Preventive effects of the deleted form of hepatocyte growth factor against various liver injuries

Abstract

The effects of a naturally occurring deleted form of hepatocyte growth factor (HGF) on hepatic disorder were studied in various models of hepatic failure. The pretreatment of rats and mice with the deleted form of HGF prevented the liver injuries and coagulopathy induced by endotoxin, dimethylnitrosamine and acetaminophen and reduced the mortality due to hepatic dysfunction induced by these hepatotoxins. The concurrent administration of the deleted form of HGF also prevented the liver injury and hepatic fibrosis in mice treated with α-naphthylisothiocyanate and in rats treated with dimethylnitrosamine. Moreover, the deleted form of HGF normalized the results of the bromosulphalein-clearance test and ameliorated jaundice in rats with periportal cholangiolitic hepatopathy induced by α-naphthylisothiocyanate. The deleted form of HGF also reversed the coagulopathy in rats with hepatic disorder induced by dimethylnitrosamine or by 70% resection of cirrhotic liver (induced by carbon tetrachloride). In Long–Evans cinnamon rats receiving vehicle, 20 out of 21 animals died within 4 days after the onset of jaundice. After infusion of the deleted form of HGF for 4 days, 7 out of 20 Long–Evans cinnamon rats survived. These results indicate that the deleted form of HGF could have therapeutic potency in patients with severe hepatic failure.

Introduction

Patients with terminal liver disease frequently have various combined symptoms associated with severe hepatic dysfunction (Atillasoy and Berk, 1995; Caraceni and Van Thiei, 1995). However, the pathogenesis of terminal-stage liver disease and effective therapeutics for the patients are not still established.

Hepatocyte growth factor (HGF) was initially isolated from rat platelets (Nakamura et al., 1986) and thereafter from the plasma of patients with fulminant hepatic failure (Gohda et al., 1988). A major variant of HGF, the deleted form of HGF, has been purified from the conditioned medium of human fibroblasts (Higashio et al., 1990). The deleted form of HGF is a heparin-binding basic protein with an apparent molecular mass of 80 kDa and is a heterodimer composed of a large α-chain with a apparent molecular mass of 52–56 kDa and a small β-chain with a apparent molecular mass of 30–34 kDa as well as HGF. The deleted form of HGF cDNA, which lacks 15 nucleotides encoding a 5-amino acid residue in the first kringle domain of HGF, has been isolated from the human fibroblast cDNA library (Shima et al., 1991a). The deleted form of HGF is different from HGF in its tertiary structure because the deleted form of HGF-specific antibodies recognize three-dimensional structures newly formed in the protein moiety by the deletion of 5 amino acids (Shima et al., 1994). Matsumoto et al. (1991)have reported that the deleted form of HGF is 1.4-fold more potent than HGF in stimulating DNA synthesis in adult rat hepatocytes in primary culture. It is recognized that the deleted form of HGF and HGF are distinguishable in their target cell specificity and stimulation of growth as follows (Shima et al., 1994). Dose–response curves for the stimulation of DNA synthesis in rat hepatocytes by the deleted form of HGF and HGF are very similar up to about 10 ng/ml, but are significantly different at higher concentrations. HGF activity is markedly decrease over a dose range of 10 to 500 ng/ml, while the deleted form of HGF has maximal activity over the same dose range. The specific activity of the deleted form of HGF is maximally 1.9-fold higher than that of HGF in this dose range. In addition to the difference between the deleted form of HGF and HGF in the stimulation of hepatocytes, the deleted form of HGF is less potent (1/10–1/20) than HGF in the stimulation of DNA synthesis in mesenchymal cells such as human umbilical vein endothelial cells and human aorta smooth muscle cells. These results indicate that the deleted form of HGF may be more specific for the growth of hepatocytes than HGF (Shima et al., 1994).

Since plasma levels of the deleted form of HGF and HGF in patients with liver diseases are higher than those of normal subjects, these growth factors appear to act as the effecter molecules responsible for repairing injured liver tissue (Shima et al., 1991b; Tsubouchi et al., 1991). However, there is no evidence that the deleted form of HGF is useful for treating the principal complication of severe liver diseases. Recently, we showed that the deleted form of HGF ameliorates disordered hepatic protein synthesis in models of liver failure (Masunaga et al., 1996). A systematic investigation should be made to evaluate the possibility of therapeutically using the deleted form of HGF for hepatic disorders generated by various causes. In this study, we examined whether the treatment with the deleted form of HGF could reduce mortality of animals with severe hepatic failure induced by the administration of a lethal dose of several hepatotoxins, ameliorate clinico-chemically hepatic dysfunction and prevent morphological liver injury and hepatic fibrosis in models of liver failure.