Jana C Mossanen1, Marlene Kohlhepp2, Alexander Wehr3, Oliver Krenkel2, Anke Liepelt2, Anjali A Roeth4, Diana Möckel5, Felix Heymann3, Twan Lammers5, Nikolaus Gassler6, Juliane Hermann7, Joachim Jankowski7, Ulf P Neumann4, Tom Luedde2, Christian Trautwein2, Frank Tacke8. 1. Department of Medicine III, RWTH-University Hospital Aachen, Aachen, Germany; Department of Intensive Care, RWTH-University Hospital Aachen, Aachen, Germany. 2. Department of Medicine III, RWTH-University Hospital Aachen, Aachen, Germany. 3. Department of Medicine III, RWTH-University Hospital Aachen, Aachen, Germany; Department of Hepatology/Gastroenterology, Charité University Medical Center, Berlin, Germany. 4. Department of General, Visceral and Transplantation Surgery, RWTH-University Hospital Aachen, Aachen, Germany. 5. Department of Nanomedicines and Theranostics, Institute for Experimental Molecular Imaging, RWTH-University Hospital Aachen, Aachen, Germany. 6. Institute of Pathology, Clinical Center Braunschweig, Braunschweig, Germany. 7. Institute for Molecular Cardiovascular Research, University Hospital Aachen, Aachen, Germany. 8. Department of Medicine III, RWTH-University Hospital Aachen, Aachen, Germany; Department of Hepatology/Gastroenterology, Charité University Medical Center, Berlin, Germany. Electronic address: frank.tacke@gmx.net.
Abstract
BACKGROUND & AIMS: Inflammation in the liver provokes fibrosis, but inflammation is also important for tumor surveillance. Inhibitors of chemokine pathways, such as CXCL16 and CXCR6 regulation of lymphocyte trafficking, are being tested as antifibrotic agents, but their effects on the development of hepatocellular carcinoma (HCC) are unclear. We assessed the roles of CXCR6-dependent immune mechanisms in hepatocarcinogenesis. METHODS: C57BL/6J wild-type (WT) mice and CXCR6-deficient mice (Cxcr6eGfp/eGfp) were given injections of diethylnitrosamine (DEN) to induce liver cancer and α-galactosylceramide to activate natural killer T (NKT) cells. We also performed studies in mice with conditional, hepatocyte-specific deletion of NEMO, which develop inflammation-associated liver tumors (NemoLPC-KO and NemoLPC-KOCxcr6eGfp/eGfp mice). We collected liver tissues from patients with cirrhosis (n = 43), HCC (n = 35), and neither of these diseases (control individuals, n = 25). Human and mouse liver tissues were analyzed by histology, immunohistochemistry, flow cytometry, RNA expression arrays (from sorted hepatic lymphocytes), and matrix-assisted laser desorption/ionization imaging. Bone marrow was transferred from Cxcr6eGfp/eGfp or WT mice to irradiated C57BL/6J mice, and spleen and liver cells were analyzed by flow cytometry. CD4+ T cells or NKT cells were isolated from the spleen and liver of CD45.1+ WT mice and transferred into CXCR6-deficient mice after DEN injection. RESULTS: After DEN injection, CXCR6-deficient mice had a significantly higher tumor burden than WT mice and increased tumor progression, characterized by reduced intrahepatic numbers of invariant NKT and CD4+ T cells that express tumor necrosis factor and interferon gamma. Livers of NemoLPC-KOCxcr6eGfp/eGfp mice had significantly more senescent hepatocytes than livers of NemoLPC-KO mice. In studies of bone-marrow chimeras, adoptive cell transfer experiments, and analyses of NemoLPC-KO mice, we found that NKT and CD4 T cells promote the removal of senescent hepatocytes to prevent hepatocarcinogenesis, and that this process required CXCR6. Injection of WT with α-galactosylceramide increased removal of senescent hepatocytes by NKT cells. We observed peritumoral accumulation of CXCR6-associated lymphocytes in human HCC, which appeared reduced compared with cirrhosis tissues. CONCLUSIONS: In studies of mice with liver tumors, we found that CXCR6 mediated NKT-cell and CD4+ T-cell removal of senescent hepatocytes. Antifibrotic strategies to reduce CXCR6 activity in liver, or to reduce inflammation or modulate the immune response, should be tested for their effects on hepatocarcinogenesis.
