Project Summary/Abstract The goal of this proposal is to extend my training as an independent scientist in the field of hepatic manifestations of Sickle cell disease (SCD). To this end, I have selected the division of Hematology-Oncology at University of Pittsburgh to continue my transition to become an independent investigator at an academic institution. This proposal outlines an extensive Research Strategy that is complemented by several areas of training, which includes several courses directly related to my studies in Specific Aims, meeting with my junior faculty research committee, and attendance and participation of multiple seminars throughout the University Of Pittsburgh Medical School. My Research Strategy will determine the molecular mechanisms of SCD induced hepatic crisis. SCD is an autosomal-recessive monogenic disorder that affects approximately 100,000 Americans and millions of people worldwide. Sinusoidal vaso-occlusion and hemolysis are considered as chief contributors of sickle hepatic crisis. Hepatic crisis affects 10-40% of hospitalized SCD patients which is characterized by liver injury and sickle cell intrahepatic cholestasis (SCIC) that can progress to fatal liver failure. The current treatment for hepatic crisis is primarily supportive, and the molecular mechanism is unknown, suggesting that preventive therapies based on the improved understanding of the molecular pathways that enable SCIC are needed. In this study, we have used a transgenic, humanized mouse model of SCD that exclusively expresses sickle human hemoglobin. Preliminary findings reveal that SCD mice developed chronic liver injury with age, which was manifested by sustained inflammation, hyperbilirubinemia and cholestasis. Using our recently developed real-time in vivo imaging of the intact liver of live mice, we discovered the presence of sinusoidal ischemia and impaired bile transport across the apical membrane of hepatocytes in SCD mice. The impaired bile transport was associated with loss of apical bile transporters (BSEP, ABCG5 and ABCG8) from hepatocytes. RNA-seq analysis identified dysregulation of genes encoding proteins responsible for inflammation and bile secretion in the liver of SCD mice. Furthermore, we observed NF-κB activation in the liver of SCD mice inhibited FXR signaling and its downstream targets, leading to impaired bile secretion. These findings form the basis for my overarching hypothesis that ischemia and hemolysis induced inflammation, tissue injury and oxidative stress promotes NF-kB activation in sickle hepatocytes which inhibits FXR signaling leading to impaired bile secretion in SCD, and activating FXR or rescuing bile secretion can ameliorate SCIC. This hypothesis will be tested in the following aims: 1) To determine whether ischemia-reperfusion injury and hemolysis promotes hepatocyte specific activation of NF-kB in SCD and 2) To determine whether hepatocyte-specific activation of NF-kB promotes loss of FXR-signaling leading to ...