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. 2017 Sep 19;8(46):80709–80721. doi: 10.18632/oncotarget.21080

Figure 2. Forced expression of miR-23a induces autophagic activity.

Figure 2

(A) MCF-7 and T47D cells were transfected with miR-24 mimics, miR-7 mimics, miR-23a mimics and miR-513a-5p mimics. Forty-eight hours later, LC3-II/I proteins were detected by Western blot. (B) EBSS induced miR-23a expression strongly in MCF-7 and T47D cells. Cells were treated with EBSS and compared cells grown in with normal medium. Shown is the qRT-PCR analysis for miR-24, miR-7, miR-513a-5p and miR-23a. U6 snRNA was used as an input control (*, P < 0.05). (C) XIAP mRNA expression in seven human mammary cell lines was analyzed by qRT-PCR. (D) Expression of miR-23a induces LC3 conversion and SQSTM1/P62 degradation. T47D cells were transiently transfected with miR-23a mimics and MCF-7 cells were transiently transfected with miR-23a ASO. Total cellular protein was isolated and subjected to Western blot analysis. GAPDH was used as input control. (E) Expression of miR-23a did not significantly induce LC3 conversion and SQSTM1/P62 degradation. MCF-10A cells were transiently transfected with miR-23a mimics and miR-23a ASO. Total cellular protein was isolated and subjected to Western blot analysis. GAPDH was used as input control. (F) GFP-LC3 puncta formation was analyzed by fluorescence microscopy (200× magnification). Black arrows indicate clusters of GFP-LC3 puncta in cells. Quantification of GFP-LC3 puncta in E (mean ± S.D of independent experiments, n = 3,*P < 0.05). (G) Autophagy was evaluated in breast cancer cells by electron microscopy. Scale bars, 200 nm.