Research Paper Volume 13, Issue 1 pp 910—932

Tanshinone IIA attenuates atherosclerosis via inhibiting NLRP3 inflammasome activation

Tan IIA inhibits oxLDL-induced macrophage lysosomal damage and cathepsin B release. (A) Fluorescence confocal assay for oxLDL-induced cathepsin B release from lysosome of mouse B6 macrophages or cathepsin B-/- B6 macrophages under the different treatments, respectively. The cells were treated with oxLDL (50 μg/mL) alone (2), oxLDL plus 2.5 μg/mL Tan IIA (3), oxLDL plus 10 μg/mL Tan IIA (4) and oxLDL plus Cathepsin B inhibitor CA-074-Me (5). The Cathepsin B-/- mouse macrophages (6) were treated with oxLDL alone. The small interfering RNA-mediated CD36 silenced-mouse B6 macrophages (7) were treated with oxLDL alone. All the cells were stained with Alexa 647-conjugated cholera toxin B (membrane staining, red), and then with DQ ovalbumin (cathepsin B staining, green) and Hoechst dye (nuclei staining, blue) after saponin treatment. (B) Western blot detection for Cathepsin B releases from cytosol of oxLDL-induced B6 macrophages or cathepsin B-/- B6 macrophages treated with digitonin and phenylmethylsulfonyl fluoride (PMSF). (C) ELISA assay for oxLDL-induced Il-1β releases from mouse B6 macrophages or cathepsin B-/- B6 macrophages under the different treatments.

Figure 6. Tan IIA inhibits oxLDL-induced macrophage lysosomal damage and cathepsin B release. (A) Fluorescence confocal assay for oxLDL-induced cathepsin B release from lysosome of mouse B6 macrophages or cathepsin B-/- B6 macrophages under the different treatments, respectively. The cells were treated with oxLDL (50 μg/mL) alone (2), oxLDL plus 2.5 μg/mL Tan IIA (3), oxLDL plus 10 μg/mL Tan IIA (4) and oxLDL plus Cathepsin B inhibitor CA-074-Me (5). The Cathepsin B-/- mouse macrophages (6) were treated with oxLDL alone. The small interfering RNA-mediated CD36 silenced-mouse B6 macrophages (7) were treated with oxLDL alone. All the cells were stained with Alexa 647-conjugated cholera toxin B (membrane staining, red), and then with DQ ovalbumin (cathepsin B staining, green) and Hoechst dye (nuclei staining, blue) after saponin treatment. (B) Western blot detection for Cathepsin B releases from cytosol of oxLDL-induced B6 macrophages or cathepsin B-/- B6 macrophages treated with digitonin and phenylmethylsulfonyl fluoride (PMSF). (C) ELISA assay for oxLDL-induced Il-1β releases from mouse B6 macrophages or cathepsin B-/- B6 macrophages under the different treatments.