Macrophage secretory products induce an inflammatory phenotype in hepatocytes

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)-beta 1 (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-beta 1 (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 Refereed/Peer-reviewed