Introduction Vascular adhesion protein (VAP)-1 is an adhesion molecule which possesses potent amine oxidase activity, and deaminates dietary amines resulting in the production of H2O2. Through this function, VAP-1 leads to activation of NFκB in hepatic sinusoidal endothelium (HSEC) resulting in the expression of mucosal-vascular cell-adhesion molecule-1 (MAdCAM-1); a mechanism proposed to contribute to the homing of gut-tropic lymphocytes expressing a4b7 to the liver. Given the putative role this pathway has in hepatic diseases complicating inflammatory bowel disease (IBD), we set out to quantify circulating/soluble (sVAP-1) and intrahepatic VAP-1 enzyme activity in primary sclerosing cholangitis (PSC), and evaluate the functional consequence of its inhibition on MAdCAM-1 dependent lymphocyte recruitment to HSEC.
Methods Total VAP-1 concentration was measured by ELISA. VAP-1 amine oxidase activity was quantified in human serum and explanted liver tissue using the amplex red assay. Flow-based adhesion assays were performed using human HSEC isolated from liver explants, activated with TNFa and methylamine (VAP-1 substrate), and treated with VAP-1 antibody or semicarbazide (VAP-1 enzyme inhibitor). FAC-sorted peripheral blood leucocytes expressing a4b7 were perfused over HSEC under flow rates simulating physiological shear (0.05kPa) and adhesion and transmigration quantified.
Results Patients with PSC had significantly higher circulating median VAP-1 enzyme activity (114.5pmol H2O2 produced/min/ml serum, IQR 100.6–134.7) than patients with IBD (60.3, IQR 38.5–73.0; P = 0.006), normal controls (84.0, IQR 77.7–105.7; P = 0.020) and individuals with PBC (53.9, IQR 33.0–90.9; P = 0.006), and trended higher than AIH (77.6, IQR 51.0–124.5; P = 0.200) (Mann-Whitney). Total sVAP-1 concentration correlated well with sVAP-1 enzyme activity (R2 = 0.75). Intrahepatic median VAP-1 activity was also significantly higher in PSC (97.6pmol H2O2/min/mg protein respectively, IQR 69.5–114.5) vs. PBC (24.6, IQR 18.7–27.8; P = 0.029) and AIH (32.3, IQR 23.3–35.6; P = 0.028) (Mann-Whitney). HSEC pretreatment with semicarbazide but not antibody led to a profound reduction in total a4b7+ lymphocyte adhesion (75%); however, both antibody and enzyme inhibition independently reduced transmigration by ~50% compared to untreated HSEC.
Conclusion sVAP-1 enzyme activity is greater in PSC compared to IBD alone, normal controls, and other immune-mediated liver diseases. Intrahepatic VAP-1 enzyme activity is significantly higher in PSC compared to AIH and PBC. Inhibition of VAP-1 leads to abrogation of a4b7-mediated adhesion to HSEC, representing a putative target for therapeutic intervention in PSC.
Disclosure of Interest None Declared.
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