BACKGROUND & AIMS:Inflammation in the liver provokes fibrosis, but inflammation is also important for tumor surveillance. Inhibitors of chemokine pathways, such as CXCL16 and CXCR6 regulation of lymphocyte trafficking, are being tested as antifibrotic agents, but their effects on the development of hepatocellular carcinoma (HCC) are unclear. We assessed the roles of CXCR6-dependent immune mechanisms in hepatocarcinogenesis. METHODS: C57BL/6J wild-type (WT) mice and CXCR6-deficientmice (Cxcr6eGfp/eGfp) were given injections of diethylnitrosamine (DEN) to induce liver cancer and α-galactosylceramide to activate natural killer T (NKT) cells. We also performed studies in mice with conditional, hepatocyte-specific deletion of NEMO, which develop inflammation-associated liver tumors (NemoLPC-KO and NemoLPC-KOCxcr6eGfp/eGfp mice). We collected liver tissues from patients with cirrhosis (n = 43), HCC (n = 35), and neither of these diseases (control individuals, n = 25). Human and mouse liver tissues were analyzed by histology, immunohistochemistry, flow cytometry, RNA expression arrays (from sorted hepatic lymphocytes), and matrix-assisted laser desorption/ionization imaging. Bone marrow was transferred from Cxcr6eGfp/eGfp or WT mice to irradiated C57BL/6J mice, and spleen and liver cells were analyzed by flow cytometry. CD4+ T cells or NKT cells were isolated from the spleen and liver of CD45.1+ WT mice and transferred into CXCR6-deficientmice after DEN injection. RESULTS: After DEN injection, CXCR6-deficientmice had a significantly higher tumor burden than WT mice and increased tumor progression, characterized by reduced intrahepatic numbers of invariant NKT and CD4+ T cells that express tumor necrosis factor and interferon gamma. Livers of NemoLPC-KOCxcr6eGfp/eGfp mice had significantly more senescent hepatocytes than livers of NemoLPC-KO mice. In studies of bone-marrow chimeras, adoptive cell transfer experiments, and analyses of NemoLPC-KO mice, we found that NKT and CD4 T cells promote the removal of senescent hepatocytes to prevent hepatocarcinogenesis, and that this process required CXCR6. Injection of WT with α-galactosylceramide increased removal of senescent hepatocytes by NKT cells. We observed peritumoral accumulation of CXCR6-associated lymphocytes in human HCC, which appeared reduced compared with cirrhosis tissues. CONCLUSIONS: In studies of mice with liver tumors, we found that CXCR6 mediated NKT-cell and CD4+ T-cell removal of senescent hepatocytes. Antifibrotic strategies to reduce CXCR6 activity in liver, or to reduce inflammation or modulate the immune response, should be tested for their effects on hepatocarcinogenesis.
Authors: Giuseppe Visani; Federica Loscocco; Mike Dennis; Eliana Zuffa; Anna Candoni; Alberto Sensi; Barbara Giannini; Gerardo Musuraca; Anna Maria Mianulli; Marino Clavio; Marco Rocchi; Davide Gibellini; Mohsen Navari; Amanda Gilkes; Pier Paolo Piccaluga; Alessandro Isidori Journal: Blood Adv Date: 2020-10-27
Authors: Turan Aghayev; Aleksandra M Mazitova; Jennifer R Fang; Sergei I Grivennikov; Ekaterina K Koltsova; Iuliia O Peshkova; Matthew Rausch; Manhsin Hung; Kerry F White; Ricard Masia; Elizaveta K Titerina; Aliia R Fatkhullina; Isabelle Cousineau; Simon Turcotte; Dmitry Zhigarev; Anastasiia Marchenko; Svetlana Khoziainova; Petr Makhov; Yin Fei Tan; Andrew V Kossenkov; David L Wiest; John Stagg; Xin Wei Wang; Kerry S Campbell; Amiran K Dzutsev; Giorgio Trinchieri; Jonathan A Hill Journal: Cancer Discov Date: 2022-08-05 Impact factor: 38.272
Authors: Guang He Ran; Yu Qing Lin; Lei Tian; Tao Zhang; Dong Mei Yan; Jian Hua Yu; You Cai Deng Journal: Signal Transduct Target Ther Date: 2022-06-29