Effects of a Brain-Engraftable Microglial Cell Line Expressing Anti-Prion scFv Antibodies on Survival Times of Mice Infected with Scrapie Prions

Abstract

We first verified that a single chain Fv fragment against prion protein (anti-PrP scFv) was secreted by HEK293T cells and prevented prion replication in infected cells. We then stably expressed anti-PrP scFv in brain-engraftable murine microglial cells and intracerebrally injected these cells into mice before or after infection with prions. Interestingly, the injection before or at an early time point after infection attenuated the infection marginally but significantly prolonged survival times of the mice. These suggest that the ex vivo gene transfer of anti-PrP scFvs using brain-engraftable cells could be a possible immunotherapeutic approach against prion diseases.

Prions disturb post-Golgi trafficking of membrane proteins

“…Uninfected N2aC24 line is a cloned N2a line overexpressesing exogenously transfected mouse PrP C (ref. 23). Infected N2aC24L1-3 line is a clone of N2aC24 cells that were infected with 22L prions 23 .…”

“…23). Infected N2aC24L1-3 line is a clone of N2aC24 cells that were infected with 22L prions 23 . PrP Sc was detectable in infected N2aC24L1-3 cells on western blotting (WB) (Fig.…”

“…N2aC24, N2aC24L1-3 and N2aC24Chm cells were established elsewhere 23 . N2a cells (ATCC CCL-131, kindly gifted from Prof Horiuchi, Hokkaido University) were permanently transfected with pEF1/Myc-His A (Invitrogen) encoding mouse PrP C .…”

Biological and Biochemical Characterization of Mice Expressing Prion Protein Devoid of the Octapeptide Repeat Region after Infection with Prions

“…Mouse neuroblastoma N2a cells persistently infected with 22L prions, designated N2aC24L1-3 [13], were transiently transfected with expression vectors using Lipofectamine 2000 reagent (Invitrogen). The cells were lysed in a buffer (150 mM NaCl, 0.5% Triton X-100, 0.5% sodium deoxycholate, 50 mM Tris-HCl, pH 7.5) 2 days after transfection and subjected to Western blotting.…”

Microglia in prion diseases

“…However, stimulation of microglia should be carefully controlled, since intracerebral or systemic administration of LPS to ME7-infected mice markedly stimulated microglial activation, but exacerbated the local inflammatory response and increased neuronal death (106). Intracerebral injection of brain-engraftable murine microglial cells that express an antiPrP single-chain variable fragment before or at an early stage after prion infection modestly but significantly prolonged the incubation times, suggesting a novel microglia-mediated immunotherapeutic approach against prion diseases (107).…”