Fig 1: Effects of U18666A on mitochondrial cholesterol trafficking and HMβCD modulation in human fibroblasts. Fibroblasts from control patients treated with U18666A (2 μg/μl, 16 h) were co-treated with HMβCD (2 mM, 16 h) to determine (A) Mitochondria and Filipin co-staining for confocal microscopy analyses. (B) Staining markers colocalization analysis using Image J software. (C) StARD1, and ACDase protein expression analyzed by Western blot. Data are presented as means ± SEM (n > 5, Unpaired Student's t-test (two-tailed)). *p < 0.05, **p < 0.01, ****p < 0.0001 vs. U18666A or control.
Fig 2: Effects of HMßCD on hepatocytes from Npc1+/+mice treated with U18666A. Primary mouse hepatocytes from Npc1+/+ treated with U18666A (2 µg/µl, 16 h) and primary mouse hepatocytes from Npc1-/- were co-treated with HMßCD (2 mM, 16 h) to determine (A–C) Mitochondria and Filipin co-staining for confocal microscopy analyses. Staining markers colocalization analysis using Image J software. (B–D) StARD1, and ACDase protein expression analyzed by Western blot. Data are presented as means ± SEM (n > 5, Unpaired Student's t-test (two-tailed)). *p < 0.05, **p < 0.01 vs. U18666A or Npc1-/-.
Fig 3: Effects of U18666A treatment on liver mitochondrial cholesterol trafficking. Primary mouse hepatocytes from Npc1+/+ were treated with U18666A (2 µg/µl, 16 h) to determine (A) Lysosomes and Filipin co-staining, (B) Mitochondria and Filipin co-staining for confocal microscopy analyses. Staining markers colocalization analysis using Image J software. (C) StARD1, ACDase and (D) ER stress markers protein expression analyzed by Western blot. Data are presented as means ± SEM (n > 5, Unpaired Student's t-test (two-tailed)). *p < 0.05, ***p < 0.001 vs. Control.
Fig 4: ACDase overexpression in fibroblasts from NPC patients. Fibroblasts from NPC patients were transfected with a sobrexpression ACDase vector or the scrambled control-GFP vector to analyze (A) GFP and ACDase mRNA expression and (B) StARD1, and ACDase protein expression by Western blot. (C) LRH expression in fibroblasts from patients with NPC disease. Data are presented as means ± SEM (n = 3, Unpaired Student's t-test (two-tailed)). (D) Mitochondria and Filipin co-staining for confocal microscopy analyses. Staining markers colocalization analysis using Image J software. Data are presented as means ± SEM (n > 5, Unpaired Student's t-test (two-tailed)). *p < 0.05, **p < 0.01 vs. control GFP.
Fig 5: ACDase and ER stress markers protein expression in livers from Npc1−/−mice and fibroblasts from NPC patients. A) ER stress markers and (B) ACDase protein expression analyzed by Western blot. Data are presented as means ± SEM (n > 3, Unpaired Student's t-test (two-tailed)). ***p < 0.001, ****p < 0.0001 vs. Npc1+/+.
Supplier Page from MilliporeSigma for Anti-ASAH1 antibody produced in rabbit