The pancreas is a heterogeneous organ mixed with both exocrine and endocrine cells. The pancreas is involved in metabolic activities with the endocrine cells participating in the regulation of blood glucose while the exocrine portion provides a compatible environment for the pancreatic islets and is responsible for secretion of digestive enzymes. The purpose of this study was to identify pancreatic proteins that are differentially expressed in normal mice and those with diet-induced type 2 diabetes (T2DM). In this study, C57BL/6J male mice fed a highfat diet became obese and developed T2DM. The pancreatic protein profiles were compared between control and diabetic mice using two-dimensional gel electrophoresis (2-DE). Differentially expressed protein ‘spots’ were identified by mass spectrometry. REG1 and REG2 proteins, which may be involved in the proliferation of pancreatic beta-cells, were up-regulated very early in the progression of obese mice to T2DM. Glutathione peroxidase, which functions in the clearance of reactive oxidative species (ROS), was found to be down-regulated in the diabetic mice at later stages. The RNA levels of Reg2 and glutathione peroxidase were compared by northern blot analysis and were consistent to the changes in protein levels between diabetic and control mice. The up-regulation of Reg1 and Reg2 suggests the effort of the pancreas in trying to ameliorate the hyperglycemic condition by stimulating the proliferation of pancreatic beta-cells and enhancing the subsequent insulin secretion. The down-regulation of glutathione peroxidase in pancreas could contribute to the progressive deterioration of beta-cell function, due to the hyperglycemia induced oxidative stress.