ca074me  (TargetMol)

Journal: Molecular Neurodegeneration

Article Title: The Parkinson’s disease risk gene cathepsin B promotes fibrillar alpha-synuclein clearance, lysosomal function and glucocerebrosidase activity in dopaminergic neurons

doi: 10.1186/s13024-024-00779-9

Figure Lengend Snippet: Cathepsin B inhibitors potentiate the effect of α-syn PFFs on dopaminergic neurons: A ) CatB activity measured by fluorogenic assay in DA neurons treated with 1 μM CA074me. B Representative immunofluorescent images from high-content confocal imaging of DA neurons treated with CA074me (1 μM) and/or α-syn PFFs (300 nM) and stained for Map2 and pSyn-S129. C Quantification of pSyn-S129 in Map2-positive cells 3-weeks after PFF and/or CA074me treatment in either Control (AIW002-2) DA neurons or isogenic neurons lacking endogenous α-syn (AIW002-2 SNCA-KO). D Quantification of pSyn-S129 in control DA neurons 2, 3, or 4 weeks after PFF (300 nM) and/or CA074me treatment. Bonferroni-corrected t-tests, ** p < 0.01, *** p < 0.001, **** p < 0.0001

Article Snippet: CA074me (Selleckchem) and PADK (Bachem) treatments were performed at the indicated final concentrations with DMSO as vehicle.

Techniques: Activity Assay, Imaging, Staining, Control

Journal: Molecular Neurodegeneration

Article Title: The Parkinson’s disease risk gene cathepsin B promotes fibrillar alpha-synuclein clearance, lysosomal function and glucocerebrosidase activity in dopaminergic neurons

doi: 10.1186/s13024-024-00779-9

Figure Lengend Snippet: Cathepsin B inhibition increases lysosome abundance but impairs function in dopaminergic neurons. A Representative live-cell confocal images of neuronal cell bodies stained with lysotracker-green 72-h after exposure to alexa-633 labelled α-syn PFFs (80 nM). B Lysosome density per cell body, measured as the percentage of lysotracker-positive area per cell soma. C PFF density per cell body, measured as the percentage of PFF-633-positive area per cell soma. D Colocalization of lysotracker and PFF-633 measured using Pearson’s coefficient per cell soma. E Representative immunofluorescent images from high-content confocal imaging of DA neurons treated with CA074me (1 μM) and/or α-syn PFFs (300 nM) for 3 weeks and stained for Map2 and LAMP1. F Quantification of LAMP1 in Map2-positive cells. G Lysosomal degradative capacity measured by fluorescence intensity of DQ-BSA fluorogenic probe 24-h after dye loading. H Quantification of lysosome velocity in neurites measured by live-cell confocal imaging and quantified using TrackMate. Points represent individual quantified image fields derived from 6 independent experiments. I Representative images of neurons stained with lysotracker deep-red and PFB-FDGlu fluorescent signal at baseline and 100 min after dye-loading. J Quantification of PFB-FDGlu fluorescence per cell in DA neurons pre-treated for 24-h with CA074me or PADK. K Quantification of the slope of PFB-FDGlu fluorescence versus time. T-test or Bonferroni-corrected t-tests, * p < 0.05, ** p < 0.01, **** p < 0.0001

Article Snippet: CA074me (Selleckchem) and PADK (Bachem) treatments were performed at the indicated final concentrations with DMSO as vehicle.

Techniques: Inhibition, Staining, Imaging, Fluorescence, Derivative Assay

Journal: Molecular Neurodegeneration

Article Title: The Parkinson’s disease risk gene cathepsin B promotes fibrillar alpha-synuclein clearance, lysosomal function and glucocerebrosidase activity in dopaminergic neurons

doi: 10.1186/s13024-024-00779-9

Figure Lengend Snippet: CTSB inhibition promotes pSyn-S129 accumulation in patient-derived midbrain organoids. A Representative immunofluorescent images of Map2 and pSyn-S129 in 5-month old SNCA-triplication (3xSNCA) and isogenic SNCA-KO midbrain organoids treated with vehicle (DMSO) or 1 μM CA074me for 60 days. Large images depict representative whole-organoids and high-magnification images depict individual Map2-positive cells. B Quantification of the pSyn-S129 positive area of the organoid relative to the total organoid size. C Quantification of the Map2-positive area relative to organoid size. D Quantification of pSyn-S129- and Map2 double-positive area in 3 × SNCA organoids. E Quantification of the percentage of cells positive for both Map2 and pSyn-S129 in 3 × SNCA organoids. Bonferroni-corrected t-tests, * p < 0.05

Article Snippet: CA074me (Selleckchem) and PADK (Bachem) treatments were performed at the indicated final concentrations with DMSO as vehicle.

Techniques: Inhibition, Derivative Assay