Fig 2: miR-564 attenuates the activation of the PTEN/Akt pathway in PTC cells. miR-564 mimics were co-transfected with pc-AEG-1 or pcDNA3.1 into TPC-1 and HTH83 cells. Western blot analysis was conducted at 72 hrs following transfection to detect the protein levels of PTEN, p-Akt, and Akt.

Fig 3: AEG-1 restoration partially reverses the tumor-suppressive effects of miR-564 overexpression in PTC cells. (A) TPC-1 and HTH83 cells were transfected with miR-564 mimics with pc-AEG-1 or pcDNA3.1. Following transfection, AEG-1 expression was measured by Western blot analysis. *P<0.05 compared with miR-NC; #P<0.05 compared with miR-564 mimics+pcDNA3.1. (B and C) MTT and cell apoptosis assays were employed to evaluate cell proliferation and apoptosis of the aforementioned cells. *P<0.05 compared with miR-NC; #P<0.05 compared with miR-564 mimics+pcDNA3.1. (D and E) The migration and invasion in TPC-1 and HTH83 cells treated as described above was investigated using transwell assays. *P<0.05 compared with miR-NC; #P<0.05 compared with miR-564 mimics+pcDNA3.1.

Fig 4: MTDH is a direct target gene of miR-877 in CRC cells. (A) A sequence complementary to that of miR-877 was discovered in the 3'-UTR of MTDH. The mutant miR-877-binding sequence is also shown. (B) miR-877 mimics or miR-NC and pGL3-WT-MTDH-3'-UTR or pGL3-MUT-MTDH-3'-UTR were co-transfected into SW480 and HCT116 cells. After 48 h of transfection, luciferase reporter assay was performed to detect luciferase activity. *P<0.05 versus miR-NC. (C) MTDH mRNA expression in 53 pairs of CRC tissues and adjacent non-tumor tissues was detected through RT-qPCR. *P<0.05 versus non-tumor tissues. (D and E) Western blotting analysis was carried out to measure MTDH protein expression in several pairs of CRC tissues and adjacent non-tumor tissues. N: non-tumor tissues; T: CRC tissues. *P<0.05 versus non-tumor tissues. (F) Spearman’s correlation analysis was applied to evaluate the relationship between miR-877 and MTDH mRNA levels in CRC tissues. r=-0.5381, P<0.0001. (G and H) SW480 and HCT116 cells were transfected with miR-877 or miR-NC. Results of RT-qPCR and western blotting analyses indicated that miR-877 upregulation decreased MTDH mRNA and protein expression. *P<0.05 versus miR-NC.

Fig 5: miR-877 directly targets MTDH to inhibit the activation of the PTEN/Akt signaling pathway in CRC cells. SW480 and HCT116 cells were co-transfected with miR-877 mimics and pc-MTDH or empty pcDNA3.1. After 72 h of transfection, western blotting analysis was performed to measure PTEN, p-Akt, and Akt protein levels.