Aim: To create HEK293 cells and murine ES cell lines that synthesize and release either GIP or the specific GIPR antagonist GIP(5-42). Methods: Using PCR, chimeric genes, consisting of the mouse growth hormone leader (mGH), the furin cleavage site, and either GIP or GIP(5-42), were generated and cloned into a lentiviral vector downstream of a constitutive promoter. After the generation of lentiviral pseudoparticles containing the chimeric genes, mGH-GIP and mGH-GIP(5-42), HEK 293 and ES cells were transduced with ~108 pseudoparticles. Pools of cells containing the co-expressed fluorescent marker ZsGreen were isolated using flow cytometry before supernates from cultured cells were collected, and a specific ELISA was employed to quantify GIP immunoreactivity. To assess GIP bioactivity in supernates, a specific bioassay consisting of reporter cells expressing GIPR and containing the LacZ gene under the control of a cAMP-responsive promoter was used. To demonstrate antagonist activity, peptides in supernates from HEK 293 cells containing mGH-GIP(5-42) were concentrated 50-fold using a C-18 reverse phase column before being mixed with 1010 M GIP and added to reporter cells. Results: HEK 293 cells containing mGH-GIP and mGH-GIP(5-42) expressed 8x10-4 and 10-4 pg/cell/h, respectively, while ES cells containing mGH-GIP and mGH-GIP(5-42) expressed 10-4 and 6x10-3 pg/cell/h, respectively. Supernates from HEK 293 and ES cells containing mGH-GIP were both shown to induce LacZ expression, demonstrating the presence of bioactive GIP. As expected, supernates from either HEK 293 cells or ES cells containing mGH-GIP(5-42) did not induce LacZ expression. However, after concentration, supernates from HEK 293 cells containing mGH-GIP(5-42) inhibited LacZ expression induced by 10-10 M GIP, indicating the presence of antagonist activity. Summary and Conclusion: The results of these studies demonstrate the successful engineering of HEK 293 and ES cells to express bioactive GIP and GIP(5-42). Although early in its development, engraftment of tissue derived fromGIP(5-42) expressing stem cells may provide a method for treating obesity.