miR-509-5p inhibits cellular proliferation and migration via targeting MDM2 in pancreatic cancer cells

Figures & data

Figure 1 Real-time quantitative reverse transcription polymerase chain reaction was performed to examine the expression of miR-509-5p in pancreatic cancer tissues and noncancerous adjacent tissues (n=10).

Notes: The results are expressed relative to the inner control value. *P<0.05 vs adjacent normal tissues.

Figure 2 Inhibition of pancreatic cancer PANC-1 cell proliferation, migration, and invasion by miR-509-5p.

Notes: (A) Expression levels of miR-509-5p were examined by real-time quantitative reverse transcription polymerase chain reaction in PANC-1 cells transfected with miR-509-5p mimic, inhibitor, or scramble, or with no transfection. (B) PANC-1 cells transfected with miR-509-5p mimic, inhibitor, or scramble control were measured by CCK-8 assay. The data are expressed as the mean ± SD of sextuplicate results per treatment group of one representative experiment. (C) Transwell analysis of the migration of PANC-1 cells after treatment with miRNA mimic, inhibitor, or scramble, or with no transfection (400× magnification). (D) Transwell analysis of invasion of PANC-1 cells after treatment with miRNA mimic, inhibitor, or scramble, or with no transfection (400× magnification). Representative images (left panel) from each group are shown. The right panel depicts the quantitative data from at least three random fields of view. *P<0.05, ^#P<0.05 (mean ± SD) vs scramble control.

Figure 3 miR-509-5p directly targeted MDM2 to exert its effect.

Notes: (A) Sequences of miR-509-5p binding sites within the human MDM2 3′UTR and schematic reporter constructs. (B) Analysis of the relative luciferase activities of MDM2-WT and MDM2-Mut in 293T cells transfected with miR-509-5p mimic or scramble control. *P<0.05 (mean ± SD) vs scramble control. (C) Real-time quantitative reverse transcription polymerase chain reaction analysis of MDM2 mRNA expression in PANC-1 cells transfected with miR-509-5p mimic or scramble control. The results are expressed relative to the control value. *P<0.05 (mean ± SD) vs scramble control.

Figure 4 miR-509-5p reversed MDM2 overexpression-induced increase in pancreatic cancer PANC-1 cell proliferation and migration.

Notes: (A) Western blot analysis of MDM2 in PANC-1 cells transfected with miR-509-5p mimic, inhibitor, or scramble or co-transfected with either pc-DNA vector control or pcDNA-MDM2, with or without miR-509-5p mimic. GAPDH was used as internal control. (B) PANC-1 cells cotransfected with the target gene and miR-509-5p mimic or scramble control were measured by CCK-8 assay. The data are expressed as the mean ± SD of sextuplicate results per treatment group of one representative experiment. (C) Transwell analysis of migration of Panc-1 cells treated with different combinations. Representative images from each group are shown. (D) The quantitative data from at least three random fields of view. *P<0.05 (mean ± SD) vs pc-DNA vector control, ^#P<0.05 (mean ± SD) vs scramble control.

Figure 5 Inhibition of tumor growth by miR-509-5p in a xenograft mouse model.

Notes: Mean tumor volumes (A), mean tumor weights (B), and representative xenograft tumor images (C) from miR-509-5p-expressing PANC-1 cells and control group are shown. (D) Tumor immunohistochemical (IHC) staining specific for MDM2. Hematoxylin and eosin (HE) staining indicates the area containing necrotic cells, cell debris, or cells undergoing cell necrosis. *P<0.05 (mean ± SD, n=5) vs scramble control group.

Figure S1 Real-time quantitative polymerase chain reaction was performed to examine the expression of miR-509-5p in the isolated islet cells from pancreas (n=3) and three pancreatic cancer cell lines.

Notes: Results are expressed relative to the control value. *P<0.05 vs islet cells.