances such as therapeutic anticoagulants [ 24 ] or IL-8 [ 12 ], respectively. This highlights the advantage of this method's approach: several conditions can be observed simultaneously for comparison.…”

“…NET formation is described as E T50 NETs in minutes, which is the time point of half-maximum effect of NET formation reached during the experiment. The sufficiency of analyzing time-resolved DAPI staining for NETosis identification has already been proven [ 22 , 24 ]. The “Spots” tool tracked each cell movement for the entire period of observation.…”

Potential Impact of Local Anesthetics Inducing Granulocyte Arrest and Altering Immune Functions on Perioperative Outcome

“…This experimental setup was based on previous experiments. [18][19][20] In brief, the cells were examined in temporal resolution by light and fluorescence microscopy over 8 hours. 3D-µ-slides (ibidi© GmbH, Martinsried, Germany) were used to perform a chemotactic assay.…”

Current Understanding of Leukocyte Phenotypic and Functional Modulation During Extracorporeal Membrane Oxygenation: A Narrative Review

“…Additional investigation into neonates and pediatrics, an even younger and thrombosis-prone population, may provide different insights. Furthermore, Bredthauer and colleagues showed that the frequently required anticoagulants used for ECMO inhibits neutrophil migration and chemotactic migration efficiency ( 100 ). The inhibitory effect of heparin and argatroban have been proposed to not only modulate neutrophil function but may also increase adhesion onto the membrane oxygenator to prompt clot formation.…”

“…Other than the discussed non-physiological stressors of changing flow dynamics and the associated shear stress generated within the circuit, patients are also frequently exposed to therapies such as multiple blood transfusions when treating bleeding complications ( 107 ), heparinization to reduce clot formations ( 11 , 12 ), supranormal administration of supplemental oxygen concerning hypoxia ( 108 ), and renal replacement therapy for ECMO-related AKI ( 9 ). These therapies have been independently reported to modulate leukocyte phenotype and function ( 44 – 46 , 100 , 109 – 111 ), which can result in a worsened prognosis. The cumulative impact of these therapies and stressors is likely to be significant.…”