Pancreatic ductal adenocarcinoma (PDAC) is one of the most malignant primary tumors in humans, with extremely high lethality. Although great efforts have been made to understand the molecular regulation of the tumorigenesis of PDAC, our current knowledge remains very limited. Previous work has shown a possible involvement of miR-21 in the growth of PDAC, whereas the underlying mechanism has not been clarified. Here, we show significant higher levels of miR-21 in PDAC, compared to the adjacent normal pancreatic tissue. Moreover, overexpression of miR-21 in PDAC cells increased cell growth, whereas inhibition of miR-21 decreased cell growth. Furthermore, miR-21 was found to inhibit nuclear retention of FoxO1 to augment the growth of PDAC cells. Thus, miR-21/FoxO1 axis appears to be a novel therapeutic target for inhibiting the growth of PDAC.