Background: It has been shown that expression of the potent angiogenic factor, vascular endothelial growth factor (VEGF), and its receptors, flt-1 (VEGFR-1) and KDR/Flk-1 (VEGFR-2), increased during the development of liver fibrosis. Aims: To elucidate the in vivo role of interaction between VEGF and its receptors in liver fibrogenesis. Methods: A model of CCl ₄ induced hepatic fibrosis was used to assess the role of VEGFR-1 and VEGFR-2 by means of specific neutralising monoclonal antibodies (R-1mAb and R-2mAb, respectively). R-1mAb and R-2mAb were administered after two weeks of treatment with CCl ₄ , and indices of fibrosis were assessed at eight weeks. Results: Hepatic VEGF mRNA expression significantly increased during the development of liver fibrosis. Both R-1mAb and R-2mAb treatments significantly attenuated the development of fibrosis associated with suppression of neovascularisation in the liver. Hepatic hydroxyproline and serum fibrosis markers were also suppressed. Furthermore, the number of α-smooth muscle actin positive cells and α1(I)-procollagen mRNA expression were significantly suppressed by R-1mAb and R-2mAb treatment. The inhibitory effect of R-2mAb was more potent than that of R-1mAb, and combination treatment with both mAbs almost completely attenuated fibrosis development. Our in vitro study showed that VEGF treatment significantly stimulated proliferation of both activated hepatic stellate cells (HSC) and sinusoidal endothelial cells (SEC). VEGF also significantly increased α1(I)-procollagen mRNA expression in activated HSC. Conclusions: These results suggest that the interaction of VEGF and its receptor, which reflected the combined effects of both on HSC and SEC, was a prerequisite for liver fibrosis development.

Tissue inhibitor of metalloproteinases–1 (TIMP–1) has been shown to be increased in liver fibrosis development both in murine experimental models and human samples. However, the direct role of TIMP–1 during liver fibrosis development has not been defined. To address this issue, we developed transgenic mice overexpressing human TIMP–1 (hTIMP–1) in the liver under control of the albumin promoter/enhancer. A model of CCl₄–induced hepatic fibrosis was used to assess the extent of fibrosis development in TIMP–1 transgenic (TIMP–Tg) mice and control hybrid (Cont) mice. Without any treatment, overexpression of TIMP–1 itself did not induce liver fibrosis. There were no significant differences of pro–(α1)–collagen–I, (α2)–collagen–IV, and α–smooth muscle actin (α–SMA) mRNA expression in the liver between TIMP–Tg and Cont–mice, suggesting that overexpression of TIMP–1 itself did not cause hepatic stellate cell (HSC) activation. After 4–week treatment with CCl₄, however, densitometric analysis revealed that TIMP–Tg–mice had a seven–fold increase in liver fibrosis compared with the Cont–mice. The hepatic hydroxyproline content and serum hyaluronic acid were also significantly increased in TIMP–Tg–mice, whereas CCl₄–induced liver dysfunction was not altered. An active form of matrix metalloproteinases–2 (MMP–2) level in the liver of TIMP–Tg–mice was decreased relative to that in Cont–mice because of the transgenic TIMP–1. Immunohistochemical analysis revealed that collagen–I and collagen–IV accumulation was markedly increased in the liver of CCl₄–treated TIMP–Tg–mice with a pattern similar to that of α–SMA positive cells. These results suggest that TIMP–1 does not by itself result in liver fibrosis, but strongly promotes liver fibrosis development.

Alcohol is the most common cause of liver disease in the world. Chronic alcohol consumption leads to hepatocellular injury and liver inflammation. Inflammatory cytokines, such as TNF- and IFN-, induce liver injury in the rat model of alcoholic liver disease (ALD). Hepatoprotective cytokines, such as IL-6, and anti-inflammatory cytokines, such as IL-10, are also associated with ALD. IL-6 improves ALD via activation of the signal transducer and activator of transcription 3 (STAT3) and the subsequent induction of a variety of hepatoprotective genes in hepatocytes. IL-10 inhibits alcoholic liver inflammation via activation of STAT3 in Kupffer cells and the subsequent inhibition of liver inflammation. Alcohol consumption promotes liver inflammation by increasing translocation of gut-derived endotoxins to the portal circulation and activating Kupffer cells through the LPS/Toll-like receptor (TLR) 4 pathways. Oxidative stress and microflora products are also associated with ALD. Interactions between pro- and anti-inflammatory cytokines and other cytokines and chemokines are likely to play important roles in the development of ALD. The present study aims to conduct a systemic review of ALD from the aspect of inflammation.

Nonalcoholic steatohepatitis (NASH) may cause fibrosis, cirrhosis, and hepatocellular carcinoma (HCC); however, the exact mechanism of disease progression is not fully understood. Angiogenesis has been shown to play an important role in the progression of chronic liver disease. The aim of this study was to elucidate the role of angiogenesis in the development of liver fibrosis and hepatocarcinogenesis in NASH. Zucker rats, which naturally develop leptin receptor mutations, and their lean littermate rats were fed a choline-deficient, amino acid–defined diet. Both Zucker and littermate rats showed marked steatohepatitis and elevation of oxidative stress markers (e.g., thiobarbital acid reactive substances and 8-hydroxydeoxyguanosine). In sharp contrast, liver fibrosis, glutathione-S-transferase placental form (GST-P)-positive preneoplastic lesions, and HCC developed in littermate rats but not in Zucker rats. Hepatic neovascularization and the expression of vascular endothelial growth factor (VEGF), a potent angiogenic factor, only increased in littermate rats, almost in parallel with fibrogenesis and carcinogenesis. The CD31-immunopositive neovessels were mainly localized either along the fibrotic septa or in the GST-P–positive lesions. Our in vitro study revealed that leptin exerted a proangiogenic activity in the presence of VEGF. In conclusion, these results suggest that leptin-mediated neovascularization coordinated with VEGF plays an important role in the development of liver fibrosis and hepatocarcinogenesis in NASH. (HEPATOLOGY 2006;44:983–991.)