Characterizing the role of WISP1 in liver pathophysiology

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Upon acute or chronic exposure to hepatotoxicants, a strong inflammatory response is elicited in the liver. The role of this complex response is not fully understood. In this proposal, we aim to unravel the role of the matricellular protein WISP1 (Wnt-induced secreted protein-1) in acute and chronic liver disease. Our preliminary data indicates that WISP1 mRNA is strongly induced in mouse liver after acute and chronic CCl4 administration, and that WISP1 expression and secretion can be induced in primary mouse hepatocytes by TGFbeta. Furthermore, WISP1 knock out mice showed enhanced liver damage upon acute CCl4 intoxication compared to wild type counterparts, acompanied by lower induction of cytokines (TNFalpha, IL-6, Ccl2) and reduced recriutment of neutrophils to the centrilobular dead cell areas. These results suggest that WISP1 may be an unrecognized modulator of inflammatory responses in liver disease. Our proposal will be focused on four aspects. First, we will determine the cell source of WISP1 on mouse liver after acute injury by CCl4, and the signaling mechanisms controlling its expression. The detection of WISP1 in liver tissue will be performed by in situ hybridization and immunostaining using liver tissue from wild type and WISP1 KO mice. Second, we will determine the role of WISP1 in mouse models of sistemic inflammatory response syndrome, T-cell-mediated hepatitis, fibrosis and hepatocellular carcinoma, using well-established mouse models for each disease. The models will be applied in wild type and WISP1 KO mice. In addition, we will determine the expression of WISP1 in human liver disease samples (tissue and serum) and correlate its expression to disease state. Third, we will investigate the mechanisms by which WISP1 generates a chemoattractant response for leukocytes upon acute liver injury. We will establish which leukocytes become differentially recruited upon liver damage in wild type and WISP1 KO mice by FACS analysis. In addition, we will assess the chemoattractant potential of WISP1 in vitro using primary leukocytes from human donors and analyze the expression of chemokines, cytokines and adhesion molecules in wild type and WISP1 KO mouse liver. Also, we will use 2-photon microscopy to track the movement of fluorescently-labeled neutrophils and macrophages to assess the migration alterations induced in WISP1 KO mice. Fourth, we will identify the signaling and transcriptional pathways mediating WISP1 effects on its target cells, by muliplex assays or immunoblot for phosphorylated signal transducer proteins, and calcium imaging on cells treated with recombinant WISP1 in vitro. We expect that our project will unveil the contribution and the mechanisms by which WISP1 influences immune responses in liver pathophysiology, setting the basis for diagnostic and therapeutic applications focused on this novel molecular player in liver disease.