Hidenao Yamamoto

Suppression of growth of hepatocellular carcinoma by sodium butyrate in vitro and in vivo

Treatment of HuH‐7 human hepatocellular carcinoma (HCC) cells with 1–10 mM sodium butyrate (SB) resulted in growth inhibition in a dose‐dependent manner. At 3 mM and higher concentrations, SB caused nuclear fragmentation and DNA ladder formation characteristic of apoptosis. In the treated cells, the expression of p21 (WAF1/CIP1) increased and that of α‐fetoprotein (AFP) decreased. These characteristic changes were also observed with 5 other human HCC cell lines with or without mutation of the p53 gene. The ability of these cells to form colonies in soft agar was suppressed by either pretreating the cells with SB prior to soft agar plating or incubating untreated cells in SB‐containing soft agar. Direct injection of SB into tumors developed from HuH‐7 cells in nude mice resulted in an increase in the p21 level, a decrease in the tumor size and an increase in the survival time of mice. When the inoculation of HuH‐7 cells into nude mice was immediately followed by subcutaneous injection of SB, development of tumors was either significantly delayed or completely suppressed. These results suggest that SB induces cellular differentiation and suppresses growth and tumorigenicity of HCC cells in vitro and in vivo by a mechanism independent of p53 but possibly dependent on p21. Int. J. Cancer 76:897–902, 1998.© 1998 Wiley‐Liss, Inc.

Persistent gene expression in rat liver in vivo by repetitive transfections using HVJ-liposome

Most viral vectors are highly immunogenic and are of limited use for somatic gene therapy that requires repetitive administrations. We have developed a highly efficient gene transduction procedure useful for repetitive transfections using liposome containing hemagglutinating virus of Japan (HVJ-liposome). The Escherichia coli β-galactosidase (β-gal) gene was embodied in HVJ-liposome, and introduced directly into the caudal lobe of rat liver that was transiently isolated from a systemic circulation. A 116 kDa β-gal protein was detected in transfected rat liver tissues by Western blot analysis and it was expressed in more than two- thirds of the liver by histological staining. It was found that the transfection efficiency was not affected by repetitive transfections. In support of these findings, antibody response to HVJ-liposome detected in the rat sera was weak and transient. Furthermore, cytotoxic T lymphocytes were not elicited against autologous rat hepatocytes that were transfected in vivo using HVJ-liposome. Thus, our results demonstrate that the isolation of a target liver from systemic circulation and the direct administration of foreign genes using HVJ-liposomes are useful for high gene transduction and persistent gene expression in the liver.

Telomerase activity in hepatocellular carcinoma and adjacent liver tissues

Activation of telomerase and stabilization of telomeres are considered necessary for immortalization of tumor cells. Telomerase activity was analyzed in 69 hepatocellular carcinomas and adjacent chronic liver disease tissues. The telomerase activity level was examined in relation to clinicopathologic features.

HVJ-liposome mediated gene transfer into hepatocytes in vivo

Abstract Background/Aims: The efficient transduction of appropriate target cells will be critical for gene therapy. We evaluated the suitability of hemagglutinating virus of Japan (HVJ)-liposome-mediated gene transfer for gene therapy of liver diseases. Methods: The Escherichia coli β-galactosidase (β-gal) gene was introduced into rat liver by HVJ-liposome to examine gene transfer efficacy and persistence of expression with or without partial hepatectomy prior to transfection. Results: About 30% of hepatocytes were transduced after portal vein injection. Gene expression was transient, with only 2% of hepatocytes expressing β-gal after 4 weeks. However, partial hepatectomy performed 24 h prior to injection resulted in persistently high levels of β-gal for 4 weeks after injection. A 247-bp β-gal polymerase chain reaction fragment transcript was detected in livers of transfected rats, but not in livers of control rats. The rat livers following gene transfer were histologically normal, and serum glutamic-pyruvic transaminase was not found to be elevated in rats. Conclusions: Our results demonstrate that HVJ-liposome-mediated gene transfer produced high gene transduction and persistent gene expression in the liver.

Genomic aberrations in early stage human hepatocellular carcinomas

Primary liver cancer, which most often takes the form of hepatocellular carcinoma (HCC), is among the 10 most common cancers in humans worldwide. In hepatocarcinogenesis, evidence of a multistep process is supported by the marked increase of HCC incidence with age; most HCCs are diagnosed in the second half of life, generally after a long period of chronic liver disease and in frequent association with cirrhosis. This long process may be correlated with the development of multiple genetic lesions, the origin of which currently remain largely unknown. In a previous study, the authors collected data on genomic DNA aberrations in primary HCC by restriction landmark genomic scanning (RLGS), a powerful screening method for the human genome.