BACKGROUND INFORMATION: Cellular prion protein (PrPC ) is infamous for its role in prion diseases. The physiological function of PrPC remains enigmatic, but several studies point to its involvement in cell differentiation processes. To test this possibility, we monitored PrPC changes during the differentiation of prion-susceptible CAD 5 cells, and then we analysed the effect of PrPC ablation on the differentiation process. RESULTS: Neuronal CAD 5 cells differentiate within 5 days of serum withdrawal, with the majority of the cells developing long neurites. This process is accompanied by an up to sixfold increase in PrPC expression and enhanced N-terminal β-cleavage of the protein, which suggests a role for the PrPC in the differentiation process. Moreover, the majority of PrPC in differentiated cells is inside the cell, and a large proportion of the protein does not associate with membrane lipid rafts. In contrast, PrPC in proliferating cells is found mostly on the cytoplasmic membrane and is predominantly associated with lipid rafts. To determine the importance of PrPC in cell differentiation, a CAD 5 PrP-/- cell line with ablated PrPC expression was created using the CRISPR/Cas9 system. We observed no considerable difference in morphology, proliferation rate or expression of molecular markers between CAD 5 and CAD 5 PrP-/- cells during the differentiation initiated by serum withdrawal. CONCLUSIONS: PrPC characteristics, such as cell localisation, level of expression and posttranslational modifications, change during CAD 5 cell differentiation, but PrPC ablation does not change the course of the differentiation process. SIGNIFICANCE: Ablation of PrPC expression does not affect CAD 5 cell differentiation, although we observed many intriguing changes in PrPC features during the process. Our study does not support the concept that PrPC is important for neuronal cell differentiation, at least in simple in vitro conditions.

• MeSH
• buněčná diferenciace * MeSH
• buněčné linie MeSH
• membránové mikrodomény MeSH
• myši MeSH
• neurony cytologie   metabolismus   MeSH
• posttranslační úpravy proteinů MeSH
• priony metabolismus   MeSH
• PrPC proteiny metabolismus   MeSH
• zvířata MeSH
• Check Tag
• myši MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH

Plasma levels of circulating platelet extracellular vesicles (PEVs) are an emerging marker of platelet activation, thrombosis, inflammation, and endothelial dysfunction. Analysis of PEVs in cord blood of preterm newborns may reflect the underlying pathology and possibly serve as a new diagnostic and prognostic tool. However, collection, preparation and analysis of cord blood samples in clinical settings is a logistically complex process. We have studied the effect of delay in sample preparation and sample freezing on the PEV analysis by flow cytometry. PEVs in the cord blood plasma were identified after double labelling with monoclonal antibodies CD36+CD41 or CD41+CD62. Both, the delay and the freezing significantly affected the count and often also fluorescence of the detected PEVs. Additionally, our pilot study utilizing fresh cord blood samples of term and preterm newborns demonstrated significantly decreased CD36 and CD62 PEV fluorescence in preterm newborns. Our data highlight the importance of pre-analytical steps in the analysis of cord blood PEVs and suggest that not only the count, but also the level of PEV fluorescence may have possible diagnostic potential.

• MeSH
• extracelulární vezikuly * MeSH
• fetální krev * MeSH
• lidé MeSH
• novorozenec MeSH
• pilotní projekty MeSH
• průtoková cytometrie MeSH
• zmrazování MeSH
• Check Tag
• lidé MeSH
• novorozenec MeSH
• Publikační typ
• časopisecké články MeSH

Prion disorders are fatal neurodegenerative diseases caused by the autocatalytic conversion of a natively occurring prion protein (PrPC ) into its misfolded infectious form (PrPTSE ). The proven resistance of PrPTSE to common disinfection procedures increases the risk of prion transmission in medical settings. Herein, we present the effective photodynamic inactivation (PDI) of prions by disulfonated hydroxyaluminum phthalocyanine (AlPcOH(SO3 )2 ) utilizing two custom-built red light sources. The treatment eliminates PrPTSE signal in infectious mouse brain homogenate with efficiency that depends on light intensity but has a low effect on the overall protein content. Importantly, singlet oxygen (O2 (1 Δg )) is the only species significantly photogenerated by AlPcOH(SO3 )2 , and it is responsible for the PDI of prions. More intensive light conditions show not only higher O2 (1 Δg ) production but also decreases in AlPcOH(SO3 )2 photostability. Our findings suggest that PDI by AlPcOH(SO3 )2 -generated O2 (1 Δg ) represents a promising approach for prion inactivation that may be useful in future decontamination strategies for delicate medical tools.

• MeSH
• fotochemoterapie * MeSH
• fotosenzibilizující látky chemie   farmakologie   MeSH
• indoly chemie   farmakologie   MeSH
• kyseliny sulfonové chemie   MeSH
• mozek účinky léků   metabolismus   účinky záření   MeSH
• myši MeSH
• prionová bílkovina metabolismus   MeSH
• singletový kyslík metabolismus   MeSH
• zvířata MeSH
• Check Tag
• myši MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

• Publikační typ
• abstrakt z konference MeSH