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. 2019 Dec 19;10:1427. doi: 10.3389/fphar.2019.01427

Figure 7.

Figure 7

Bavachin suppresses β-GP-induced calcification and apoptosis via induction of autophagy. Effects of autophagy inhibitor WM in β-GP/bavachin-treated HASMCs. (A) A representative plate view of VONKOSSA staining was shown after application of WM in β-GP/bavachin-treated HASMCs. The black spots were observed in cytoplasm and nuclear regions of cells. (B) Immunofluorescence staining of calcification and autophagy-related proteins. Green GFP signal represents the calcified protein OPN, whereas red TRITC signal represents the autophagy protein LC3, and blue DAPI signal represents the nuclear region (scalebar = 25 μm). (C) Western blot analysis of autophagy proteins (LC3 and beclin1) in HASMCs treated with 10 mM β-GP and 25 μM bavachin for 72 h (**P < 0.01, ***P < 0.001). The full-length images of Western blot are shown in Figure S7C . (D) Western blot analysis of apoptotic proteins (Caspase3, Caspase9, Bak, Bcl-2, and Bax). The full-length images of Western blot are shown in Figures S1A, S7D . (E) Western blot analysis of calcification-related proteins: Wnt3a, β-catenin, OPN, BMP2, and Runx2. The full-length images of Western blot are shown in Figures 1B , S7E . Statistical significance was analyzed by the one-way ANOVA (**P < 0.01, ***P < 0.001). The data is represented as mean ± S.D. (n = 3).