Soluble vimentin decreases neutrophil adhesion to platelets by inhibiting P‐selectin glycoprotein ligand‐1‐P‐selectin interactions
Authors: Lam Fong Wilson , Da Qi , Cruz Miguel , Smith C Wayne 
Published: November 2023

Background

Vimentin is an intermediate filament protein mainly thought to be involved in maintaining the cytoskeleton. Recently, there has been increasing evidence of vimentin's role during inflammation, although its precise role is unclear. In a model of murine peritonitis, vimentin‐deficient mice have an increased ability to clear bacteria, suggesting that vimentin somehow inhibits inflammation. This effect may be through inhibiting the interaction between platelets and leukocytes (WBC), since platelets are known to play an important role in the capture and activation of WBC at sites of injury and infection. We and others have described the important role of P‐selectin glycoprotein ligand‐1 (PSGL‐1)‐P‐selectin (P‐sel) interactions on WBC capture by platelets and neutrophil (PMN) transmigration. We hypothesize that soluble vimentin (sVim) decreases PMN‐platelet interactions via interrupting PSGL‐1‐P‐selectin interactions.

Methods

All research was approved by the Institutional Review Board and Institutional Animal Care and Use Committee at Baylor College of Medicine. Recombinant sVim was expressed in E. coli and His‐purified using affinity chromatography. Blood was collected from healthy adult donors and mepacrine‐labelled whole blood or isolated PMN were perfused over fibrinogen, washed platelet monolayers, or P‐selectin/Fc chimeric protein at 2 dyn/cm2 in the presence of varying concentrations of sVim using a microfluidic parallel‐plate flow system (Bioflux). Finally, to determine the role of vimentin on WBC rolling in vivo, mean WBC rolling velocities in both wild type (C57Bl/6) and vimentin−/−mice were measured in cremaster muscle venules (46 ± 0.5 μm) using intravital video microscopy. In vitro data were analyzed as paired t‐test or repeated‐measured analysis of variance, where appropriate, whereas in vivo data were compared using t‐test.

Results

Perfusion of whole blood over fibrinogen resulted in platelet adherence to the fibrinogen coating. WBC adhesion to platelets was reduced in the presence of sVim (50 μg/mL; 29±15% of vehicle control; p<0.05, mean±SEM, n=3). To determine whether this effect was directly related to PMN‐platelet interactions, isolated PMN in the presence of varying concentrations of sVim was perfused over washed platelet monolayers. sVim decreased the number of rolling and adhered PMN on washed platelets in a dose‐dependent fashion (88±22, 35±6, 11±2, 4±2, & 0±0% of vehicle control values for 2.5, 5, 10, 20, & 40 μg/mL, respectively; p<0.05, n=5). To determine whether these observations were due to PSGL‐1‐P‐sel interactions, isolated PMN in the presence varying concentrations of sVim were perfused through P‐sel/Fc chimeric protein (10 μg/mL)‐coated channels. Similar to their effects on PMN‐platelet interactions, sVim significantly decreased PMN adhesion to P‐sel/Fc in a dose‐dependent manner (53±15, 21±3, 16±2, 10±2, 2±1% of vehicle control values for 2.5, 5, 10, 20, & 40 μg/mL, respectively; p<0.05, n=4). In vivo, mean WBC rolling velocities were lower in venules of mice lacking vimentin as compared to wild type mice (42.5±8.8 vs 122.5±23.3 μm/sec; p<0.05, mean±SEM, n=3–4).

Conclusion

Soluble vimentin decreases neutrophil adhesion to platelets by inhibiting PSGL‐1‐P‐selectin interactions. Further studies need to take place to determine how vimentin disrupts WBC capture and rolling and whether vimentin can be used to modulate inflammation caused by platelet‐leukocyte interactions.

Support or Funding Information

NIH grants HL116524, GM0112806, & EY018239 and a Merit Review Grant from the Department of Veterans Affairs