BACKGROUND.: Accumulated evidence has shown that insulin producing β-cells in pancreatic islets have the limited potential to regenerate. Adenoviruses have been widely employed to deliver genes of interest into pancreatic islets. This study was aimed at investigating whether adenovirus infection has any impact on the potential of β-cell proliferation. METHODS.: Human adenovirus type 5 (Ad5) encoding rat insulin promoter driven reporter genes were used to infect freshly isolated pancreatic islets. Western blotting assays were performed to evaluate the expression and activation of key molecules involved in cell survival and proliferation following Ad5 infection. Immunofluorescence staining was employed to identify proliferating cells after culturing the infected and control islets in the presence of BrdU, an analog of thymidine that can be incorporated into the genome of proliferating cells. RESULTS.: Ad5 infection of the islets resulted in expression and activation of Akt1, a key molecule in the PI3 kinase signaling pathway. Accordingly, a higher frequency of islet cell proliferation was detected in Ad5-infected islets than in control islets. DISCUSSION.: These data suggest adenovirus infection can activate β-cell survival and proliferation machinery, in particular operating through the PI3K/Akt signaling pathway. This information has significant ramification for the use of adenovirus as a gene delivery vehicle for pancreatic islet cells. © 2009 by Lippincott Williams & Wilkins.