Scrapie in vitro: agent replication and reduced cell yield

Abstract

Exposure of PAM cells (see below) to brain homogenates from mice infected with scrapie caused a relative decrease in total cell yield, which persisted from passage 2 or 3 to passage 18 after treatment. The effect was elicited by each of the eight independent scrapie isolates tested. Lysates prepared from cultures 16 passages after treatment with scrapie caused the decrease when applied to fresh PAM cultures. Mice inoculated with passage 14 and 18 lysates developed a reduced percentage of polymorphonuclear neutrophils by 5 wk and scrapie disease by 6 to 9 months after inoculation. Based on the total dilution from treatment of the PAM cultures with scrapie material to the preparation of the lysates, it is concluded that the agent(s) responsible for the reduced PAM cell yield, the decreased percentage of polymorphonuclear neutrophils, and the induction of scrapie disease had replicated in the PAM cells. By filtration, the diameter of the agent causing the reduction in cell yield was estimated to be between 25 and 50 nm. PAM cells originated several yr ago as a spontaneously transformed, continuous cell line derived from mouse embryo fibroblasts. The cells were subcultured as monolayers at approximately weekly intervals at a split ratio of 1:10. The line was used at passages 175 to 210 for these experiments. This line contains the mouse leukemia virus, as determined by the presence of virus group specific complement fixation antigen, the appearance of [3H]uridine labeled virus in culture medium, and the presence of reverse transcriptase in the culture medium.