In this study, we investigated the expression and function of thymosinα1 (Thyα1) in human pancreatic cancer. We found that human pancreatic cancer cell lines Panc-1, Panc03.27, ASPC-1, and PL45 cells significantly over-expressed the mRNA of Thyα1 as compared to the normal human pancreatic ductal epithelium (HPDE) cells.. Thyα1 mRNA and protein levels were also over-expressed in clinical pancreatic adenocarcinoma specimens. In addition, synthetic Thyα1 significantly promoted Panc-1 cell proliferation and increased phosphorylation of ERK1/2 and JNK. Furthermore, Thyα1 increased the secretion of multiple cytokines including IL-10, IL-13, and IL-17 in Panc-1 cells. Thus, Thyα1 may have a new role in pancreatic cancer pathogenesis.