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©2012 Baishideng Publishing Group Co., Limited. All rights reserved.
Macrophage secretory products induce an inflammatory phenotype in hepatocytes
Michelle Melino, Victoria L Gadd, Gene V Walker, Richard Skoien, Helen D Barrie, Dinesh Jothimani, Leigh Horsfall, Alun Jones, Matthew J Sweet, Gethin P Thomas, Andrew D Clouston, Julie R Jonsson, Elizabeth E Powell
Michelle Melino, Victoria L Gadd, Gene V Walker, Richard Skoien, Helen D Barrie, Dinesh Jothimani, Andrew D Clouston, Julie R Jonsson, Elizabeth E Powell, Centre for Liver Disease Research, School of Medicine, The University of Queensland, Princess Alexandra Hospital, Brisbane 4102, Queensland, Australia
Richard Skoien, Leigh Horsfall, Elizabeth E Powell, Department of Gastroenterology and Hepatology, Princess Alexandra Hospital, Brisbane 4102, Queensland, Australia
Alun Jones, Matthew J Sweet, Institute for Molecular Bioscience and Australian Infectious Diseases Research Centre, The University of Queensland, Brisbane 4072, Queensland, Australia
Gethin P Thomas, Diamantina Institute, The University of Queensland, Brisbane 4102, Queensland, Australia
Author contributions: Powell EE, Jonsson JR and Clouston AD conceived the study; Powell EE, Jonsson JR, Melino M and Clouston AD designed the study; Melino M, Powell EE, Jonsson JR and Clouston AD wrote the manuscript and gave final approval; Melino M, Gadd VL, Walker GV, Skoien R, Barrie HD and Jothimani D performed the laboratory analyses, analysed and interpreted the results and approved the manuscript; Horsfall L recruited patients and processed and managed the clinical data; Jones A designed, performed, analysed and interpreted the HPLC/MS/MS experiments; Thomas GP designed, analysed and interpreted the microarray experiments and contributed to the writing of the manuscript; Melino M performed the microarray experiments; Sweet MJ contributed to the design of the study and writing, and approval of the manuscript.
Supported by The National Health and Medical Research Council of Australia, No. APP1003108; the Queensland Government’s Smart State Health and Medical Research Fund; The Princess Alexandra Hospital Research and Development Foundation and The Sasakawa Foundation (Royal Children’s Hospital, Brisbane); an Unrestricted Education Grant from MSD (to Powell EE); a Lions Medical Research Foundation Senior Research Fellowship (to Thomas GP)
Correspondence to: Elizabeth Powell, Professor, Department of Gastroenterology and Hepatology, Princess Alexandra Hospital, Woolloongabba 4102, Queensland, Australia. e.powell@uq.edu.au
Telephone: +61-7-32402035 Fax: +61-7-32402337
Received: June 23, 2011
Revised: September 20, 2011
Accepted: September 27, 2011
Published online: April 21, 2012
AIM: To investigate the influence of macrophages on hepatocyte phenotype and function.
METHODS: Macrophages were differentiated from THP-1 monocytes via phorbol myristate acetate stimulation and the effects of monocyte or macrophage-conditioned medium on HepG2 mRNA and protein expression determined. The in vivo relevance of these findings was confirmed using liver biopsies from 147 patients with hepatitis C virus (HCV) infection.
RESULTS: Conditioned media from macrophages, but not monocytes, induced a transient morphological change in hepatocytes associated with upregulation of vimentin (7.8 ± 2.5-fold, P = 0.045) and transforming growth factor (TGF)-β1 (2.6 ± 0.2-fold, P < 0.001) and downregulation of epithelial cadherin (1.7 ± 0.02-fold, P = 0.017) mRNA expression. Microarray analysis revealed significant upregulation of lipocalin-2 (17-fold, P < 0.001) and pathways associated with inflammation, and substantial downregulation of pathways related to hepatocyte function. In patients with chronic HCV, real-time polymerase chain reaction and immunohistochemistry confirmed an increase in lipocalin-2 mRNA (F0 1.0 ± 0.3, F1 2.2 ± 0.2, F2 3.0 ± 9.3, F3/4 4.0 ± 0.8, P = 0.003) and protein expression (F1 1.0 ± 0.5, F2 1.3 ± 0.4, F3/4 3.6 ± 0.4, P = 0.014) with increasing liver injury. High performance liquid chromatography-tandem mass spectrometry analysis identified elevated levels of matrix metalloproteinase (MMP)-9 in macrophage-conditioned medium, and a chemical inhibitor of MMP-9 attenuated the change in morphology and mRNA expression of TGF-β1 (2.9 ± 0.2 vs 1.04 ± 0.1, P < 0.001) in macrophage-conditioned media treated HepG2 cells. In patients with chronic HCV infection, hepatic mRNA expression of CD163 (F0 1.0 ± 0.2, F1/2 2.8 ± 0.3, F3/4 5.3 ± 1.0, P = 0.001) and MMP-9 (F0 1.0 ± 0.4, F1/2 2.8 ± 0.3, F3/4 4.1 ± 0.8, P = 0.011) was significantly associated with increasing stage of fibrosis.
CONCLUSION: Secreted macrophage products alter the phenotype and function of hepatocytes, with increased expression of inflammatory mediators, suggesting that hepatocytes actively participate in liver injury.
