title: L-Arginine ethylester enhances in vitro amplification of PrP(Sc) in macaques with atypical L-type bovine spongiform encephalopathy and enables presymptomatic detection of PrP(Sc) in the bodily fluids.
abstract: Protease-resistant, misfolded isoforms (PrP(Sc)) of a normal cellular prion protein (PrP(C)) in the bodily fluids, including blood, urine, and saliva, are expected to be useful diagnostic markers of prion diseases, and nonhuman primate models are suited for performing valid diagnostic tests for human Creutzfeldt-Jakob disease (CJD). We developed an effective amplification method for PrP(Sc) derived from macaques infected with the atypical L-type bovine spongiform encephalopathy (L-BSE) prion by using mouse brain homogenate as a substrate in the presence of polyanions and L-arginine ethylester. This method was highly sensitive and detected PrP(Sc) in infected brain homogenate diluted up to 10(10) by sequential amplification. This method in combination with PrP(Sc) precipitation by sodium phosphotungstic acid is capable of amplifying very small amounts of PrP(Sc) contained in the cerebrospinal fluid (CSF), saliva, urine, and plasma of macaques that have been intracerebrally inoculated with the L-BSE prion. Furthermore, PrP(Sc) was detectable in the saliva or urine samples as well as CSF samples obtained at the preclinical phases of the disease. Thus, our novel method may be useful for furthering the understanding of bodily fluid leakage of PrP(Sc) in nonhuman primate models.
authors.affiliation: Influenza Prion Disease Research Center, National Institute of Animal Health, Tsukuba, Ibaraki, Japan. Electronic address: ymura@affrc.go.jp.
authors.forename: Y
authors.initials: Y
authors.lastname: Murayama
authors.affiliation: Chiba Institute of Science Faculty of Risk and Crisis Management, Choshi, Chiba, Japan.
authors.forename: F
authors.initials: F
authors.lastname: Ono
authors.affiliation: Influenza Prion Disease Research Center, National Institute of Animal Health, Tsukuba, Ibaraki, Japan.
authors.forename: N
authors.initials: N
authors.lastname: Shimozaki
authors.affiliation: Tsukuba Primate Research Center, National Institutes of Biomedical Innovation, Health and Nutrition, Tsukuba, Ibaraki, Japan.
authors.forename: H
authors.initials: H
authors.lastname: Shibata
mesh_terms: D000818:Animals; D001120:Arginine / Q000008:administration & dosage / Q000031:analogs & derivatives*; D001826:Body Fluids / Q000378:metabolism*; D001921:Brain / Q000187:drug effects / Q000378:metabolism*; D002417:Cattle; D016643:Encephalopathy, Bovine Spongiform / Q000175:diagnosis* / Q000378:metabolism*; D008252:Macaca fascicularis; D008297:Male; D018620:PrPSc Proteins / Q000032:analysis*; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity
chemical_list: D018620:PrPSc Proteins; D001120:Arginine; C489258:arginine ethyl ester
keywords: Arginine ethylester; Atypical bovine spongiform encephalopathy; Bodily fluid; Nonhuman primate; Protein misfolding cyclic amplification