title: A system, using neural cell lines, to characterize HSV-1 vectors containing genes which affect neuronal physiology, or neuronal promoters.
abstract: Among the potential uses of defective herpes simplex virus (HSV-1) vectors are to study neuronal physiology, neuronal gene regulation, and to perform gene therapy of neuronal diseases. The prototype HSV-1 vector, pHSVlac, stably expresses Escherichia coli beta-galactosidase from the HSV-1 immediate early (IE) 4/5 promoter in cultured rat peripheral and CNS neurons, and in neurons in the adult rat brain. The LacZ gene and the IE 4/5 promoter in pHSVlac can be replaced with genes which affect neuronal physiology or cellular promoters, respectively. A system is required to characterize these HSV-1 vectors; cultured neurons, a mixture of different kinds of neurons and glia, cannot be used. In contrast, neural cell lines represent a homogenous population of neural cells available in virtually unlimited quantities. A system, using neural cell lines, to characterize HSV-1 vectors carrying other genes or promoters is now reported: First, 4 assays are described to detect HSV-1 vector DNA, RNA transcribed from the vector, and to quantitate beta-galactosidase expression. Second, 8 cell lines derived from rodents, primates, and humans were infected with pHSVlac virus and shown to express beta-galactosidase. The cell lines tested included adrenergic and cholinergic mouse neuroblastoma cells, rat pheochromocytoma cells, rodent pituicytes, and human neuroblastoma cells. Infection of these cell lines should prove useful for characterizing HSV-1 vectors with molecular and biochemical assays. Third, differentiated rat pheochromocytoma and mouse neuroblastoma cells, which resemble neurons, were infected with pHSVlac virus and shown to stably express beta-galactosidase. Infection of these cells should be useful for determining the effect of various HSV-1 vectors on neuronal physiology. Thus, HSV-1 vectors containing various genes or promoters can be characterized using the system described in this study.
authors.affiliation: Department of Neurology, Massachusetts General Hospital, Boston 02114.
authors.forename: A I
authors.initials: AI
authors.lastname: Geller
mesh_terms: D000818:Animals; D015152:Blotting, Northern; D015139:Blotting, Southern; D002454:Cell Differentiation; D002460:Cell Line; D004279:DNA, Viral / Q000032:analysis; D004926:Escherichia coli / Q000201:enzymology; D005696:Galactosidases / Q000235:genetics / Q000276:immunology; D015967:Gene Expression Regulation, Viral / Q000502:physiology; D005814:Genes, Viral; D005822:Genetic Vectors / Q000502:physiology*; D000882:Haplorhini; D006801:Humans; D051379:Mice; D009474:Neurons / Q000502:physiology / Q000648:ultrastructure*; D011401:Promoter Regions, Genetic / Q000502:physiology*; D014176:Protein Biosynthesis; D012367:RNA, Viral / Q000032:analysis; D051381:Rats; D018139:Simplexvirus / Q000235:genetics*; D014407:Tumor Cells, Cultured
chemical_list: D004279:DNA, Viral; D012367:RNA, Viral; D005696:Galactosidases