Distinctive Properties of Plaque-Type Dura Mater Graft-Associated Creutzfeldt–Jakob Disease in Cell-protein Misfolding Cyclic Amplification

Atsuko Takeuchi; Atsushi Kobayashi; Piero Parchi; Masahito Yamada; Masanori Morita; Shusei Uno; Tetsuyuki Kitamoto


Lab Invest. 2016;96(5):581-587. 

In This Article

Abstract and Introduction


There are two distinct subtypes of dura mater graft-associated Creutzfeldt–Jakob disease (dCJD) with methionine homozygosity at codon 129 of the PRNP gene. The majority of cases is represented by a non-plaque-type (np-dCJD) resembling sporadic CJD (sCJD)-MM1 or -MV1, while the minority by a plaque-type (p-dCJD). p-dCJD shows distinctive phenotypic features, namely numerous kuru plaques and an abnormal isoform of prion protein (PrPSc) intermediate in size between types 1 and 2. Transmission studies have shown that the unusual phenotypic features of p-dCJD are linked to the V2 prion strain that is associated with sCJD subtypes VV2 or -MV2. In this study, we applied protein misfolding cyclic amplification (PMCA) using recombinant human prion protein as a substrate and demonstrated that p-dCJD prions show amplification features that are distinct from those of np-dCJD. Although no amplification of np-dCJD prions was observed with either 129 M or 129 V substrate, p-dCJD prions were drastically amplified with the 129 V substrates, despite the PRNP codon 129 incompatibility between seed and substrate. Moreover, by using a type 2 PrPSc-specific antibody not recognizing PrPSc in p-dCJD, we found that type 2 products are generated de novo from p-dCJD prions during PMCA with the 129 V substrates. These findings suggest that our cell-PMCA is a useful tool for easily and rapidly identifying acquired CJD associated with the transmission of the V2 CJD strain to codon 129 methionine homozygotes, based on the preference for the 129 V substrate and the type of the amplified products.


Creutzfeldt–Jakob disease (CJD) is a fatal neurodegenerative disease. The large majority of CJD cases are thought to be caused by a spontaneous conformational change of the normal monomeric isoform of the prion protein (PrPC) into an abnormal isoform (PrPSc), as in sporadic CJD (sCJD) or genetic/familial CJD. On the other hand CJD can also be acquired through PrPSc infection, as in variant CJD or iatrogenic CJD (iCJD). The wide heterogeneity of sCJD clinico-pathological phenotypes depends on both the genotype (methionine (M) or valine (V)) at the polymorphic codon 129 of the PRNP gene and the type (1 or 2) of PrPSc accumulating in the brain.[1] Type 1 and type 2 can be distinguished according to the size of the proteinase K-resistant core of the protein (21 and 19 kDa, respectively) on western blots. Based on the polymorphism at codon 129 of the PRNP gene and the type of PrPSc, sCJD patients are classified into six major subtypes: MM1/MV1, MM2 cortical, MM2 thalamic, VV1, VV2, and MV2.[1]

iCJD is caused by the transmission of prions via cadaveric pituitary hormones, dura mater, and corneal grafts, or contaminated neurosurgical instruments. Many cases of dura mater graft-associated CJD (dCJD) have been reported in Japan.[2–4] There are two distinct phenotypes in dCJD with methionine homozygosity at codon 129. The first is a major group represented by a non-plaque-type dCJD (np-dCJD), the second a minor group represented by a plaque-type dCJD (p-dCJD).[5–7] np-dCJD shares phenotypic characteristics such as diffuse synaptic-PrP deposition and type 1 PrPSc with sCJD-MM1 or -MV1, the most common sCJD phenotype (denoted as M1 strain in transmission studies).[4,8] In contrast, p-dCJD is characterized by unusual phenotypic features such as the presence of numerous kuru plaques and a unique PrPSc type with an electrophoretic mobility of about 20 kDa, which is intermediate in size between types 1 and 2 (type i PrPSc).[9]

We previously demonstrated that the transmission properties of p-dCJD prions resemble those of sCJD-VV2 or -MV2 prions and, in particular, that the transmission of sCJD-VV2 prions to humanized knock-in mice carrying the 129 MM genotype produce type i PrPSc and kuru plaques, indicating that this subtype of dCJD is caused by cross-sequential transmission of sCJD-VV2 or -MV2 (denoted as V2 strain),[8] the second most common CJD strain, to individuals with codon 129 methionine homozygosity.[9–11] Interestingly, type i PrPSc and kuru plaques in patients homozygous for methionine at codon 129 have only been observed in acquired prion diseases, especially in p-dCJD.[9,12–15]

A correct diagnosis and classification of cases is of critical importance for CJD surveillance centers worldwide aiming to identify potentially new disease forms or risk factors. Although PrPSc typing may be considered a tool for identification of the CJD etiology (eg, sporadic vs acquired) when no reliable evidence of CJD infection is available from clinical records, it is rather difficult to differentiate type i from type 1 PrPSc in routine western blot analysis due to the subtle differences in their electrophoretic mobility. In a transmission study with knock-in humanized mice, we have recently demonstrated, for example, that two cases with atypical CJD-MM with plaques, reported as sCJD in the literature, were actually acquired CJD cases caused by the V2 strain.[16] Therefore, the most reliable current approach to identify acquired CJD is to carry out expensive and time-consuming experimental transmission studies. Under these circumstances, this study was undertaken to establish a new, rapid, and reliable method for the differentiation of acquired CJD from sCJD with PMCA using cell lysates containing exogenously expressed human PrPC as a substrate (denoted as cell-PMCA).[17] We investigated the relative amplification efficiency of PrPSc and characterized PrPSc amplified products in patients with p-dCJD, np-dCJD, and sCJD. Based on the results obtained, we propose that p-dCJD (V2 strain) can be reliably distinguished from np-dCJD (M1 strain) according to distinctive amplification properties.