TRP X14 Role of the Xase complex and prothrombotic platelet function in small intestinal mucositis

Funding period: 01.08.2016. - 31.12.2016

Project Summary

The risk of venous thrombosis is increased by 3-fold in patients with IBD. Under normal steady-state conditions, microbial signatures leak into the portal circulation, but even more if the epithelial barrier is perturbed as in IBD. In our previous experiments we identified that FIX activity is increased in the plasma of conventionally-raised (CONV-R) mice compared with their germ-free (GF) counterparts, whereas total FIX protein levels were unchanged. Furthermore, we found that FVIIIa levels are increased in CONV-R mice suggesting increased formation of the Xase complex (FVIIIa/FIXa) forming an important amplification loop that augments conversion of FX into FXa. The microbial signaling that augments the release of FVIII from liver endothelial cells is unexplored. We hypothesize that his microbiota-dependent regulation of components that constitute the Xase complex might be of particular relevance for the prothrombotic state observed in the acute phases of IBD, since in particular FVIIIa levels were found increased in blood of Crohn’s disease patients  and hemophiliacs are protected against development of IBD. Although Tissue Factor (TF)-dependent coagulation activation was related to disease active IBD, the role of the Josso pathway (amplification loop of Xa formation via the Xase complex) and Xase-dependent augmentation of thrombin formation under conditions of acute intestinal Inflammation  has not been studied. Primary liver endothelial cells will be isolated from livers of GF and CONV-R mice and the release of FVIII will be studied by stimulation with bacterial pathogen-associated molecular patterns. To study the role of FVIII and FIX for the d velopment of acute DSS-induced mucositis, we will take advantage of F8-/- and F9-/- mice and WT littermate controls that we have established as mouse colonies at the Translational Animal Research Facility (TARC). It will be interesting to study, if mice that show impaired thrombin formation due to their inability of Xase complex formation are protected against DSS-induced mucosal injury and acute intestinal inflammation. To investigate the role of the Xase complex formed on tissue factor-activated platelets we will investigate the thrombin generation potential in PRP and PPP of GF and CONV-R mice, as well as in untreated and 3.5% DSS-treated F8-/- and F9-/- mice, as we hypothesize that Xase complex formation is reduced under germ-free conditions and under non-inflamed conditions. Collectively, our experiments with F8-/- and F9-/- mice will contribute to understand the role of the gut microbiota, inflammation that is induced by perturbed intestinal barrier function and   the Xase complex for hypercoagulability and the increased risk of venous thrombosis in IBD. 

Principle Investigator