Figure 4.

QS-21 accumulates in lysosomes and promotes their destabilization. (A) Differentiated THP-1 cells were pulsed with ¹⁴C-QS-21 for 4 h followed by an overnight chase, and postnuclear particles were resolved by density gradients fractionation. Na+/K+-ATPase identifies the plasma membrane and N-acetyl-β-hexosaminidase activity identifies lysosomes. The abscissa axis represents the different fractions. The ordinate reflects enrichment (C/C_i > 1) or depletion (C/C_i < 1) vs. initial abundance (C_i = 1), indicated by dotted lines. (B) Differentiated THP-1 cells were co-incubated with BODIPY-QS-21 and Lysotracker Red then analyzed by vital confocal microscopy. (C) moDCs were incubated with 1 µg/ml acridine orange (AO), washed, and stimulated for 2 h with QS-21 (10 µg/ml). AO fluorescence was quantified by flow cytometry and a 610 nm filter. Cells with decreased AO fluorescence are indicated with the double arrow. One representative donor of 3 is shown. (D) moDCs were incubated with Lucifer Yellow, 10 kDa dextran-Alexa633 and 40 kDa dextran-Texas Red for 16 h then stimulated for QS-21 for 4 h. The localization of fluorescent markers was observed by confocal microscopy. The proportion of cells with punctate (lysosomal), diffuse (cytoplasmic) or both (punctate + diffuse) signal for the different fluorescent markers was determined with ImageJ. The data are representative of four donors.