PMID- 33870156 OWN - NLM STAT- PubMed-not-MEDLINE LR - 20220422 IS - 2589-5559 (Electronic) IS - 2589-5559 (Linking) VI - 3 IP - 3 DP - 2021 Jun TI - Genetic or pharmacological reduction of cholangiocyte senescence improves inflammation and fibrosis in the Mdr2 (-/-) mouse. PG - 100250 LID - 10.1016/j.jhepr.2021.100250 [doi] LID - 100250 AB - BACKGROUND & AIMS: Cholangiocyte senescence is important in the pathogenesis of primary sclerosing cholangitis (PSC). We found that CDKN2A (p16), a cyclin-dependent kinase inhibitor and mediator of senescence, was increased in cholangiocytes of patients with PSC and from a PSC mouse model (multidrug resistance 2; Mdr2 (-/-)). Given that recent data suggest that a reduction of senescent cells is beneficial in different diseases, we hypothesised that inhibition of cholangiocyte senescence would ameliorate disease in Mdr2 (-/-) mice. METHODS: We used 2 novel genetic murine models to reduce cholangiocyte senescence: (i) p16(Ink4a) apoptosis through targeted activation of caspase (INK-ATTAC)xMdr2 (-/-), in which the dimerizing molecule AP20187 promotes selective apoptotic removal of p16-expressing cells; and (ii) mice deficient in both p16 and Mdr2. Mdr2 (-/-) mice were also treated with fisetin, a flavonoid molecule that selectively kills senescent cells. p16, p21, and inflammatory markers (tumour necrosis factor [TNF]-alpha, IL-1beta, and monocyte chemoattractant protein-1 [MCP-1]) were measured by PCR, and hepatic fibrosis via a hydroxyproline assay and Sirius red staining. RESULTS: AP20187 treatment reduced p16 and p21 expression by ~35% and ~70% (p >0.05), respectively. Expression of inflammatory markers (TNF-alpha, IL-1beta, and MCP-1) decreased (by 60%, 40%, and 60%, respectively), and fibrosis was reduced by ~60% (p >0.05). Similarly, p16 (-/-) xMdr2 (-/-) mice exhibited reduced p21 expression (70%), decreased expression of TNF-alpha, IL-1beta (60%), and MCP-1 (65%) and reduced fibrosis (~50%) (p >0.05) compared with Mdr2 (-/-) mice. Fisetin treatment reduced expression of p16 and p21 (80% and 90%, respectively), TNF-alpha (50%), IL-1beta (50%), MCP-1 (70%), and fibrosis (60%) (p >0.05). CONCLUSIONS: Our data support a pathophysiological role of cholangiocyte senescence in the progression of PSC, and that targeted removal of senescent cholangiocytes is a plausible therapeutic approach. LAY SUMMARY: Primary sclerosing cholangitis is a fibroinflammatory, incurable biliary disease. We previously reported that biliary epithelial cell senescence (cell-cycle arrest and hypersecretion of profibrotic molecules) is an important phenotype in primary sclerosing cholangitis. Herein, we demonstrate that reducing the number of senescent cholangiocytes leads to a reduction in the expression of inflammatory, fibrotic, and senescence markers associated with the disease. CI - (c) 2021 The Authors. FAU - Alsuraih, Mohammed AU - Alsuraih M AD - Department of Biochemistry and Molecular Biology, Mayo Clinic, Rochester, MN, 55905, USA. AD - Mayo Clinic Graduate School of Biomedical Sciences, Mayo Clinic, Rochester, MN, 55905, USA. FAU - O'Hara, Steven P AU - O'Hara SP AD - Division of Gastroenterology and Hepatology, and the Mayo Clinic Center for Cell Signaling in Gastroenterology, Mayo Clinic, Rochester, MN, 55905, USA. FAU - Woodrum, Julie E AU - Woodrum JE AD - Division of Gastroenterology and Hepatology, and the Mayo Clinic Center for Cell Signaling in Gastroenterology, Mayo Clinic, Rochester, MN, 55905, USA. FAU - Pirius, Nicholas E AU - Pirius NE AD - Division of Gastroenterology and Hepatology, and the Mayo Clinic Center for Cell Signaling in Gastroenterology, Mayo Clinic, Rochester, MN, 55905, USA. FAU - LaRusso, Nicholas F AU - LaRusso NF AD - Division of Gastroenterology and Hepatology, and the Mayo Clinic Center for Cell Signaling in Gastroenterology, Mayo Clinic, Rochester, MN, 55905, USA. LA - eng GR - P30 DK084567/DK/NIDDK NIH HHS/United States GR - R01 DK057993/DK/NIDDK NIH HHS/United States PT - Journal Article DEP - 20210127 PL - Netherlands TA - JHEP Rep JT - JHEP reports : innovation in hepatology JID - 101761237 PMC - PMC8044431 OTO - NOTNLM OT - ALP, alkaline phosphatase OT - AP, AP20187 OT - Apoptosis resistance OT - BCL2, B cell lymphoma 2 OT - Bcl-xL, B-cell lymphoma-extra large OT - Biliary epithelial cell OT - CCA, cholangiocarcinoma OT - CKI, cyclin-dependent kinase inhibitor OT - Cellular senescence OT - Cholestatic liver disease OT - Col.1A, collagen 1A OT - D, dasatinib OT - EVs, extracellular vesicles OT - FKBP-Casp8, FK506-binding-protein-caspase 8 OT - IF, immunofluorescence OT - INK-ATTAC, p16Ink4a apoptosis through targeted activation of caspase OT - IR, irradiation OT - MCL1, myeloid cell leukemia 1 OT - MCP-1, monocyte chemoattractant protein-1 OT - MMP, matrix metalloproteinase OT - NHC, normal human cholangiocyte OT - PSC, primary sclerosing cholangitis OT - Primary sclerosing cholangitis OT - Q, quercetin OT - RT, reverse transcription OT - SA-beta-gal, senescence-associated beta-gal OT - SASP, senescence-associated secretory phenotype OT - Senescence-associated secretory phenotype OT - Senolytics OT - TNF, tumour necrosis factor OT - WT, wild-type OT - mdr2, multidrug-resistance 2 OT - qPCR, quantitative PCR OT - alpha-SMA, alpha-smooth muscle actin OT - beta-Gal, beta-galactosidase COIS- The authors have no conflict of interest related to the manuscript. Please refer to the accompanying ICMJE disclosure forms for further details. EDAT- 2021/04/20 06:00 MHDA- 2021/04/20 06:01 PMCR- 2021/01/27 CRDT- 2021/04/19 06:41 PHST- 2020/07/28 00:00 [received] PHST- 2021/01/11 00:00 [revised] PHST- 2021/01/12 00:00 [accepted] PHST- 2021/04/19 06:41 [entrez] PHST- 2021/04/20 06:00 [pubmed] PHST- 2021/04/20 06:01 [medline] PHST- 2021/01/27 00:00 [pmc-release] AID - S2589-5559(21)00026-4 [pii] AID - 100250 [pii] AID - 10.1016/j.jhepr.2021.100250 [doi] PST - epublish SO - JHEP Rep. 2021 Jan 27;3(3):100250. doi: 10.1016/j.jhepr.2021.100250. eCollection 2021 Jun.