NR AJWV
AU Raymond,G.J.; Hope,J.; Kocisko,D.A.; Priola,S.A.; Raymond,L.D.; Bossers,A.; Ironside,J.; Will,R.G.; Chen,S.G.; Petersen,R.B.; Gambetti,P.; Rubenstein,R.; Smits,M.A.; Lansbury,P.T.Jr.; Caughey,B.W.
TI Molecular assessment of the potential transmissibilities of BSE and scrapie to humans
QU Nature 1997 Jul 17; 388(6639): 285-8
KI Nature. 1997 Jul 17;388(6639):228-9. PMID: 9230425
PT journal article
AB More than a million cattle infected with bovine spongiform encephalopathy (BSE) may have entered the human food chain. Fears that BSE might transmit to man were raised when atypical cases of Creutzfeldt-Jakob disease (CJD), a human transmissible spongiform encephalopathy (TSE), emerged in the UK. In BSE and other TSE diseases, the conversion of the protease-sensitive host prion protein (PrP-sen) to a protease-resistant isoform (PrPres) is an important event in pathogenesis. Biological aspects of TSE diseases are reflected in the specificities of in vitro PrP conversion reactions. Here we show that there is a correlation between in vitro conversion efficiencies and known transmissibilities of BSE, sheep scrapie and CJD. On this basis, we used an in vitro system to gauge the potential transmissibility of scrapie and BSE to humans. We found limited conversion of human PrP-sen to PrPres driven by PrPres associated with both scrapie (PrP[Sc]) and BSE (PrP[BSE]). The efficiencies of these heterologous conversion reactions were similar but much lower than those of relevant homologous conversions. Thus the inherent ability of these infectious agents of BSE and scrapie to affect humans following equivalent exposure may be finite but similarly low.
VT
(Erste Seite) Experimental transmission of BSE to mice[13], cattle[14] and two sheep PrP genotypes (A136 Q171 (ov-AQ) and V136 Q171 (ov-VQ))[15] has been reported, but hamsters (J. D. Foster and J-H., unpublished data) and at least one PrP genotype of sheep (A136 R171 (ov-AR))[16] seen to be resistant to clinical disease. To test for a correlation between in vitro cell-free conversion and in vivo transmission, we included types of 35S-labelled PrP-sen from each susceptible or resistant group of species in PrPBSE-driven conversion experiments. ProteinaseK resistant 35S-labelled conversion products that were the characteristic 6K to 7K smaller than the 35S-PrP-sen substrate[8-12,17] were generated in the reactions with both glycosylated and aglycosyl 35S-PrP-sen molecules from BSE-susceptible, but not BSE-resistant, hosts (Fig. 1). For example, the 25K aglycosyl bovine and sheep (ov-AQ and ov-VQ) 35S-PrP-sen proteins generated 18K 35S-PrPres bands (Fig. 1, lanes 1,6,7,13,15,16), whereas the aglycosyl hamster and ov-AR gave no proteinaseK-resistant product (Fig, 1, lanes 17, 20, 21). Thus, the efficiency of the PrPBSE-induced conversion of PrP-sen of a given host was found to be correlated with the in vivo transmissibility of BSE to that host. This correlation encouraged us to look for an in vitro indication of the transmissibility of BSE to humans, using a PrPBSE/human PrP-sen convesion assay.
Wild-type human (h) PrP has two common allelic forms that encode either methionine (hPrP-M) or valine (hPrP-V) at codon 129 (ref, 18). Hence we tested both types of hPrP-sen in conversion experiments. PrPBSE converted the ~25K aglycosyl forms of 35S-hPrP-M and 35S-hPrP-V to ~18K proteinaseK-resistant forms compatible with the proteinaseK-resistant core of PrP found in the human diseases (Fig. 1, lanes 2, 3, 4, 14; Fig, 2, lane 8). When more glycosylated 35S-hPrP-M was used, the proteinaseK-resistant conversion product was still largely restricted to an ~18K band, suggesting preferential conversion of the 25K aglycosylated form of 35S-hPrP-M (Fig, 1, lane 9). Little or no spontaneous formation of proteinaseK-resistant PrP was observed in the absence of PrPres (Fig, 1, bottom). The 35S-hPrP-V labels were converted by PrPBSE roughly threefold less efficiently than the 35-hPrP-M labels (Fig. 1, lanes 2-5,9, 10; Fig. 2, lanes 8, 9; Fig. 4; P = 0.0025). So far the new variant CJD has been found only in patients homozygous for methionine at codon 129 (ref. 2) and, although it may be premature ...
Figure I Phosphor autoradiographic analysis of SDS-PAGE gels of cell-free conversion products formed on incubation of PrPBSE with 35S-PrP-sen from different species. The molecular size range of 35S-radiolabelled PrP proteins following incubation with PrPBSE is seen before (-PK) or after (+PK) digestion with proteinase K in the top two panels (+PrPBSE, -PK and +PrPBSE, +PK). The effect of omitting PrPBSE from the incubation is shown in the bottorn panels (-PrPBSE, +PK). The migration of Mr markers is shown on the right. The ~18K PK-resistant 35S-PrP product is marked by the arrowhead. Equal amounts of PrPBSE (~0.5 µg) were used in each reaction with an approximately equivalent amount of radioactive PrP-sen (~25,000-40,000 c.p.m. put reaction; apart from boPrP-sen reactions. where ~10,000 c.p.m. was used owing to inefficient labelling in the bovine cells). The reaction sample was divided for analysis ~1 : 10(-PK; +PK) between the gels shown in the top two panels (and also between the top and bottom panels). Results are shown using differing concentrations of GdnHCl in the pretreatment and conversion phases of the reaction (see Methods). Conversions using 35S-PrP-sen molecules labelled in the presence (+) or absence (-) of tunicamycin are shown. The relative intensities of bands in the +PrPBSE, +PK panel need not reflect the efficiency of conversion as the specific activities of each label are not always the same: the percentage of label converted to proteinase K-resistant fragments (summarized in Fig. 4) is a better indicator of this efficiency than absolute intensities of the PK-resistant bands in the +PK, +PrPBSE panel.
