Retrovirus-mediated gene transfer and galactocerebrosidase uptake into twitcher glial cells results in appropriate localization and phenotype correction

Galactocerebrosidase (GALC) is deficient in all tissues from human patients and animal models with globoid cell leukodystrophy (GLD) or Krabbe disease . The deficiency results in decreased lysosomal catabolism of certain galac tolipids including galactosylceramide and psychosine that are synthesized m aximally during myelination. According to current theories, the accumulatio n of psychosine in humans and animals with GLD induces oligodendrocyte dege neration and myelination ceases. Transduction of oligodendrocytes from twit cher mice with a retroviral vector containing the GALC cDNA can correct the enzyme deficiency in these cells. Our data show that twitcher astrocytes a nd oligodendrocytes can internalize exogenous GALC, as well as donate the e nzyme to the mutant glial cells. Antibodies against human GALC localized th e GALC antigen in retrovirally transduced cells and cells receiving enzyme via cell to cell secretion and uptake to the lysosomal fraction. In fact im munocytochemical studies in transduced oligodendrocytes revealed that the G ALC colocalizes in vesicles lysosomal-associated membrane protein-2 (LAMP2) (+). Moreover, labeling cells with anti-GALC and a marker for oligodendroc ytes demonstrated that, upon differentiation, transduced, twitcher oligoden drocytes attained the normal branched process configuration, while untransd uced calls show only abnormal morphology. Phenotype correction in mutant ol igodendrocytes has also been observed after enzyme transfer. These studies indicate that GALC activity supplied to cultured oligodendrocytes from twit cher mice by different methods can correct the pathological phenotype of th ese cells. (C) 2001 Academic Press.