ABSTRACTAn ideal cell type for ex vivogene therapy should be easy to biopsy, propagate, and genetically engineer in culture, should be transplantable using simple procedures, and should express therapeutic proteins at useful levels. The mesothelial cell appears to satisfy these criteria. Several thousand proliferative mesothelial cells were present in typical specimens of nonpathologic human peritoneal fluid obtained by needle aspiration. These divided rapidly in a specialized medium to yield pure cultures of ~107cells within 2 weeks. The replicative lifespan of mesothelial cells cultured from adults was ~4252 population doublings, permitting expansion and cryopreservation of a lifetime supply of autologous cells from one fluid sample. Cells transduced with a human growth hormone (hGH) adenoviral vector secreted 100300 g of hGH/106cells per day for at least 6 weeks in culture when maintained at quiescence. Intraperitoneal injection of transduced cells into athymic mice resulted in rapid systemic delivery of hGH, with peak plasma levels of 0.11 g/ml declining over 3 weeks to <1 ng/ml. Mice receiving a second injection of engineered cells displayed the same plasma hGH levels and duration as naive mice. Cells labeled with a -galactosidase vector were identifiable by in situenzymatic staining as clusters attached to peritoneal surfaces at multiple sites for at least 19 days after injection. Cells serially passaged through about three-quarters of their lifespan before transduction and injection were as effective at hGH delivery as earlier-passage cells. These results indicate the clinical potential for ex vivogene therapy using mesothelial cells.