Abstract
Abstract Thrombin is the main effector protease in the coagulation cascade. It can also signal via protease activating receptors (PAR). The presence of these receptors on the surface of innate immune cells has been well reported but the functional consequence of activation has yet to be fully defined. This study aims to investigate the role thrombin has on innate immune cell function using murine bone marrow macrophages (BMM). In Vitro, stimulating mature MCSF-cultured BMM with thrombin did not affect gross markers of macrophage polarisation (iNOS or CD206). The stimulated cell supernatants contained increased amounts of Interferon gamma (IFNγ) but reduced IL10. Thrombin increased IFNγ receptor expression at the cell surface. Thrombin-treated cells had increased lipid rich microdomains by Cholera Toxin B staining and increased co-localisation of the LPS receptor within the lipid rafts. Compared to untreated cells, thrombin stimulated cells were highly sensitive to low dose M1 polarising stimuli, as evidenced by iNOS expression. The thrombin treated cells down regulated surface ABCA1 expression, but this was prevented by transfecting cells with siRNA against Cullin 3. This preserved ABCA1 expression and prevented the increase in lipid rich microdomains after thrombin stimulation and was associated with the loss of heightened sensitivity to low dose M1 stimuli. Taken together this shows a clear pro inflammatory signal on the thrombin treated cells and to date the first description of ABCA1’s key role in thrombin mediated inflammatory signalling.
Published Version
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