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Hepatocyte growth factor/scatter factor (HGF/SF) is one of the most important humoral mediators of liver regeneration. It is potentially related to molecular mechanisms of hepatocarcinogenesis via a paracrine system involving its cellular receptor, c-met. In this study, the expression patterns of HGF and c-met were evidenced by multiplex RT-PCR in different specimens of human hepatic tissues (n = 71). A significant increase of c-met mRNA expression was detected in hepatitis (P = 0.001), cirrhosis (P = 0.006), and hepatocellular carcinoma (HCC) tissue (P = 0.003) compared with normal parenchyma and steatosis. HGF mRNA expression was significantly higher only in hepatitis (P = 0.01). Over-expression of c-met mRNA and under-expression of HGF mRNA were detected in the HCCs compared with the corresponding peri-tumoral tissues. Neither HGF nor c-met expression was related to age, sex, tumor size, grading, presence of pseudocapsula, and proliferative activity of the malignant hepatocytes. A significant inverse correlation was found between c-met mRNA expression level and survival (in months) of patients (P = 0.007), as previously shown for urokinase-type plasminogen activator (u-PA) mRNA (P = 0.027). In addition, c-met mRNA expression was strictly associated with u-PA mRNA level in HCC samples (P = 0.001). These data show that a loss of balance concerning HGF, c-met, and u-PA mRNA expression occurs during hepatocarcinogenesis. Particularly, up-regulation of c-met and u-PA mRNA transcription appears to be coordinately regulated, and their levels of expression are inversely correlated with survival; they must therefore play an important role in the development and progression of human HCC and may also be relevant prognostic markers.

Alternative splicing of fibronectin pre-mRNA has been shown to be independently regulated at the EDA and EDB regions in a tissue and developmental stage-specific manner. In this study, RT-PCR approaches were developed for the detection of EDA and EDB FN mRNA isoforms in hepatocarcinoma cells (SK-Hep-I) grown in vitro and in human liver biopsies. While EDA+ and EDB+ isoforms were not present in control adult liver, they were detectable in the hepatocarcinoma cells and in fetal liver. The RT-PCR analysis, extended to biopsies of malignant and non-malignant hepatic tissues, showed that FN mRNAs containing the EDA and EDB sequences were present in the 14 hepatocellular carcinomas (HCCs) tested but absent in the non-tumorous liver tissues (i.e., normal parenchyma, non-specific reactive and chronic hepatitis, steatosis). The EDB+ FN mRNA isoforms were also detected in 3 cases of benign neoplasm (hepatocellular adenoma, HCA, I; nodular focal hyperplasia, NFH, 2), while the EDA+ was only detectable in I of the 2 cases of NFH. In addition, both EDA+ and EDB+ isoforms were expressed in 5 out of 9 cirrhotic livers surrounding the tumors. This molecular analysis, which can also be performed on small liver biopsies (2 mg), may therefore be a useful additional tool in the diagnosis of HCC.