An adenovirus vector encoding tyrosine hydroxylase activity may enter human cns cells in primary dissociated cultures

Abstract

An adenovirus encoding tyrosine hydroxylase (TH) activity was inserted in neuronal and glial cultured cells obtained from human fetal central nervous system (CNS) tissue. Using a double fluorescence immunostaining, we characterized inoculated CNS cells, with a TH antiserum and one of the following antibodies: microtubule-associated protein (MAP2) and GABA for neuronal cells, vimentin (Vim) for glial cells and glial fibrillary acidic protein (GFAP) for astrocytes. The characterization of inoculated neuronal cells was established by the detection of TH-MAP2-stained neurons in cultures obtained from the thoracic and lumbar parts of the spinal cord where no intrinsic TH cells are described. Inoculated glial cells were characterized by the detection of TH-Vim and TH-GFAP-stained CNS cultured cells. We also observed GABA neurons expressing TH immunoreactivity which could be considered as inoculated neurons expressing the GABA phenotype. Whatever the time of inoculation, transfection was observed in both neuronal and glial cells, after up to 4 months of culture. Although no precise quantitation was performed, the percentage of inoculation was found on microscopic inspection to be greater in glia than in neurons, as previously reported. We concluded that a gene coding for a key neuronal enzyme can be incorporated in embryonic human glial and neuronal cells through the use of a recombinant adenovirus.