Establishment of a Line of Human Fetal Glial Cells That Supports JC Virus Multiplication

Primary cultures of human fetal brain cells were transfected with plasmid DNA pMK16, containing an origin-defective mutant of simian virus 40 (SV40). Several weeks after DNA treatment, proliferation of glial cells was evident in the culture, allowing passage of the cells at low split ratios. Initial...

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Bibliographic Details
Published in:	Proceedings of the National Academy of Sciences - PNAS Vol. 82; no. 4; pp. 1257 - 1261
Main Authors:	Major, Eugene O., Miller, Andra E., Mourrain, Pascale, Traub, Renee G., De Widt, Erik, Sever, John
Format:	Journal Article
Language:	English
Published:	Washington, DC National Academy of Sciences of the United States of America 01-02-1985 National Acad Sciences
Subjects:	Animal cells Antibodies Antigens, Polyomavirus Transforming Antigens, Viral, Tumor - biosynthesis Astrocytes - microbiology Biological and medical sciences Biotechnology Cell culture techniques Cell cultures. Hybridization. Fusion Cell Line Cell lines Cell Transformation, Viral Cultured cells Defective Viruses DNA DNA probes DNA Replication Establishment of new cell lines, improvement of cultural methods, mass cultures Eukaryotic cell cultures Fetus Fundamental and applied biological sciences. Psychology Humans JC virus JC Virus - growth & development Kidney cells Methods. Procedures. Technologies Molecular and cellular biology Neuroglia Neurons Polyomavirus - growth & development Simian virus 40 Viral Proteins - biosynthesis Virus Replication Human Cell culture Virus multiplication Parvovirus Neuroglia Polyomavirus Papovaviridae Long term Virus Astroglia Cell line Parvoviridae JC virus Production Fetus SV40 virus Cell
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Summary:	Primary cultures of human fetal brain cells were transfected with plasmid DNA pMK16, containing an origin-defective mutant of simian virus 40 (SV40). Several weeks after DNA treatment, proliferation of glial cells was evident in the culture, allowing passage of the cells at low split ratios. Initially, only 10% of the cells demonstrated nuclear fluorescence staining using a hamster tumor antibody to the SV40 T protein. By the sixth passage, however, 100% of the cells reacted positively to the same antibody. During these early passages, the cells designated SVG began growing very rapidly and acquired a homogenous morphology. Cell division required only low serum concentrations, was not contact-inhibited, and remained anchorage dependent. These characteristics of the SVG cells have been stable through 25 passages or ≈ 80 cell generations. The SV40 T protein is continuously produced in the cells and can direct the replication of DNA inserts in the pSV2 vector, determined by in situ hybridization using biotin-labeled DNA probes, which contains the SV40 replication origin. More importantly, SVG cells support the multiplication of the human papovavirus JCV at levels comparable to primary cultures of human fetal glial cells, producing infectious virus as early as 1 week after viral adsorption. Their brain-cell derivation has been established as astroglial, based on their reactivity with a monoclonal antibody to glial fibrillary acid protein and lack of activity with an anti-galactocerebroside antibody, which identifies oligodendroglial cells. The SVG cells represent a unique line of continuous rapidly growing human fetal astroglial cells that synthesizes a replication-proficient SV40 T protein. Their susceptibility to JC virus (JCV) infection obviates a host restriction barrier that limited JCV studies to primary cultures of human fetal brain and thus should allow for more detailed molecular studies of human brain cells and JCV that infects them.
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ISSN:	0027-8424 1091-6490
DOI:	10.1073/pnas.82.4.1257