HGF, MET, and Matrix-Related Proteases in Hepatocellular Carcinoma, Fibrolamellar Variant, Cirrhotic and Normal Liver

Abstract

Fibrolamellar variant is an uncommon subcategory of hepatocellular carcinoma with a better prognostic outcome. Proteinases and growth factors that are involved in the remodeling of extracellular matrix may influence the behavior of cancers. To determine whether these factors contribute to the distinct etiologies of fibrolamellar hepatocellular carcinoma and traditional hepatocellular carcinoma, we assayed hepatocyte growth factor, the hepatocyte growth factor receptor, and two hepatocyte growth factor activators, hepatocyte growth factor activator and urokinase-type plasminogen activator, in hepatocellular carcinoma, fibrolamellar hepatocellular carcinoma, cirrhotic liver and normal liver. In addition, we examined the urokinase-type plasminogen activator receptor, the type 1 plasminogen activator inhibitor, plasmin, fibrinogen, and the type IV matrix metalloproteinases. Eighteen hepatocellular carcinomas and 11 fibrolamellar hepatocellular carcinomas were obtained as paraffin embedded sections from the University of Pittsburgh Department of Pathology. Frozen tissues from a subset of cases (9 hepatocellular carcinomas, 4 fibrolamellar hepatocellular carcinomas, 12 cirrhotic livers and 2 normal livers) were also available for analysis. Antibodies against urokinase-type plasminogen activator, urokinase-type plasminogen activator receptor, hepatocyte growth factor and hepatocyte growth factor receptor were used to analyze immunoperoxidase stained slides from the paraffin blocks. Western blot analyses using antibodies against hepatocyte growth factor, hepatocyte growth factor receptor, phosphotyrosine, hepatocyte growth factor activator, urokinase-type plasminogen activator receptor, urokinase-type plasminogen activator, plasminogen activator inhibitor-1, fibrinogen and plasmin were performed on membrane-enriched fractions from the frozen tissue, as was collagen zymography for matrix metalloproteinase-2 and matrix metalloproteinase-9. The most notable findings are as follows: hepatocyte growth factor activator was only detected in malignant tissue but not cirrhotic liver or normal liver. Although hepatocyte growth factor was detected in most samples, it was significantly elevated in 5/9 hepatocellular carcinomas. Furthermore, 8/9 fibrolamellar hepatocellular carcinomas demonstrated hepatocyte growth factor receptor levels similar to normal, whereas 8/9 hepatocellular carcinomas and 11/12 cirrhotic livers exhibited either an increase or decrease. In contrast, active matrix metalloproteinase-2, which was absent in normal liver, was elevated in fibrolamellar hepatocellular carcinoma as compared to cirrhotic liver and conventional hepatocellular carcinoma. Surprisingly, 10/12 cirrhotic livers and 2/4 fibrolamellar hepatocellular carcinomas but only 1/9 hepatocellular carcinomas were enriched for plasmin. The combined data suggest that the hepatocyte growth factor and plasmin systems tend to be operative in hepatocellular carcinoma and cirrhotic liver, more than fibrolamellar hepatocellular carcinoma. Furthermore, matrix turnover appears to be a more prominent feature of fibrolamellar hepatocellular carcinoma. These findings provide insight into the behavioral differences between hepatocellular carcinoma and the fibrolamellar variant.