Yukio Kuwada

Expressions of Angiogenic Factors in Pancreatic Ductal Carcinoma : A Correlative Study with Clinicopathologic Parameters and Patient Survival

Introduction It has been reported that angiogenic factors play an important role in proliferation and metastasis in various cancers. Aim To investigate the expression of angiogenic factors and microvessel density (MVD) in pancreatic ductal carcinoma and to examine the correlations among expression of angiogenic factors, clinicopathologic parameters, and clinical prognosis. Methodology Paraffin-embedded specimens from 55 patients with pancreatic ductal carcinoma were immunostained for vascular endothelial growth factor (VEGF), fibroblast growth factor-2 (FGF-2), platelet-derived endothelial cell growth factor (PD-ECGF), and CD34. The correlations among the expression of individual angiogenic factors and MVD, the clinicopathologic features, and the clinical prognoses were analyzed statistically. Results Immunostaining demonstrated that 70.8% of pancreatic ductal carcinomas were positive for VEGF, 60.9% for FGF-2, and 57.2% for PD-ECGF. A significant correlation was observed between VEGF expression and MVD (p < 0.05) but not between FGF-2 or PD-ECGF and MVD. Although the expression of each angiogenic factor had no correlation with clinicopathologic features, the patients with tumors that showed high expression of VEGF and FGF-2 had significantly shorter survival times than those with low or no such expression (p < 0.05). Conclusions These observations suggest that the expression of VEGF closely correlates with MVD and with a poor prognosis in pancreatic ductal carcinoma.

Irsogladine Malate Up-regulates Gap Junctional Intercellular Communication Between Pancreatic Cancer Cells Via PKA Pathway

Introduction Gap junctions (GJs) are intercellular channels that aid communication between coupling cells and may play a critical role in cell differentiation and growth. Connexins (Cxs) are structural proteins of GJs. Though several reports have demonstrated that Cx expression decreases in various malignant tumors, a pancreatic cancer cell line, PANC-1, was reported to express Cx43 mRNA. It is known that irsogladine malate (IM) can up-regulate gap junctional intercellular communication (GJIC). We examined the effects of IM on GJ between pancreatic cancer cells (PC cells) and the mechanism of GJ up-regulation. Methodology GJIC between PC cells (PANC-1) was evaluated by dye transfer methods. The expression of Cx43 was estimated by Western blot analysis with immunoprecipitation sample and immunohistochemical analysis. Intracellular cAMP level was estimated by enzyme-linked immunoassay. Results IM increased cell coupling in a dose-dependent manner (0M-10−4M). Western blot analysis of Cx43 revealed that PANC-1 cells expressed Cx43 protein. Treatment with IM was found to move localization of Cx43 immunoreactive spots from the cytoplasm to boundary lesions with neighboring cells, but no major change was seen in the phosphorylation state of Cx43. Intracellular cAMP level was increased by IM. The PKA inhibitor H-89 and adenylyl cyclase inhibitor SQ22536 inhibited the effects of IM. Conclusion These results suggest that IM up-regulates GJIC between PC cells via regulation of the PKA pathway. It also suggests a useful adjuvant of IM to pancreatic cancer therapy.

Activation of Peroxisome Proliferator-Activated Receptor Gamma Inhibits the Growth of Human Pancreatic Cancer

Objective: In the present study, we examined the expression of peroxisome proliferator-activated receptor gamma (PPARγ) in human pancreatic cancer and the possible effects of its ligand engagement on cell growth. Methods: Seven human pancreatic cancer cell lines and 7 surgically resected human pancreatic cancer tissues were used as samples. The expression of PPARγ was analyzed with reverse transcription-polymerase chain reaction and immunoblotting. The interaction between PPARγ and PPAR-responsive element (PPRE) was examined by gel shift assay. Growth inhibition by thiazolidinediones was confirmed with anchorage-dependent and anchorage-independent growth assays. Results: PPARγ was detected in all cell lines tested and in 5 out of 7 cancer tissues (71%), but was not found in adjacent normal pancreatic tissues. Gel shift analysis revealed that the proteins in nuclear extracts of the pancreatic cancer cell line PANC-1 specifically bind to the PPRE. Cell growth was significantly inhibited by treatment with troglitazone and rosiglitazone in a dose- and time-dependent manner (p < 0.01). In contrast, a nonfunctional metabolic analog of troglitazone did not affect cell growth. Conclusion: These observations suggest that PPARγ plays an important role in human pancreatic cancer growth and that ligand-induced activation of PPARγ would be a useful strategy for treatment of human pancreatic cancer.

Expression of interleukin-8 correlates with invasiveness in human pancreatic cancer

Background Interleukin S (IL-S), a member of the CXC chemokine family, is a multifunctional cytokine shown to induce angiogenesis, haptotactic migration, and proliferation of certain cancer cells. IL-S is presumed to exert its action after binding to its specific receptors (IL-SRA and IL-SRB). Previously, we have reported that matrix metalloproteinase-9 (MMP-9) was increased by treatment with IL-S in gastric cancer cells. The aims of this study was to examine whether IL-S and IL-S receptors were expressed in human pancreatic cancer and to evaluate the role of IL-S on metastatic potential in pancreatic cancer. Methods The expression of IL-S, IL-SRA and IL-SRB in human pancreatic cancer cells (PANC-I, MIA PaCa-2, Capan-2) were examined by northern blot analysis and western blot analysis. The expressions of IL-S and its receptors were also investigated by immunohistochemistry in human pancreatic cancer tissues using specific antibody. Cytokine induced secretions of IL-S in supernatants of cultured medium from cells were investigated with ELISA (R&D). After treatment with various concentrations of recombinant IL-S (Sigma), the cell proliferations were assayed using Cell Counting KitTM (Dojindo Labs., Japan). IL-S induced production and gelatinolytic activity of MMP-2 and MMP-9 were analyzed by western blot analysis and gelatin zymogram using conditioned mediums from each cell as samples. Results The expression of IL-S, IL-SRA and IL-SRB were detected in all investigated pancreatic cancer cells. Moreover, the secretion of IL-S was increased with cytokine stimuli. In addition, immunoreactive signals of IL-8 and its receptors were observed in cancer tissues.· Exogenous adding of IL-S did not induce proliferation of human pancreatic cancer cell lines. Surprisingly, the production and gelatinolytic activity ofMMP-2 and MMP-9 were increased by the treatment of recombinant IL-S in a dose-dependent manner. Conclusion These results suggest that IL-S regulate MMP activity and may play an important role in the invasiveness of human pancreatic cancer.