Figure 4.

K‐80003 decreases p62/sequestosome 1 expression through promoting autophagic flux. Bone marrow‐derived macrophages (BMDMs; a) or RAW264.7 cells (b) were treated with 7‐keto‐cholesterol (7‐KC; 40 μM) together with or without K‐80003 (20 μM) for 16 hr, and bafilomycin A1 (BafA1; 50 nM) or chloroquine (CQ; 15 μg·ml⁻¹) was added to cultures for the final 4 hr. Lysates were prepared and examined by immunoblotting (n = 5). BMDMs (c) or RAW264.7 cells (d) were treated with 7‐KC (40 μM) in the absence or presence of the indicated concentrations of K‐80003 for 16 hr. Lysates were prepared and analysed by immunoblotting using appropriate antibodies (n = 5). (e) BMDMs were treated with 7‐KC (40 μM) with or without K‐80003 (20 μM) for 16 hr, and BafA1 (50 nM) or chloroquine (15 μg·ml⁻¹) was added to cultures for the final 4 hr. Lysates were prepared and examined by immunoblotting (n = 5). (f) BMDMs were pretreated with 7‐KC (40 μM) alone or together with K‐80003 (20 μM) for 12 hr and then co‐treated with chloroquine (15 μg·ml⁻¹) for another 4 hr. Immunoblotting was applied to examine IκBα expression (n = 5). ^* P < .05, significantly different as indicated; ns, P > .05