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. 2022 Apr 27;13:849279. doi: 10.3389/fendo.2022.849279

Figure 4.

Figure 4

Effects of IGF1R localization over phospholipid and triglyceride synthesis. U87Empty, U87WT, and U87Mut cells were cultured 24 h in serum-free medium or with 50-nM IGF1 stimulation (IGF1) or pre-incubated (1 h) with 0.5 μM OSI906 (IGF1+OSI) in the presence of 1-[14C] acetic acid. Lipids were extracted and resolved by thin-layer chromatography, and the radioactivity associated with each condition was quantified. Three independent experiments were performed. (A) 1-[14C] acetic acid incorporated to newly synthesized phospholipids. The results are expressed as percentage of 1-[14C] acetic acid incorporated into phospholipid fraction normalized by the number of cells seeded/well. Values are presented as mean ± SD (*p < 0.05, **p < 0.005, ANOVA, Tukey’s post-test). (B) Changes in the amount of phospholipids in response to 24-h IGF1 stimulation in each cell line. The results are presented as fold change compared to control conditions (serum-free). Values are expressed as mean ± SD (*p < 0.05, Mann–Whitney test). (C) U87Empty, U87WT, and U87Mut cells were cultured 24 h in serum-free medium. CCTα mRNA expression was calculated by rqPCR by the relative quantitation method, and the results are presented as fold change compared to U87Empty cells. Values are expressed as mean ± SD of three independent experiments performed in triplicates (*p < 0.05, ANOVA, Tukey’s post-test). (D) Representative picture of lipid droplets visualized by oil red O (ORO) in U87Empty, U87WT, and U87Mut cells after 24 h of incubation in serum-free medium or with 50-nM IGF1 stimulation (IGF1). (E) DGAT mRNA expression of U87Empty, U87WT 5, and U87Mut 5 cells incubated in serum-free medium (left) or with 50-nM IGF1 stimulation (IGF1) (right). The results are presented as fold change compared to U87Empty cells. Values are expressed as ± SD of three independent experiments performed in triplicates (***p < 0.001, ANOVA, Tukey’s post-test).