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Fig. 2 | Journal of Experimental & Clinical Cancer Research

Fig. 2

From: Inhibition of Stearoyl-CoA desaturase 1 reverts BRAF and MEK inhibition-induced selection of cancer stem cells in BRAF-mutated melanoma

Fig. 2

Higher SCD1 expression correlates with increased stemness and drug resistance. a) Evaluation of proliferation performed on A375, M14, M14-R, WM115 by MTT assay after 72 h of vemurafenib exposure; b) MITF and SCD1 protein expression examined in BRAF-mutated melanoma stable cell lines (A375, M14, M14-R, WM115) grown in adhesion by WB; c) qRT-PCR analyses performed on A375, M14, M14-R and WM115 in basal condition; d) Western blotting analysis of SCD1 in A375, LOXIMV1, M14 and WM266 sensitive and their counterpart resistant to vemurafenib; e-f) Western blotting performed on A375 transfected with empty or SCD1 vector in adherent conditions and treated or not with MAPKi combination (50 + 50 nM) for 48 h. Densitometric analyses relative to SCD1, pERK and YAP/TAZ protein levels were expressed as a fold-change relative to empty (CTRL); g) Evaluation of BRAFi/MEKi effects on proliferation performed on 2D A375-transfected with empty or SCD1 vector and treated for 72 h. The dose–effect curves shows a decreased sensitivity of A375 SCD1-overpressing compared to this of A375-transfected with empty vector; h) Stemness gene expression in stable (left panel) and primary (right panel) cell lines by qRT-PCR; i) Representative images of melanoma cell lines grown in 2D and 3D condition taken on day 4. Scale bars: 50 μM (3D) and 100 μM (2D); j) Gene expression of SCD1 performed on 3D and 2D cultures by qRT-PCR analyses. The results indicate an enrichment of SCD1 mRNA expression in 3D spheroids compared with 2D parental cells; k) Western blotting analysis of SCD1 protein in a large panel of melanoma stable and primary cell lines grown in 3D and 2D conditions; l) MUFA levels in A375 and M14 cell lines analysed by GS/MS in 2D and 3D cultures. Data represent the means and SD of 3 independent experiments and are statistically significant if *p < 0.05 (ANOVA test)

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