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. 2015 Jul 1;6(19):16926–16938. doi: 10.18632/oncotarget.4700

Figure 4. RNA-mediated silencing of Itch impairs lung CSCs proliferation.

Figure 4

A. Western blot analysis of Itch expression in lung CSC LC1 transfected with short-hairpin RNAs non-targeting (Scrambled) or direct against Itch (sh Itch). B. Colony formation in soft-agar culture of lung CSC LC1 transfected as above. Bars represent mean ± S.D.; **P-value <0.01 (n = 3). C. Size of colonies formed in soft-agar assay by lung CSC LC1 transfected as above. Bars represent mean ± S.D.; *P-value <0.05 and **P-value <0.01 (n = 3). D. Viability of lung CSC LC1 transfected as above and treated with 50 μM gemcitabine for 96 h. Bars represent mean ± S.D.; *P-value <0.05 and **P-value <0.01 (n = 3). E. Fold variation of ALDEFLUOR-positive cells in treated lung CSCs with 50 μM gemcitabine for 96 h. Bars represent mean ± S.D.; **P-value <0.01. F. Western blot analysis of lung CSC LC1 transfected as above and treated with 50 μM gemcitabine for 96 h. All whole cell extracts were analyzed by IB using antibodies to the indicated proteins. G. GEO lung adenocarcinoma data set (GSE31210 and GSE11969) were analyzed for the expression of Itch with computation estimation of Kaplan-Maier. Green line represents patients displaying high Itch expression levels while red line represents those with low expression. The R statistical package was used to perform survival analyses and to draw the KM plots.