A versatile prion replication assay in organotypic brain slices.
Methods enabling prion replication ex vivo are important for advancing prion studies. However, few such technologies exist, and many prion strains are not amenable to them. Here we describe a prion organotypic slice culture assay (POSCA) that allows prion amplification and titration ex vivo under conditions that closely resemble intracerebral infection. Thirty-five days after contact with prions, mouse cerebellar slices had amplified the abnormal isoform of prion protein, PrP(Sc), >10(5)-fold. This is quantitatively similar to amplification in vivo, but fivefold faster. PrP(Sc) accumulated predominantly in the molecular layer, as in infected mice. The POSCA detected replication of prion strains from disparate sources, including bovines and ovines, with variable detection efficiency. Pharmacogenetic ablation of microglia from POSCA slices led to a 15-fold increase in prion titers and PrP(Sc) concentrations over those in microglia-containing slices, as well as an increase in susceptibility to infection. This suggests that the extensive microglial activation accompanying prion diseases represents an efficacious defensive reaction.
Falsig
Jeppe
J
Institute of Neuropathology, University of Zürich, Schmelzbergstrasse 12, Zürich, Switzerland.
Julius
Christian
C
Margalith
Ilan
I
Schwarz
Petra
P
Heppner
Frank L
FL
Aguzzi
Adriano
A
D000704:Analysis of Variance; D000818:Animals; D000831:Animals, Newborn; D017209:Apoptosis / Q000235:genetics; D039481:CD11b Antigen / Q000235:genetics; D053148:Caspase 3; D016923:Cell Death / Q000235:genetics; D002478:Cells, Cultured; D002531:Cerebellum / Q000166:cytology / Q000378:metabolism*; D005786:Gene Expression Regulation; D049452:Green Fluorescent Proteins / Q000378:metabolism; D051379:Mice; D008810:Mice, Inbred C57BL; D008822:Mice, Transgenic; D017628:Microglia / Q000378:metabolism; D009447:Neuroblastoma / Q000473:pathology; D009924:Organ Culture Techniques; D018620:PrPSc Proteins / Q000378:metabolism*; D017096:Prion Diseases / Q000209:etiology* / Q000635:transmission; D000072002:Prion Proteins; D011328:Prions / Q000235:genetics / Q000378:metabolism*; D011419:Propidium; D013997:Time Factors
D039481:CD11b Antigen; D018620:PrPSc Proteins; D000072002:Prion Proteins; D011328:Prions; C512657:Prnp protein, mouse; D049452:Green Fluorescent Proteins; D011419:Propidium; D053148:Caspase 3