Rapid and highly sensitive detection of variant creutzfeldt - jakob disease abnormal prion protein on steel surfaces by protein misfolding cyclic amplification: application to prion decontamination studies

The prevalence of variant Creutzfeldt-Jakob disease (vCJD) in the population remains uncertain, although it has been estimated that 1 in 2000 people in the United Kingdom are positive for abnormal prion protein (PrPTSE) by a recent survey of archived appendix tissues. The prominent lymphotropism of vCJD prions raises the possibility that some surgical procedures may be at risk of iatrogenic vCJD transmission in healthcare facilities. It is therefore vital that decontamination procedures applied to medical devices before their reprocessing are thoroughly validated. A current limitation is the lack of a rapid model permissive to human prions. Here, we developed a prion detection assay based on protein misfolding cyclic amplification (PMCA) technology combined with stainless-steel wire surfaces as carriers of prions (Surf-PMCA). This assay allowed the specific detection of minute quantities (10(-8) brain dilution) of either human vCJD or ovine scrapie PrPTSE adsorbed onto a single steel wire, within a two week timeframe. Using Surf-PMCA we evaluated the performance of several reference and commercially available prion-specific decontamination procedures. Surprisingly, we found the efficiency of several marketed reagents to remove human vCJD PrPTSE was lower than expected. Overall, our results demonstrate that Surf-PMCA can be used as a rapid and ultrasensitive assay for the detection of human vCJD PrPTSE adsorbed onto a metallic surface, therefore facilitating the development and validation of decontamination procedures against human prions

Rapid and highly sensitive detection of variant Creutzfeldt - Jakob Disease prions binded on steel surfaces by PMCA: Application to Prion decontamination studies.

International audienc

Rapid and highly sensitive detection of variant Creutzfeldt - Jakob Disease prions binded on steel surfaces by PMCA: Application to Prion decontamination studies.

International audienc

Artificial neural network prediction of clozapine response with combined pharmacogenetic and clinical data.

Although one third to one half of refractory schizophrenic patients responds to clozapine, however, there are few evidences currently that could predict clozapine response before the use of the medication. The present study aimed to train and validate artificial neural networks (ANN) , using clinical and pharmacogenetic data, to predict clozapine response in schizophrenic patients. Five pharmacogenetic variables and five clinical variables were collated from 93 schizophrenic patients taking clozapine, including 26 responders. ANN analysis was carried out by training the network with data from 75% of cases and subsequently testing with data from 25% of unseen cases to determine the optimal ANN architecture. Then the leave-one-out method was used to examine the generalization of the models. The optimal ANN architecture was found to be a standard feed-forward, fully- connected, back-propagation multilayer perceptron. The overall accuracy rate of ANN was 83.3%, which is higher than that of logistic regression (LR) (70.8%). By using the area under the receiver operating characteristics curve as a measure of performance, the ANN outperformed the LR (0.821 + /- 0.054 versus 0.579 +/- 0.068; p < 0. 001). The ANN with only genetic variables outperformed the ANN with only clinical variables (0.805 +/- 0.056 versus 0.647 +/- 0.066; p = 0.046). The gene polymorphisms should play an important role in the prediction. Further validation of ANN analysis is likely to provide decision support for predicting individual response

Prion amplification techniques for the rapid evaluation of surface decontamination procedures.

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Prion amplification techniques for the rapid evaluation of surface decontamination procedures.

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Prion replication occurs in endogenous adult neural stem cells and alters their neuronal fate: involvement of endogenous neural stem cells in prion diseases.

International audiencePrion diseases are irreversible progressive neurodegenerative diseases, leading to severe incapacity and death. They are characterized in the brain by prion amyloid deposits, vacuolisation, astrocytosis, neuronal degeneration, and by cognitive, behavioural and physical impairments. There is no treatment for these disorders and stem cell therapy therefore represents an interesting new approach. Gains could not only result from the cell transplantation, but also from the stimulation of endogenous neural stem cells (NSC) or by the combination of both approaches. However, the development of such strategies requires a detailed knowledge of the pathology, particularly concerning the status of the adult neurogenesis and endogenous NSC during the development of the disease. During the past decade, several studies have consistently shown that NSC reside in the adult mammalian central nervous system (CNS) and that adult neurogenesis occurs throughout the adulthood in the subventricular zone of the lateral ventricle or the Dentate Gyrus of the hippocampus. Adult NSC are believed to constitute a reservoir for neuronal replacement during normal cell turnover or after brain injury. However, the activation of this system does not fully compensate the neuronal loss that occurs during neurodegenerative diseases and could even contribute to the disease progression. We investigated here the status of these cells during the development of prion disorders. We were able to show that NSC accumulate and replicate prions. Importantly, this resulted in the alteration of their neuronal fate which then represents a new pathologic event that might underlie the rapid progression of the disease

Optimization of steel wire contamination in 96-well microplates.

<p>Wires were contaminated with either 1% 127S-scrapie or 10% human vCJD brain homogenates at different times (1 min, 10 min or 1 hr) and subjected to one Surf-PMCA round. Protease-resistant prion protein was detected with 6D11 (127S-scrapie) or 3F4 (human vCJD) monoclonal antibodies. NEG: Negative control (wires mock-contaminated in normal brain homogenate). M: molecular mass marker.</p

Evaluation of “standard” decontamination procedures on 127S scrapie prions.

<p>Steel wires contaminated with 1% 127S scrapie brain homogenate were treated with “standard” prion decontamination procedures and subjected to three serial PMCA rounds. Three independent series of PMCA were performed on 4 wires per treatment (Black cross (<b>Χ)</b> Series 1, Red square (■) Series 2 and Green circle (●) Series 3). PAA: Per acetic acid 1.2%—60 min; NaOH: sodium hydroxide 0.1N—15 min or 1N—60 min; Auto: steam sterilization 121°C -20 min or 134°C -20 min; NaOCl: Sodium hypochlorite 0.2% (2000ppm)– 15 min or 2% (20000ppm)– 60 min; H₂O: untreated positive control; NEG: negative control (wire mock-contaminated with normal brain homogenate).</p

Earliest detection of prion seeding activity in blood of human PRP transgenic mice infected with vCJD

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