SUMMARY:Recent evidence indicates that loss of centrosome integrity may be a major cause of genetic instability underlying various human cancers. The aim of this study was to define the role of centrosome defects during the in vivo tumor progression of pancreatic carcinoma using an orthotopic implantation model. Injection of Suit-2 human pancreatic cancer cells into the pancreata of nude mice reproduced the pattern of local tumor growth and distant metastasis observed in humans. Pancreatic xenografts, peritoneal disseminations, and hepatic metastases were harvested, and tumor cells were examined for centrosomes by immunofluorescence microscopy. Centrosome abnormalities, characterized by increased numbers of centrosomes, were detected in only a small fraction of parental Suit-2 cells in culture, whereas the frequency was markedly increased in cells isolated from the pancreatic xenografts. Abnormal centrosome numbers were found at higher frequencies in metastatic foci than in pancreatic xenografts. A significant positive correlation existed between the fraction of cells with multiple centrosomes and that with multipolar mitotic spindles, suggesting a functional involvement of aberrant centrosomes in spindle disorganization and chromosome missegregation. In addition, the increased frequency of abnormal centrosomes was associated with an enhanced degree of chromosomal instability. These findings suggest a novel model of pancreatic tumor progression whereby a stepwise increase in the magnitude of centrosomal abnormalities confers an increased chance for aberrant mitotic events, thus accelerating genetic instability and causing the tumor to progress to a more advanced stage. (Lab Invest 2001, 81:945-952).
Epidermal growth factor (EGF) stimulated proliferation of gastric mucous epithelial cells from guinea pigs in serum-free culture conditions. Western blot analysis with antiphosphotyrosine antibody showed that EGF initiated tyrosine phosphorylation of a 170-kDa protein, and this protein was identical to the EGF receptor. Insulin was not mitogenic, but it potentiated the mitogenic effect of EGF. Tyrosine phosphorylation of additional proteins was not induced by the combined actions of insulin and EGF. Stimulation by EGF and/or insulin did not cause a calcium response. However, when insulin was added to cells pretreated with EGF for > 6 h, it elicited a rapid intracellular calcium concentration rise that was reproducible in both cell suspension and single cell analyses. This calcium response coincided with the translocation of protein kinase C (PKC) from the cytosolic to the particulate fraction. Phorbol 12-myristate 13-acetate also caused the translocation and stimulated proliferation of the cells. These results suggest that the calcium-dependent activation of PKC may participate in the potentiation of the mitogenic effect of EGF by insulin.
These results suggest that the increased level of chromosomal instability may play a critical role in the development of aggressive tumor phenotype during pancreatic cancer progression. J. Surg. Oncol. 2001;76:181-187.
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