IN
Jeweils etwa 500 ng aus einer millionenfach größeren Menge BSE-infektiösen Gehirnes isoliertes proteaseresistentes Prionprotein wurden gemischt mit etwa gleich großen Mengen radioaktiv markierten normalen Prionproteins von verschiedenen Spezies. In den Gefäßen mit normalem Prionproteinen von Rind, Schaf (Typ AQ und VQ) und Mensch bildete sich binnen 3 Tagen proteaseresistentes markiertes Prionprotein, während das Schafsprionprotein vom Typ AR, sowie Hamster-Prionprotein nicht meßbar umgewandelt wurde. Allerdings entstand fast nur nicht glykosyliertes proteaseresistentes Prionprotein. Durch aus BSE-Gehirnen gereinigtes proteaseresistentes Prionprotein wurde normales Rinderprionprotein etwa 10-fach effektiver als menschliches PrP-129-Met und 30-fach effektiver als menschliches PrP-129-Val umgewandelt.
Infektiöses Agens von 129-Methionin-homozygoten CJD und vCJD-Patienten wandelten normales menschliches Prionprotein vom Typ 129-Methionin etwa dreimal so effizient wie normales menschliches Prionprotein vom Typ 129-Valin um. Dabei entstand wiederum hauptsächlich nicht glykosyliertes proteaseresistentes Prionprotein.
Scrapie-infektiöses Material mit dem Prionprotein PrP-VQ wandelte normales menschliches PrP-129-Met ebenso effektiv wie normale Prionproteine von Schafen vom relativ resistenten Typ PrP-AR sowie von Mäusen um. Normales Prionprotein von Hamstern wurde nicht erkannbar umgewandelt, während normales Prionprotein von Schafen der empfänglicheren Typen Prp-VQ und PrP-AQ deutlich effizienter als das menschliche umgewandelt wurde.
Diese in vitro Ergebnisse korrelieren exakt mit entsprechenden in vivo Beobachtungen und sprechen daher dafür, dass die Speziesbarrieren ebenso wie die unterschiedlichen Empfänglichkeiten verschiedener Schafrassen und menschlicher Genotypen bei den Prionkrankheiten hauptsächlich auf den Sequenzunterschieden der Prionproteine im infektiösen Agenz bzw im infizierten Organismus beruhen. Dies unterstützt wiederum ebenso wie Möglichkeit einer in vitro Umwandlung die Priontheorie.
Die Ergebnisse bedeuten aber auch, dass der Mensch für Scrapie etwa genauso empfänglich wie für BSE ist, bzw. das bisher nur Opfer von BSE-Infektionen entdeckt wurden, weil Menschen sehr viel mehr infektiöses Material von Rindern als von Schafen aufgenommen haben.
Andere in vitro Experimente wurden beschrieben in AODT,ABHU,ACJQ,AGQC,AGQD,ABNP,ABHT [AJWV].
ZR 26 Zitate
MH Animal; Cattle; Cell-Free System; Creutzfeldt-Jakob Syndrome/transmission; Disease Susceptibility; Encephalopathy, Bovine Spongiform/*transmission; Endopeptidase K/metabolism; Hamsters; Human; Mesocricetus; Mice; Nerve Tissue Proteins/metabolism; PrPc Proteins/genetics/*metabolism; PrPsc Proteins/*metabolism; Prions/metabolism; Scrapie/*transmission; Sheep; Species Specificity; Support, Non-U.S. Gov't; Support, U.S. Gov't, P.H.S.; Tumor Cells, Cultured; Zoonoses/*transmission
AD
Gregory J. Raymond*, James Hope+, David A. Kocisko*%, Suzette A. Priola*, Lynne D. Raymond*, Alex Bossers§, James lronside!, Robert G. Will!, Shu G. Chen), Robert B. Petersen), Pierluigi Gambetti), Richard Rubenstein#, Mari A. Smits§, Peter T. Lansbury Jr%°& Byron Caughey*
*Rocky Mountain Laboratories, NIAID, National Institutes of Health, Hamilton,
Montana 59840, USA
+ BBSRC Institute for Animal Health, Compion Laboratory, Compton, Newbury, Berkshire RG20 7NN, UK
% Massachusetts Institute of Technology, Department of Chemistry, Cambridge, Massachusetts 02139, USA
§ Department of Bacteriology, DLO-Institute for Animal Science and Health, P.O. Box 65. NL-8200 AB Lelystad, The Netherlands
! Western General Hospital, CJD Surveillance Unit, Edinburgh EH4 2XU, UK
) Case Western Reserve University, Institute of Pathology, 2085 Adelbert Road, Cleveland, Ohio 44106, USA
# NYS Institute for Basic Research, 1050 Forest Hill Road, Staten Island, New York 10314, USA
°Center for Neurological Diseases, Brigham & Womens Hospital, Harvard Medical School, Boston, Masachusetts 02115, USA
SP englisch
PO England
OR Prion-Krankheiten 7
ZF kritische Zusammenfassung von Roland Heynkes