Recombinant Human Soluble Thrombomodulin Suppresses Monocyte Adhesion by Reducing Lipopolysaccharide-Induced Endothelial Cellular Stiffening

Abstract

Endothelial cellular stiffening has been observed not only in inflamed cultured endothelial cells but also in the endothelium of atherosclerotic regions, which is an underlying cause of monocyte adhesion and accumulation. Although recombinant soluble thrombomodulin (rsTM) has been reported to suppress the inflammatory response of endothelial cells, its role in regulating endothelial cellular stiffness remains unclear. The purpose of this study was to investigate the impact of anticoagulant rsTM on lipopolysaccharide (LPS)-induced endothelial cellular stiffening. We show that LPS increases endothelial cellular stiffness by using atomic force microscopy and that rsTM reduces LPS-induced cellular stiffening not only through the attenuation of actin fiber and focal adhesion formation but also via the improvement of gap junction functionality. Moreover, post-administration of rsTM, after LPS stimulation, attenuated LPS-induced cellular stiffening. We also found that endothelial cells regulate leukocyte adhesion in a substrate- and cellular stiffness-dependent manner. Our result show that LPS-induced cellular stiffening enhances monocytic THP-1 cell line adhesion, whereas rsTM suppresses THP-1 cell adhesion to inflamed endothelial cells by reducing cellular stiffness. Endothelial cells increase cellular stiffness in reaction to inflammation, thereby promoting monocyte adhesion. Treatment of rsTM reduced LPS-induced cellular stiffening and suppressed monocyte adhesion in a cellular stiffness-dependent manner.

Recombinant Human Soluble Thrombomodulin Suppresses Arteritis in a Mouse Model of Kawasaki Disease

“…In fact, the C-type lectin-like domain of TM, which lacks the thrombin-binding ability, inhibits the alamin high-mobility group box 1 protein [23] and lipopolysaccharide (LPS) [24] and suppresses neutrophil adhesion to endothelial cells [17], during which rTM promotes the inhibition of the mitogen-activated protein kinase pathway and activation of nuclear factor-κB [17]. Recently, Okamoto et al [25] demonstrated the mechanism underlying rTM inhibition of monocyte adhesion to endothelial cells by suppressing LPS-induced cell sclerosis in a cell stiffness-dependent manner. This may reduce endothelial in ammation and atherosclerosis.…”

Paradigms of endothelial stiffening in cardiovascular disease and vascular aging

“…Furthermore, after cells were returned to normoglycemic conditions for several days, both endothelial stiffness and barrier integrity improved in parallel even though some stiffening and gaps persisted. Endothelial stiffening was also observed in response to lipopolysaccharide or TNFα inflammatory signals with stiff stress fibers identified both by staining and by AFM mapping and 2D rendering ( Okamoto et al, 2020 ). In this study, the integrity of adherens junctions was not evaluated but interestingly they showed that endothelial stiffening is accompanied with decreased functionality of the gap junctions, inter-cellular channels that allow passage of intracellular small molecules between adjacent cells suggesting that endothelial stiffening affects cell-cell communication/interaction via several mechanisms.…”

Advances in mechanical biomarkers

“…Moreover, mechanics play a role early in the immune system maturation. Recent studies have shown how substrate stiffness drives dendritic cell functions, 247 macrophage metabolism, 248 and monocyte maturation and recruitment 249,250 …”

“…Recent studies have shown how substrate stiffness drives dendritic cell functions, 247 macrophage metabolism, 248 and monocyte maturation and recruitment. 249,250 Cell mechanics studies in inflammation started in the past century, when neutrophil stiffening stimulated with chemoattractants, 14 granulocytes cortical tension, 251 and neutrophil kinetics, [252][253][254] among others were described. Since then, further discoveries in diseases have…”