Ex vivo and in vitro studies of transgene expression in rat astrocytes transduced with lentiviral vectors.

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Abstract

Implantation of cells genetically modified to express therapeutic genes into the brain has been proposed as a potential treatment for neurodegenerative diseases. In the current study embryonic rat-derived astrocytes were cultured and transduced with a lentiviral vector expressing the reporter gene green fluorescent protein (GFP) and subsequently grafted into the adult rat brain. The proportion of GFP expressing cells was stable, albeit small (1%), at all survival times, up to 6 weeks, the longest time point studied. In parallel in vitro studies, the astrocytes were lentivirally transduced to express either one of the two isoforms of glutamate decarboxylase (GAD(65) or GAD(67)) or glial cell line-derived neurotrophic factor (GDNF). When transducing 293T cells with the two GAD vectors, released GABA could be measured using high-performance liquid chromatography. Further studies of rat astrocytes transduced with the same vectors resulted in a level of GAD activity about 10 times higher than the activity of an intact rat striatum. One hundred thousand astrocytes transduced with LV-GDNF released approximately 27 ng of GDNF per hour. Thus, taken together, our observations provide support for the use of rat astrocytes in ex vivo gene transfer of these proteins in animal models of CNS disorders, e.g., Parkinson's disease or epilepsy.

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Research areas and keywords

Subject classification (UKÄ) – MANDATORY

  • Neurology

Keywords

  • Gene Expression : physiology, gamma-Aminobutyric Acid : metabolism, Transgenes : physiology, Transduction Genetic, Support Non-U.S. Gov't, Rats Sprague-Dawley, Rats, Luminescent Proteins : biosynthesis : genetics, Lentivirus : genetics, Kidney : metabolism, Immunohistochemistry, Human, Genes Reporter, Genetic Vectors : genetics : metabolism, Corpus Striatum : cytology : metabolism, Cells Cultured, Brain Tissue Transplantation, Astrocytes : cytology : metabolism : transplantation, Animal
Original languageEnglish
Pages (from-to)22-30
JournalExperimental Neurology
Volume173
Issue number1
Publication statusPublished - 2002
Publication categoryResearch
Peer-reviewedYes