GENE-THERAPY OF DIABETES - GLUCOSE-STIMULATED INSULIN-SECRETION IN A HUMAN HEPATOMA-CELL LINE (HEP G2INSLG)

In order to design a feasible somatic cell gene delivery system for th e treatment of type I diabetes, a suitable cell type needs to be deter mined. We have previously shown that the stable transfection of the fu ll-length insulin cDNA into the human liver cell line, (HEP G2ins) res ulted in synthesis, storage and acute regulated release of insulin to analogues of cAMP, but not to the physiological stimulus glucose. In a ttempting to explain the lack of glucose responsiveness of the HEP G2i ns cells we have stably transfected these cells with the human islet g lucose transporter GLUT 2 (HEP G2inslg cells). The HEP G2inslg cell cl ones exhibit glucose-stimulated insulin secretion and glucose potentia tion of the secretory response to nonglucose secretagogues. While gluc ose responsiveness commenced at a lower concentration than normal isle ts, a secretion curve approaching normal physiological conditions was generated. Immunoelectron microscopy revealed the presence of insulin- containing granules, similar in size and appearance to those of the no rmal beta cell. These results demonstrate that while it is most likely that the HEP G2inslg cell line predominately secretes insulin via the constitutive pathway, significant acute regulated release was seen in response to glucose, and thus represents significant progress in the creation of a genetically-engineered 'artificial beta cell' from a hum an hepatocyte cell line.