Hedgehog signaling pathway regulates human pancreatic cancer cell proliferation and metastasis.

Pancreatic cancer is one of the most aggressive and devastating malignancies. The Hedgehog (Hh) pathway has been reported to play an important role in pancreatic cancer development and progression. The aim of this study was to examine the activation of the Hh pathway in human pancreatic cancer tissue samples and pancreatic cancer cell lines, and the molecular mechanisms involved in the Hh pathway mediated effects on pancreatic cancer cell proliferation and invasion. The expression levels of Hh molecules in human pancreatic cancer tissue samples and pancreatic cancer cell lines were evaluated using RT-PCR. The role of the Hh pathway in cell proliferation and invasion was evaluated using flow cytometry, MTT, colony formation assays and transwell invasion assays, and the expression of cancer stem cell markers and epithelial-mesenchymal transition (EMT) were evaluated using flow cytometry and RT-PCR. Tumorigenicity assays were used to further investigate the role of the Hh pathway in vivo. Hh molecules were highly expressed in human pancreatic cancer tissue samples and pancreatic cancer cell lines. Inhibition of the Hh pathway notably decreased cell proliferation and induced apoptosis through inhibition of the PI3K/AKT pathway and cancer stem cells. Furthermore, inhibition of the Hh signaling pathway significantly inhibited EMT by suppressing the activation of transcription factors Snail and Slug, which are correlated with significantly reduced pancreatic cancer cell invasion, suggesting that the Hh signaling pathway is involved in early metastasis. These results indicate that activation of the Hh pathway is a common event. Inhibition of the Hh pathway may be a potential molecular target of new therapeutic strategies for pancreatic cancer.

Combined effects of EGFR and Hedgehog signaling pathway inhibition on the proliferation and apoptosis of pancreatic cancer cells.

In the present study, we established a new experimental model to investigate the effects of EGFR targeting by RNAi, and the synergistic actions between the hedgehog (Hh) and EGFR signaling pathways on the proliferation and apoptosis in pancreatic cancer cells. Three human pancreatic cancer cell lines expressing EGFR shRNA were established, and gene expression inhibition was assessed in these lines using RT-PCR and western blot analysis. The effects of EGFR RNAi and Hh inhibition on cell proliferation and apoptosis were explored in vitro and in vivo. We observed that EGFR RNAi notably inhibited cell proliferation and colony formation, induced apoptosis and markedly decreased xenograft tumor growth. Furthermore, EGFR RNAi significantly enhanced cyclopamine sensitivity both in vitro and in vivo, and a synergistic decrease of both AKT and ERK phosphorylation was observed. The present study demonstrates that combined inhibition of both EGFR and Hh signaling pathways could establish a more promising antitumor approach than inhibiting each singly, and that there is a possible synergistic effect for Hh and EGFR signaling pathways on ERK and AKT phosphorylation.

Suppression of the epidermal growth factor receptor inhibits epithelial-mesenchymal transition in human pancreatic cancer PANC-1 cells.

BACKGROUND: Aberrant expression of epidermal growth factor receptor (EGFR) has been detected in pancreatic cancer; however, the mechanisms of EGFR in inducing pancreatic cancer development have not been adequately elucidated. The objective of this study was to determine the role of EGFR in mediating epithelial-mesenchymal transition (EMT) in pancreatic cancer cells. METHODS: Pancreatic cancer cell line PANC-1 was transfected with small interfering RNA of EGFR by use of a lentiviral expression vector to establish an EGFR-knockdown cell line (si-PANC-1). PANC-1 cells transfected with lentiviral vector expressing negative control sequence were used as negative control (NC-PANC-1). Scratch assay and transwell study were used to analyze cell migration and invasion. Real-time PCR and Western blotting were used to detect the expression of EMT markers E-cadherin, N-cadherin, vimentin, and fibronectin and transcription factors snail, slug, twist1, and sip1 in PANC-1, NC-PANC-1, and si-PANC-1 cells. Immunofluorescent staining with these antibodies and confocal microscopy were used to observe their cellular location and morphologic changes. RESULTS: After RNA interference of EGFR, the migration and invasion ability of si-PANC-1 cells decreased significantly. The expression of epithelial phenotype marker E-cadherin increased and the expression of mesenchymal phenotype markers N-cadherin, vimentin, and fibronectin decreased, indicating reversion of EMT. We also observed intracellular translocation of E-cadherin. Expression of transcription factors snail and slug in si-PANC-1 cells decreased significantly. CONCLUSION: Suppression of EGFR expression can significantly inhibit EMT of pancreatic cancer PANC-1 cells. The mechanism may be related with the down-regulation of the expression of transcription factors snail and slug.

[Surgical complications after pancreatoduodenectomy: risk factors and treatments].

OBJECTIVE: To explore the impact factors and treatment of post pancreatoduodenectomy complications. METHODS: The clinical data of 412 cases between January 1995 and April 2010 underwent pancreatoduodenectomy were analyzed retrospectively. There were 232 male, 180 female. Univariate and multivariate logistic regression model were used to identify the risk factors related to occurrence of postoperative complications. RESULTS: The overall postoperative morbidity rate was 37.1% (153/412), and mortality rate was 4.6% (19/412). Total uncinate process resection, type of pancreatic-enteric anastomosis, duct diameter and pancreatic texture had effects on postoperative pancreatic fistula statistically. Total uncinate process resection, the amount of intra-operative blood loss ≥ 600 ml and pancreatic fistula were identified as significant risk factors for post pancreatoduodenectomy hemorrhage by means of univariate analysis. Delayed gastric empting occurrence in the patients with pylorus-preserving pancreaticoduodenectomy was higher than those with standard pancreaticoduodenectomy significantly. The multivariate Logistic regression analysis revealed that duct diameter and pancreatic texture were the independent risk factors of pancreatic fistula. Total uncinate process resection, the amount of intra-operative blood loss ≥ 600 ml and pancreatic fistula were independent risk factors of bleeding. There were no statistically significant differences between the radical group and the standard group when postoperative complication rates were analyzed (P < 0.05). CONCLUSIONS: Pancreaticojejunal anastomoses by means of duct-to-mucosa is fit for the patients with dilated pancreatic duct and end-to-end invaginated pancreaticojejunostomy is fit for the patients with undilated pancreatic duct. The prevention of postoperative bleeding depends on total uncinate process resection and meticulous hemostatic technique during operation. The pancreatic fistula is one of the most important factors which can result in postoperative bleeding. Pancreaticoduodenectomy combines with SMV/PV resection and extended lymphadenectomy do not significantly increase the morbidity rates.