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. 2019 Apr 12;43:270–281. doi: 10.1016/j.ebiom.2019.04.010

Fig. 5.

Fig. 5

Involvement of autophagy or EMT pathway in the miR-338-5p-related CRC migration in vitro. (a) LC3 puncta were labelled with anti-LC3 antibody and fluorescent isothiocyanate-conjugated goat anti-rabbit IgG (green fluorescence). SW480 cell nuclei were stained with Hoechst 33258 (blue fluorescence). Images were photographed using florescent microscopy (the scale bar represents 10 μm). Data are presented as median (IQR) (n = 5) (P value was analysed using Mann-Whitney tests). (b) SW480 cells were treated with 10 μM of amiodarone for 48 h and then transfected with 100 nM of miR-338-5p or N.C. PIK3C3 and LC3 expression was measured using Western blot. (c) For transwell assay, SW480 cells were first treated with 10 μM of amiodaron and then transfected with/without 100 nM of miR-338-5p. Data are presented as median (IQR) (n = 5) (P value was analysed using Mann-Whitney tests).The scale bar represents 50 μm. (d) After infection with lentivirus carrying miR-338-5p or sh-GFP, HCT116 stable cells were treated with amiodarone (10 μM) for 48 h. Expression of PIK3C3 and LC3 was assayed by Western blot. (e) Transwell invasion assay was performed on HCT116 cells after infection with miR-338-5p or shGFP lentivirus and counted for invaded cells for 48 h. The scale bar represents 50 μm. Data are expressed as means ± SEM (n = 8) (P value was analysed using t tests). (f) SW480 cells were infected with sh-ATG-5 lentivirus and then transfected with 100 nM of miR-338-5p or N.C. Expression of ATG5-ATG12 and LC3 was measured using Western blot. Representative results of transwell assays. The scale bar represents 50 μm. The number of migrated cells are shown as means ± SEM (n = 8) (P values were analysed using t tests). (g) After infection with lentivirus carrying miR-338-5p, western blot was used to analyse the expression of PIK3C3, LC3, p62, Snail, Twist, N-cadherin, E-cadherin, vimentin, fibronectin and β-actin, respectively, in HCT116 cells. (For interpretation of the references to colour in this figure legend, the reader is referred to the web version of this article.)