PMID- 35462860 OWN - NLM STAT- PubMed-not-MEDLINE LR - 20220716 IS - 2589-5559 (Electronic) IS - 2589-5559 (Linking) VI - 4 IP - 5 DP - 2022 May TI - High precision-cut liver slice model to study cell-autonomous antiviral defense of hepatocytes within their microenvironment. PG - 100465 LID - 10.1016/j.jhepr.2022.100465 [doi] LID - 100465 AB - BACKGROUND & AIMS: Increased sensitivity towards tumor necrosis factor (TNF)-induced cell death in virus-infected hepatocytes has revealed a so far unrecognized hepatocyte-intrinsic antiviral immune surveillance mechanism, for which no in vitro or ex vivo model is available. We aimed to establish precision-cut liver slices (PCLS) as a model system to study hepatocyte-intrinsic regulation of apoptosis. METHODS: Preparation of PCLS from mouse and human liver tissue was optimized for minimal procedure-associated apoptosis. Functionality of liver cells in PCLS was characterized using extracellular flux analysis to determine mitochondrial respiration, and viral infection with recombinant adenovirus and lymphocytic choriomeningitis virus (LCMV) was used to probe for hepatocyte-intrinsic sensitivity towards apoptosis in PCLS. Apoptosis was detected by immunohistochemical staining for cleaved-caspase 3 and quantified by detection of effector caspase activity in PCLS. RESULTS: We established an optimized protocol for preparation of PCLS from human and mouse models using agarose-embedding of liver tissue to improve precision cutting and using organ-protective buffer solutions to minimize procedure-associated cell death. PCLS prepared from virus-infected livers showed preserved functional metabolic properties. Importantly, in PCLS from adenovirus- and LCMV-infected livers we detected increased induction of apoptosis after TNF challenge ex vivo. CONCLUSION: We conclude that PCLS can be used as model system to ex vivo characterize hepatocyte-intrinsic sensitivity to cell death. This may also enable researchers to characterize human hepatocyte sensitivity to apoptosis in PCLS prepared from patients with acute or chronic liver diseases. LAY SUMMARY: Virus-infected hepatocytes in vivo show an increased sensitivity towards induction of cell death signaling through the TNF receptor. Studying this hepatocyte-intrinsic antiviral immune surveillance mechanism has been hampered by the absence of model systems that reciprocate the in vivo finding of increased apoptosis of virus-infected hepatocytes challenged with TNF. Herein, we report that an optimized protocol for generation of precision-cut liver slices can be used to study this hepatocyte-intrinsic surveillance mechanism ex vivo. CI - (c) 2022 The Authors. FAU - Brugger, Marcus AU - Brugger M AD - Institute of Molecular Immunology and Experimental Oncology, School of Medicine, Technical University of Munich, Germany. AD - Department of Internal Medicine I, School of Medicine, University Hospital Munchen rechts der Isar, Technical University of Munich, Germany. FAU - Laschinger, Melanie AU - Laschinger M AD - Department of Surgery, School of Medicine, University Hospital Munchen rechts der Isar; Technical University of Munich, Germany. FAU - Lampl, Sandra AU - Lampl S AD - Institute of Molecular Immunology and Experimental Oncology, School of Medicine, Technical University of Munich, Germany. FAU - Schneider, Annika AU - Schneider A AD - Institute of Molecular Immunology and Experimental Oncology, School of Medicine, Technical University of Munich, Germany. FAU - Manske, Katrin AU - Manske K AD - Institute of Molecular Immunology and Experimental Oncology, School of Medicine, Technical University of Munich, Germany. FAU - Esfandyari, Dena AU - Esfandyari D AD - Institute of Pharmacology and Toxicology, Technical University Munich, Munich, Germany. FAU - Huser, Norbert AU - Huser N AD - Department of Surgery, School of Medicine, University Hospital Munchen rechts der Isar; Technical University of Munich, Germany. FAU - Hartmann, Daniel AU - Hartmann D AD - Department of Surgery, School of Medicine, University Hospital Munchen rechts der Isar; Technical University of Munich, Germany. FAU - Steiger, Katja AU - Steiger K AD - Institute of Pathology, Technical University of Munich, Germany. FAU - Engelhardt, Stefan AU - Engelhardt S AD - Institute of Pharmacology and Toxicology, Technical University Munich, Munich, Germany. FAU - Wohlleber, Dirk AU - Wohlleber D AD - Institute of Molecular Immunology and Experimental Oncology, School of Medicine, Technical University of Munich, Germany. FAU - Knolle, Percy A AU - Knolle PA AD - Institute of Molecular Immunology and Experimental Oncology, School of Medicine, Technical University of Munich, Germany. LA - eng PT - Journal Article DEP - 20220306 PL - Netherlands TA - JHEP Rep JT - JHEP reports : innovation in hepatology JID - 101761237 PMC - PMC9019249 OTO - NOTNLM OT - IP3, inositol-3-phosphate OT - LCMV, lymphocytic choriomeningitis virus OT - PCLS, precision-cut liver slices OT - PLCg, phospholipase C gamma OT - ROS, reactive oxygen species OT - TNF, tumor necrosis factor OT - TNF-induced apoptosis OT - anti-viral immunity OT - precision-cut liver slices COIS- The authors declare no conflict of interest. Please refer to the accompanying ICMJE disclosure forms for further details. EDAT- 2022/04/26 06:00 MHDA- 2022/04/26 06:01 PMCR- 2022/03/06 CRDT- 2022/04/25 05:12 PHST- 2021/10/21 00:00 [received] PHST- 2022/02/07 00:00 [revised] PHST- 2022/02/14 00:00 [accepted] PHST- 2022/04/25 05:12 [entrez] PHST- 2022/04/26 06:00 [pubmed] PHST- 2022/04/26 06:01 [medline] PHST- 2022/03/06 00:00 [pmc-release] AID - S2589-5559(22)00037-4 [pii] AID - 100465 [pii] AID - 10.1016/j.jhepr.2022.100465 [doi] PST - epublish SO - JHEP Rep. 2022 Mar 6;4(5):100465. doi: 10.1016/j.jhepr.2022.100465. eCollection 2022 May.