Background and purpose: Pancreatic intraepithelial neoplasia (PanIN) may be a precursor lesion of infiltrating pancreatic ductal adenocarcinoma. The mutation of the phenotypic impact of K-ras G12D alone

silencing of p53 and p16 could promote this process. The role of Smad4 in this progression was poorly understood. In our previous studies

we investigated that RNA interference silence of Smad4 to promote the PanIN cell malignant transformation. In the present study

we investigate. The further explores the siRNA interference of Smad4 expression on PanIN cells could lead to proliferation and metastasis in vitro and in vivo. Methods: Smad4 knock-down PanIN cells (PanIN-S) were established by stable transfection with lentiviral-mediated Smad4 RNA interference. In vitro

silence of Smad4 enhanced the proliferation of PanIN cells as determined by cell counting. A soft agar assay was used to assess the anchorage-independent growth ability of cells. Cell migration and invasion assays were performed using transwell chambers with or without Matrigel. In xenograft model experiments

PCNA

VEGF and MMP-9 staining was separately used to evaluate cell proliferation and angiogenesis and migration (VEGF and MMP-9). Results: Effect of siRNA of Smad4 gene in PanIN cells was confirmed by real-time RT-PCR and western blot. In vitro

silence of Smad4 enhanced the proliferation of PanIN cells as determined by cell counting. Soft agar assay showed that there were more colony cell numbers in PanIN-S cells compared with PanIN cells (P0.05). Using the transwell assay

we observed that PanIN-S cells migrated faster than PanIN cells and similar results were obtained by Matrigel assay (P0.05). Furthermore

immunohistochemical analysis of the harvested tumors suggested that Smad4 silencing was associated with cell proliferation (PCNA reactivity) and angiogenesis and migration (VEGF and MMP-9)

and the expressions of PCNA

VEGF and MMP-9 in PanIN-S group were significantly increased (P0.05). Conclusion: Silence of Smad4 in PanIN cells enhanced progression to invasive adenocarcinoma of the pancreas by promoting cell growth

migration and invasion. Smad4 might be a new diagnostic marker in pancreatic cancer and prove to be a feasible and novel target for therapeutic intervention.