Human proinsulin production in primary rat hepatocytes after retroviral vector gene transfer

The development of autologous somatic cells, engineered for the synthesis a nd release of human insulin under physiological stimuli, would certainly re present a major breakthrough in the therapy of insulin-dependent diabetes m ellitus. We generated a retroviral vector containing the human proinsulin c DNA and the gene coding for the human nerve growth factor receptor for quan titative analysis of transduced cells. Primary rat hepatocytes were selecte d as target cells because of the constitutive expression of the pancreatic p-cen glucose transporter GLUT-2 and the glycolitic enzyme glucokinase. App ropriate conditions for culture and retroviral transduction are described. The highest transduction efficiency,evaluated as percentage of LNGFr expres sing cells was obtained by repeated infection cycles (40+/-10%). Human proi nsulin accumulated in the culture medium of transduced rat hepatocytes (mea n+/-SD): 18.1+/-7.9 (range 8.7-36.4) ng/24h/10(6) cells. Primary rat hepato cytes can be efficiently transduced by a retroviral vector and the de novo synthesis of human proinsulin can be induced. Primary cultured hepatocytes represent an useful model to test retroviral constructs engineered for the glucose-inducible expression of insulin under the control of liver-specific promoters.