The OVERALL OBJECTIVE of this renewal is to clarify the pathogenesis of Primary Sclerosing Cholangitis (PSC) and develop effective treatments. PSC affects 30,000 Americans, accounts for 6% of liver transplants, and has a median survival of 13-20 years. Characterized by a fibroinflammatory peribiliary microenvironment, it progresses to cirrhosis, is associated with inflammatory bowel disease, and has an increased risk of colon, bile duct, and gallbladder cancers. There are no approved drugs for PSC because its pathogenesis is obscure. While cholangiocytes are important in pathogenesis, how they contribute to PSC initiation and progression, and what cellular processes and molecules are involved is unclear. We reported that: i) cellular senescence (cell cycle arrest) and the senescence associated secretory phenotype (SASP) are features of cholangiocytes in PSC patients and animal models; and ii) key regulatory mechanisms include a central molecular pathway (NRAS/MEK/ERK), upregulation of a transcription factor, ETS1, and epigenome modifications (loss of H3K27me3). These results suggest that ETS1 functions as an essential mediator of cholangiocyte senescence. We also found that: i) induction of cholangiocyte senescence in vitro causes hypersecretion of fibroinflammatory molecules (i.e., SASP) and apoptosis resistance from an imbalance of pro- and anti- apoptotic proteins. RNAseq on cholangiocytes from PSC patients showed upregulation of genes associated with SASP and apoptosis resistance and enrichment of ETS1 target sites within their promoters. By ChIP- qPCR, we found that ETS1 and the histone acetyltransferase, p300, interact and localize at senescence, prosurvival, and SASP associated loci. Finally: i) in vitro, pro-apoptotic, BH3-only mimetics, kill senescent but not non-senescent cholangiocytes; and ii) in vivo, killing senescent cholangiocytes using either novel, genetic cross-breeding (Mdr2-/--INK-ATTAC mouse) or pharmacologic treatment (BH3 mimetic) reduces peribiliary fibrosis. Thus, our data support the CENTRAL HYPOTHESIS that in PSC, ETS1 promotes chromatin remodeling and selective transcription inducing cholangiocyte senescence, apoptosis resistance, and SASP resulting in hypersecretion of bioactive molecules that induce a fibroinflammatory peribiliary microenvironment. We will test this hypothesis using novel in vitro biochemical and molecular techniques, and in vivo using animal models. We have 3 SPECIFIC AIMS. First, we test the hypothesis that ETS1 and p300 mediate cholangiocyte senescence. Second, we test the hypothesis that ETS1 mediates apoptosis resistance by upregulation of pro-survival proteins, including BCL- XL. Third, we test the hypothesis that ETS1 mediates cholangiocyte SASP promoting a fibroinflammatory peribiliary microenvironment through upregulation of CXCL12. Results from our innovative experiments will clarify the pathways and molecules involved in the development of PSC and could lead to novel therapies targeting a PSC cholangiocyte signature of senescence, apoptosis resistance, and SASP.

Public Health Relevance

This project explores mechanisms by which the cells that line bile ducts (cholangiocytes) influence progression of Primary Sclerosing Cholangitis (PSC), a progressive liver disease with no effective therapy except liver transplant. We explore three interconnected and druggable cellular pathways: i) senescence (cell cycle arrest); ii) resistance to cell death; and, iii) over-production of inflammatory molecules. If successful, the results could lead to new and effective treatments.

National Institute of Health (NIH)
National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK)
Research Project (R01)
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Hepatobiliary Pathophysiology Study Section (HBPP)
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Sherker, Averell H
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Mayo Clinic, Rochester
United States
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Guicciardi, Maria Eugenia; Trussoni, Christy E; Krishnan, Anuradha et al. (2018) Macrophages contribute to the pathogenesis of sclerosing cholangitis in mice. J Hepatol 69:676-686
Moncsek, Anja; Al-Suraih, Mohammed S; Trussoni, Christy E et al. (2018) Targeting senescent cholangiocytes and activated fibroblasts with B-cell lymphoma-extra large inhibitors ameliorates fibrosis in multidrug resistance 2 gene knockout (Mdr2-/- ) mice. Hepatology 67:247-259
Cheung, Angela C; Lorenzo Pisarello, Maria J; LaRusso, Nicholas F (2018) Pathobiology of biliary epithelia. Biochim Biophys Acta Mol Basis Dis 1864:1220-1231
Cheung, Angela C; LaRusso, Nicholas F; Gores, Gregory J et al. (2017) Epigenetics in the Primary Biliary Cholangitis and Primary Sclerosing Cholangitis. Semin Liver Dis 37:159-174
Loarca, Lorena; De Assuncao, Thiago M; Jalan-Sakrikar, Nidhi et al. (2017) Development and characterization of cholangioids from normal and diseased human cholangiocytes as an in vitro model to study primary sclerosing cholangitis. Lab Invest 97:1385-1396
Cheung, Angela C; Lazaridis, Konstantinos N; LaRusso, Nicholas F et al. (2017) Emerging pharmacologic therapies for primary sclerosing cholangitis. Curr Opin Gastroenterol 33:149-157
O'Hara, Steven P; Splinter, Patrick L; Trussoni, Christy E et al. (2017) ETS Proto-oncogene 1 Transcriptionally Up-regulates the Cholangiocyte Senescence-associated Protein Cyclin-dependent Kinase Inhibitor 2A. J Biol Chem 292:4833-4846
Gradilone, Sergio A; Pisarello, Maria J Lorenzo; LaRusso, Nicholas F (2017) Primary Cilia in Tumor Biology: The Primary Cilium as a Therapeutic Target in Cholangiocarcinoma. Curr Drug Targets 18:958-963
O'Hara, Steven P; La Russo, Nicholas F (2017) Cellular senescence, neuropeptides and hepatic fibrosis: Additional insights into increasing complexity. Hepatology 66:318-320
Lazaridis, Konstantinos N; LaRusso, Nicholas F (2016) Primary Sclerosing Cholangitis. N Engl J Med 375:2501-2502

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