Figure 3.

miR-199a-3p Affected MTOR-Associated Pathway In Vivo

(A) Immunoblotting analyses of hepatocellular carcinoma (HCC) tissues treated or untreated with miR-199a-3p were performed to investigate the molecular effects of miR-199a-3p on its target genes. We found that proteins involved in the MTOR pathway (phospho-AKT, phospho-forkhead box O3 [pFOXO3], and FOXM1) were downregulated in tumors treated with miR-199a-3p. Phosphatase and tensin homolog (PTEN) protein levels were higher in treated tumors than controls. Glyceraldehyde-3-phosphate dehydrogenase (GAPDH) was used as an internal normalizer. (B) A hierarchical cluster analysis of differentially expressed genes in HCC and normal liver samples (NL) (experiment described in Figure 1D) was performed. Specifically, HCC of scrambled-treated mice, HCC of miR-199a-3p mice, and NL of untreated mice were compared. The analysis showed that several genes controlled by FOXM1 (Ccnb1, Cdc25b, Cdc20, cyclin-dependent kinase 1 [Cdk1], transforming growth factor-beta 2 [Tgfb2], survivin, and Foxm1 itself) were downregulated in miR-199a-3p-treated samples. The colors of the genes represent the expression values normalized using the mean expression across all samples (green, downregulated; red, upregulated). (B) miR-199a-3p is indicated as miR-199.