Journal: Cell & Bioscience

Article Title: Cellular prion protein and calcium ions trigger the neurotoxicity of α-synuclein aggregates

doi: 10.1186/s13578-025-01479-7

Figure Lengend Snippet: PrP C partially mediates the binding of αS species to neuronal membranes. A Representative STED microscopy images showing primary rat cortical neurons treated for 60 min with OB* and SF at 0.3 µM (monomer equivalents). Red and green fluorescence: MAP-2 and αS species detected by rabbit anti-MAP-2 and mouse monoclonal 211 Abs, respectively. The histograms show the Pearson’s and Manders' coefficients (M1 and M2) determined using the ImageJ software and JACOP plugin (n = 3). B Representative confocal microscopy images of SH-SY5Y cells treated for 10 min with 488-labelled OB* and SF at 0.3 μM (monomer equivalents). Green and red fluorescence: 488-labelled αS and PrP C detected with the mouse monoclonal Ab anti-PrP c , respectively. The histograms show the Pearson’s and Manders' coefficients (M1 and M2) determined as in ( A ) (n = 10 for OB* and n = 6 for SF). C Representative confocal microscopy images showing the immunostaining of PrP C in SH-SY5Y cells pre-transfected with control siRNA, or with PrP C siRNA. The histogram shows the semi-quantitative analysis of the PrP C -derived fluorescence (n = 3). D Western blot analysis of PrP C levels in SH-SY5Y cells pre-transfected with control siRNA or with PrP C siRNA. The histogram shows the semi-quantitative analysis of PrP C levels as the percentage relative to cells transfected with control siRNA, after normalizing each band to vinculin, taken as housekeeping (n = 3). E Representative confocal microscopy images of the apical sections of SH-SY5Y cells pre-transfected with control siRNA or with PrP C siRNA, and then treated for 10 min with OB* and SF at 0.3 μM (monomer equivalents). Red and green fluorescence: cell membranes and αS species detected by WGA and rabbit polyclonal anti-αS Ab , respectively. The histograms show the Pearson’s and Manders' coefficients (M1 and M2) determined as in ( A ); (n = 14 for OB* and n = 9 for SF in cells pre-treated with control siRNA; n = 11 for OB* and n = 10 for SF in cells pre-treated with PrP C siRNA). In all panels, experimental errors are S.E.M. Samples were analyzed by Student t test (** P < 0.01 and *** P < 0.001 relative to cells treated with OB* in panels A, B); °° P < 0.01 and °°° P < 0.001 relative to corresponding cells treated with control siRNAs in panels C, D, E

Article Snippet: Membranes were blocked with the EveryBlot blocking buffer (12010020, Bio-Rad) and then incubated overnight with 1:800 diluted rabbit anti-PrP C polyclonal Ab (12555-1-AP, Proteintech) at 4 °C.

Techniques: Binding Assay, Microscopy, Fluorescence, Software, Confocal Microscopy, Immunostaining, Transfection, Control, Derivative Assay, Western Blot

Journal: Cell & Bioscience

Article Title: Cellular prion protein and calcium ions trigger the neurotoxicity of α-synuclein aggregates

doi: 10.1186/s13578-025-01479-7

Figure Lengend Snippet: PrP C partially mediates the binding of αS species to neuronal membranes and the neuronal uptake of αS oligomers released by mature fibrils A Characterization of human iPSC-derived dopaminergic neurons. Representative confocal microscopy images of human iPSC-derived dopaminergic neurons expressing MAP-2 (red) and TH (green). Nuclei were stained with DAPI (blue). B Representative confocal microscopy images of the apical sections of human iPSC-derived dopaminergic neurons and primary rat cortical neurons treated for 10 min with OB* at 0.3 μM (monomer equivalents) in the absence or presence of a pre-treatment for 30 min with anti-PrP C Ab at an OB*:Ab molar ratio of 1:1 (monomer equivalents). Red and green fluorescence: cell membranes detected by WGA and OB* detected with the rabbit polyclonal anti-αS Ab, respectively. The histogram shows the Pearson’s coefficient determined using the ImageJ software and JACOP plugin (n = 3). C Representative confocal microscopy images showing human iPSC-derived dopaminergic neurons treated for 6 h with OB* and SF at 0.3 µM (monomer equivalents) in the absence or in the presence of a pre-treatment for 30 min with anti-PrP C Ab at a αS:Ab molar ratio of 1:0.5. Red and green fluorescence: cell membranes detected by WGA and the A11-positive prefibrillar oligomers, respectively. The semi-quantitative analysis of the A11-derived fluorescence was expressed as the percentage of the value for untreated cells. Experimental errors are S.E.M. (n = 3). Samples were analysed by Student t test (*** P < 0.001 relative to untreated cells); (° P < 0.05, °° P < 0.01, and °°° P < 0.001 relative to cells treated with αS species in the absence of pre-treatment with anti-PrP C Ab)

Article Snippet: Membranes were blocked with the EveryBlot blocking buffer (12010020, Bio-Rad) and then incubated overnight with 1:800 diluted rabbit anti-PrP C polyclonal Ab (12555-1-AP, Proteintech) at 4 °C.

Techniques: Binding Assay, Derivative Assay, Confocal Microscopy, Expressing, Staining, Fluorescence, Software