Figure 6.

Exogenous acid SMase decreases cholesterol accumulation in LSOs in human NPC fibroblasts. Parallel sets of human WT (GM05659) and NPC (GM03123) Fbs were incubated in medium alone or, in the case of the NPC fibroblasts, medium containing 3 μg/mL recombinant human acid SMase (rhASM). Two days later, the cells were washed thoroughly with PBS and either lysed and assayed for acid SMase activity (A) or fixed and stained with filipin for imaging and quantification (B). The images are displayed with the same gray scale range. Scale bar, 15 μm. The quantified data in the bar graph represent LSO ratios normalized to the WT Fb values (average of 60–66 images from three independent experiments ±SEM). C) Another human NPC Fbs (GM18453) was incubated with 3 μg/mL rhASM for 24 h, unlike 48 h in NPC1 (GM03123). Quantified data shown in bar chart are the normalized LSO ratios (normalized to untreated NPC Fbs) in the presence and absence of recombinant enzyme. Data represents an average from two independent experiments and 30–36 images ±SEM, p < 0.0001. The values for NPC fibroblasts in panels A, B and C were significantly different from both the WT Fbs and the NPC fibroblasts treated with rhASM (p < 0.0001).