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1The impact of interferon-alpha on RBC-endothelium interaction
assessed with optical tweezers
M. Maksimov1*, P. Ermolinskiy1, D. Umerenkov1, O. Scheglovitova2, A. Lugovtsov1, A. Priezzhev1
1-Faculty of Physics, Lomonosov Moscow State University, 1-2 Leninskiye Gory, Moscow, Russia, 119991 2- The Gamaleya National Center, 18 Gamaleya St., Moscow, Russia, 123098
Physiological functions of red blood cells (RBCs) include oxygen and carbon dioxide transport, immune response, blood coagulation and regulation of blood rheology. The inner surface of human vessels is covered with endothelial cells, which serve as a semipermeable barrier between the blood and the tissues. Endothelial cells release signaling molecules, which can change the properties of other cells, RBCs included. Thus, RBCs can interact with endothelium not only through adhesion to these cells, but also via biochemical signals. One of such signaling biomolecules is nitric oxide (NO), which is known to increase RBC deformability and decrease RBC aggregation [1].
Interferon alpha-2b (IFN-a) is an approved antiviral drug against hepatitis C and B. Lately it was used to treat patients with COVID-19 [2]. It is known that IFN-a impacts endothelial cells, decreasing their permeability and reducing NO production [3,4]. While endothelium is badly damaged in some cases of COVID-19, it is of interest, whether IFN-a alters RBC-endothelium interaction, either direct (adhesion) or indirect (NO production).
The main aim of this study was to investigate in vitro RBC-endothelium interaction and endothelial impact on RBC aggregation at different concentrations of IFN-a (0, 10, 30, 100, 300, 1000 units/ml). To achieve these, optical tweezers trapping technique was applied to measure forces of interaction between single RBCs as well as between RBC and the monolayer of endothelial cells.
The human umbilical vein endothelial cells (HUVEC) were incubated with corresponding concentrations of IFN-a for approximately 24 hours. Lithium heparin anticoagulant was used for blood drawing. The studied samples represented microcuvettes with endothelial monolayer at the bottom, filled with blood plasma containing small number of RBCs (with the hematocrit of approximately 0.05%). Laser tweezers provide the ability to trap and manipulate single cells, allowing to measure their interaction forces in the pN range. In this work, homemade dual-beam laser tweezers setup with two traps based on Nd:YAG laser (1064 nm) was used [5].
Our results demonstrate statistically significant decrease in RBC aggregation in the presence of endothelium and IFN-a. The degree of the effect is higher if IFN-a was added to the plasma without preliminary incubation. On the contrary, samples with endothelium incubated with IFN-a for 24 hours, but lacking IFN-a in the plasma during the measurements, showed weaker tendencies in decreasing RBC aggregation. The force of RBC-endothelium adhesion was in the range of 1 -2 pN in almost all of the experiments and demonstrated weak tendencies to decreasing with the increase in IFN-a concentration. These results may witness in favor of short-time IFN-a effect on the RBC-endothelium system. Further investigation of RBC-endothelium-IFN-a interplay may spread light on the underlying mechanisms and provide clues for new therapeutic targets.
This work was supported by the Russian Science Foundation (Grant No. 22-15-00120).
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[2] R. Pereda, et al, Therapeutic effectiveness of interferon alpha 2b treatment for COVID-19 patient recovery, Journal of Interferon & Cytokine Research, Vol. 40, № 12, pp. 578-588, (2020).
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[5] A. Lugovtsov, et al, Optical assessment of alterations of microrheologic and microcirculation parameters in cardiovascular diseases, Biomed Opt Express, Vol. 10, № 8, pp. 3974-3986, (2019).
