ABSTRACT
To elucidate metastasis mechanisms, we established a Panc-1H5 subline with a highly liver metastatic cell line and a Panc-1P4a with a highly peritoneal metastatic cell line, which were sequentially selected from the parental pancreatic cancer cell line Panc-1. Using these three cell lines, we investigated several biological properties and mRNA levels of differentially-expressed genes involved in cancer metastasis with a cDNA macroarray. The tumorigenicity, motile activity, adhesive activity and cytokine production of metastatic sublines were higher than those of parental Panc-1 cells. Particularly, in Panc-1H5 cells, adhesive activity to the extracellular matrix and angiogenetic factors increased, whereas in Panc-1P4a cells, motile activity was extremely enhanced compared with Panc-1 cells. Histopathological findings for the three cell lines were the same. In cDNA macroarray analysis of Panc-1H5 cells, 11 genes were up-regulated and 20 genes were down-regulated compared with parental Panc-1 cells. In Panc-1P4a cells, 7 genes were up-regulated and 13 genes were down-regulated compared with parental Panc-1 cells. This study provides a demonstration of global gene expression analysis of pancreatic cancer cells with liver and peritoneal metastasis and these results provide new insight into the study of human pancreatic cancer metastasis.
Subject(s)
Gene Expression Profiling , Gene Expression Regulation, Neoplastic , Liver Neoplasms/genetics , Liver Neoplasms/pathology , Pancreatic Neoplasms/pathology , Peritoneal Neoplasms/genetics , Peritoneal Neoplasms/pathology , Animals , Cell Adhesion , Cell Differentiation , Cytokines/biosynthesis , Female , Humans , Liver Neoplasms/secondary , Mice , Mice, Inbred BALB C , Mice, Nude , Neoplasm Metastasis/genetics , Neoplasm Metastasis/pathology , Neoplasm Transplantation , Peritoneal Neoplasms/secondary , Tumor Cells, CulturedABSTRACT
We established a new cell line, NUGC-3P4T, with high peritoneal metastatic disseminating potential in nude mice. NUGC-3P4T cells were derived from the human gastric carcinoma line NUGC-3, which has low capacity for peritoneal dissemination. NUGC-3P4T cells developed peritoneal dissemination in 10 / 10 (100%) mice, whereas the parental NUGC-3 cells developed dissemination in 1 / 5 (20.0%) mice. The metastatic foci in the peritoneum showed essentially the same histological appearance as those induced by parental cells. The tumorigenicity, the motile activity and the adhesive activity to the laminin of NUGC-3P4T cells were stronger than those of NUGC-3 cells. Production of IL-8 was significantly higher in NUGC-3P4T than in NUGC-3. cDNA macroarrays analysis showed that a variety of cytokines, interleukins, and other immunomodulators and their receptors were up- or down-regulated at the mRNA level in NUGC-3P4T cells, compared with NUGC-3 cells. Thus, this unique cell line and in vivo model might be useful to study the biology of peritoneal dissemination of human gastric cancer.
Subject(s)
Oligonucleotide Array Sequence Analysis , Peritoneal Neoplasms/secondary , Stomach Neoplasms/pathology , Tumor Cells, Cultured , Animals , Cell Adhesion/physiology , Cell Movement/physiology , Disease Models, Animal , Endothelial Growth Factors/biosynthesis , Endothelial Growth Factors/genetics , Female , Gene Expression Regulation, Neoplastic , Humans , Interleukin-8/biosynthesis , Interleukin-8/genetics , Lymphokines/biosynthesis , Lymphokines/genetics , Mice , Mice, Inbred BALB C , Mice, Nude , Peritoneal Neoplasms/genetics , Peritoneal Neoplasms/metabolism , RNA, Messenger/biosynthesis , RNA, Messenger/genetics , Stomach Neoplasms/genetics , Stomach Neoplasms/metabolism , Vascular Endothelial Growth Factor A , Vascular Endothelial Growth FactorsABSTRACT
Retinoblastoma protein (pRb) functions as a tumor suppressor, and certain proteins are known to bind to pRb in the C-terminal region. Although the N-terminal region of pRb may also mediate interaction with some proteins, no such protein has been identified yet. We demonstrated previously the in vivo protein association between pRb and 73-kDa heat shock cognate protein (hsc73) in certain human tumor cell lines. In this report we analyzed the interaction between these two proteins in vitro. Our data showed that hsc73 interacts with the novel N-terminal region of pRb; that is, pRb binds directly to hsc73 and dissociates from hsc73 in an ATP-dependent manner. By using deletion mutants of cDNA encoding pRb, the hsc73 binding site of pRb was determined to be located in the region (residues 301-372) outside the so-called A pocket (residues 373-579) of this tumor suppressor protein. This finding was compatible with the fact that the adenovirus E1A oncoprotein, which is known to bind to the E2F binding pocket region of pRb, could not compete with hsc73 for the binding. Furthermore, phosphorylation of pRb by cyclin-dependent kinase inhibited the binding of pRb to hsc73. These data suggest that hsc73 may act exclusively as the molecular chaperone for nonphosphorylated pRb. As a result, hsc73 may function as a molecular stabilizer of nonphosphorylated pRb.
