Abstract
Most of the congenital errors of liver metabolism affect one single pathway and spare the general function and structure of the liver. Gene transfer into hepatocytes might be an alternative treatment of many inborn errors of the liver metabolism. We study gene therapy of the congenital unconjugated hyperbilirubinemia (Crigler-Najjar type 1) in a rat model (Gunn-rat). The gene for UDPGT has been cloned. We have constructed an expression vector containing the B-UGT-1 cDNA. We are busy to characterize the expression of the B-UDPGT cDNA by western-blotting and in a functional assay for bilirubin glucuronidation in microsomes prior to the use for gene transfer into hepatocytes in vitro and in vivo.
Preliminary experiments have been performed for ex vivo gene transfer and hepatocyte transplantation. After isolation the hepatocytes were transfected with the pCMV-LacZ construct using DOTAP in culture and injected into the spleen of rats. The marker gene was expressed by 10 % of the cultured cells. Five days after injection blue hepatocytes were observed in the spleen (counter staining with PAS}. To improve the viability of hepatocyte in vitro we started to culture the cells in suspension by using collagen microspheres. The hepatocytes stay viable for > 10 days in culture. To optimize the gene transfer into the hepatocytes it will be necessary to test the different avaible transfection systems and to develop new methods for vector delivery.
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Wilke, M., Bijma, A., Timmers-Reker, A. et al. 254 GENE THERAPY FOR CONGENITAL HYPERBILIRUBINEMIA. Pediatr Res 36, 45 (1994). https://doi.org/10.1203/00006450-199407000-00254
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DOI: https://doi.org/10.1203/00006450-199407000-00254