Establishment of a line of human fetal glial cells that supports JC virus multiplication.

Abstract

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 biotinlabeled 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.

Human neurotropic polyomavirus, JCV, and its role in carcinogenesis

“…However, several strains of JCV have been successfully propagated in a number of primary cultures and established cell lines ( Table 1). As might be anticipated, the virus is able to grow in the astrocytic cell lines, SVG and POJ, which were generated via transformation of primary fetal glial cells with replication-defective SV40 and JC virus, respectively (Major et al, 1985;Mandl et al, 1987). Also, primary cultures of Schwann cells, the myelin-producing cells in the peripheral nervous system, are reported to be capable of supporting JCV replication (Assouline and Major, 1991).…”

Viral variant nucleotide sequences help expose leukocytic positioning in the JC virus pathway to the CNS

“…The persistently infectible SVG cells expressed phenotypes typical of astrocytes rather than oligodendrocytic precursor cells [44,48]. The heightened mitotic activity of these possible glial progenitors along with their less robust production of JCV, as compared with oligodendrocytes, may have been key factors in the successful production of SVG cells.…”

“…Transformed COS cells were the result of a mutant SV40 plasmid, 1-11, transfected into the CV-1 monkey kidney cell line [42]. This same technique was used to immortalize human embryonic kidney cells [43] and later, HFGC [44,45]. Mutant 1-11 was replicatively defective due to the deletion of its origin of replication and solely expressed elevated levels of T Fig.…”

“…The HFGC transformed by SV40 T protein, termed SVG cells [44], were susceptible to JCV infection and proved not only valuable for virus production, but also the study of antiviral drugs [46]. The development of other susceptible HFGC lines, termed POJ-19 and POJ-27 [47], employed replicatively defective JCV plasmids.…”

Interferon β1-a and selective anti-5HT2a receptor antagonists inhibit infection of human glial cells by JC virus

“…SVG-A cells are a subclone of the SVG human glial cell line established by transformation of human fetal glial cells with an origin-defective SV40 mutant (Major et al, 1985). Cells were cultured in EMEM supplemented with 10% fetal calf serum (Mediatech, Inc.) and maintained in a humidified 37°C CO 2 incubator.…”