The overarching goal of this fellowship application is to develop the skills required to achieve my career goal of becoming an independent investigator with a research focus on cross-talk between the blood coagulation cascade and inflammation. In accordance with this goal, my mentors and I have developed a Research Training Plan to build a strong foundation for conducting research with this focus in the field of liver toxicology. Macrophage-mediated tissue repair is a critical step in the resolution of acute liver injury induced by acetaminophen (APAP) overdose; however, the mechanism by which macrophages attain a pro-repair phenotype in the APAP-injured liver is unknown. We recently discovered that the blood coagulation factor fibrin(ogen) engages leukocyte ?2 integrins, driving repair of the APAP-injured liver. The research training component of this fellowship application will determine the precise mechanisms linking fibrin(ogen)-?2 integrin engagement to pro-repair macrophage activity in the injured liver. My central hypothesis is that cross-linking of fibrin(ogen) by tissue transglutaminase-2 (TGM2) converts fibrin(ogen) to a novel ?2 integrin ligand capable of driving pro-repair macrophage activity in acute liver injury. To test this central hypothesis, I propose two Specific Aims. First, I will determine the mechanism driving pro-repair macrophage function in the APAP- injured liver. My working hypothesis is that fibrin(ogen) engagement of ?2 integrins is critical for intrahepatic pro-repair macrophage programming that drives repair of the APAP-injured liver. To determine the effects of fibrin(ogen) engagement of ?2 integrins on pro-repair macrophage function following APAP overdose, I will use transgenic mice that are deficient in fibrinogen, as well as mice expressing a novel form of fibrinogen lacking the integrin ?2 binding motif. I will also determine the exact heterodimeric ?2 integrin complex required for fibrin(ogen)-directed liver repair by measuring intrahepatic macrophage activation and liver repair in APAP- challenged mice selectively lacking integrin ?M?2 or ?X?2. Second, I will determine the mechanism converting soluble fibrinogen to a ?2 integrin ligand capable of eliciting pro-repair macrophage function. My working hypothesis is that cross-linking of fibrin(ogen) by TGM2 converts fibrin(ogen) to a novel macrophage ?2 integrin ligand that uniquely drives a pro-repair phenotype. I will compare the relative pro-repair effect of soluble fibrinogen, TGM2-cross-linked fibrin(ogen), and fibrin on macrophage pro-repair function in vitro. The studies presented here will identify novel mechanisms of tissue repair, ultimately providing the groundwork for development of targeted therapeutics that expedite patient recovery after APAP overdose. Through completion of these Specific Aims and guidance from my highly-qualified mentoring team, this fellowship application will enable me to develop the technical, intellectual, and professional development skills required to achieve my career goal of being an independent investigator at a research-intensive institution.
Overdose of the commonly used analgesic drug acetaminophen is a leading cause of liver toxicity and liver failure, in part because there have been no major advances in specifc treatments to improve repair of the liver after acute injury. Inflammatory cells known as macrophages are necessary for repair of the injured liver, and macrophage interactions with the blood clotting factor fibrinogen in the injured liver are necessary for tissue repair. The primary goal of this research proposal is to uncover precisely how fibrinogen-macrophage interactions drive repair of the damaged liver so that new therapies can be designed to promote liver repair in patients with liver toxicity.
