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Figures
- Fig. 1.
The levels of miR-96-5p in OSCC tissues and cell lines. (A) Quantitative RT-PCR analysis of miR-96-5p level in OSCC tissues and adjacent normal tissues (n=40). Transcript levels were normalized to U6 level. (B) Relative miR-96-5p level analyzed via quantitative RT-PCR in five OSCC cell lines normalized to U6 (n=6). All data are presented as means±s.e.m. *P<0.05, **P<0.01, ***P<0.001 versus normal tissues or NHOK.
- Fig. 2.
The effects of miR-96-5p on proliferation and related molecules in OSCC cells. Tca8113 and Cal-27 cells were transfected with miR-96-5p mimic or inhibitor for 48 h. (A) The level of miR-96-5p was detected by quantitative RT-PCR. (B) Cell proliferation was assessed by a BrdU-ELISA assay. (C) The mRNA expression of CDK4, cyclin D1 and p27 were determined by quantitative RT-PCR. All data are presented as means±s.e.m., n=6. #P<0.05, ##P<0.01, ###P<0.001 versus NC.
- Fig. 3.
The effects of miR-96-5p on the invasion and related molecules expression in OSCC cells. Tca8113 and Cal-27 cells were transfected with miR-96-5p mimic or inhibitor for 48 h. (A) The invasion was assessed by Transwell assay. (B) Total secretions of MMP-2, MMP-9 and TIMP-1 in the culture supernatants were detected by ELISA assays. (C) The mRNA expression of MMP-2, MMP-9 and TIMP-1 were examined by qRT-PCR. All data are presented as means±s.e.m., n=6. #P<0.05, ##P<0.01 versus NC.
- Fig. 4.
Effects of miR-96-5p on the EMT of OSCC cells. Tca8113 and Cal-27 cells were transfected with miR-96-5p mimic or inhibitor for 48 h. The expression of E-cadherin, Vimentin and N-cadherin were detected by western blot assays. All data are presented as mean±s.e.m., n=6. #P<0.05, ##P<0.01 versus NC.
- Fig. 5.
FOXF2 is a direct target of miR-96-5p. Tca8113 and Cal-27 cells were transfected with miR-96-5p mimic or inhibitor for 48 h. (A) Schematic representation of FOXF2 3′UTRs showing putative miRNA target site. (B) The analysis of the relative luciferase activities of FOXF2-WT and FOXF2-MUT. (C) The protein expression of FOXF2 were determined by western blot assay. (D) Quantitative RT-PCR analysis of FOXF2 expression in OSCC tissues (n=40) and adjacent normal tissues (n=40). Transcript levels were normalized to GAPDH expression. (E) Relative FOXF2 expression analyzed via quantitative RT-PCR in five OSCC cell lines normalized to GAPDH (n=6). (F) Pearson's correlation analysis of the relative expression levels of miR-96-5p and the relative FOXF2 mRNA levels in OSCC tissues. All data are presented as means±s.e.m., n=6. ##P<0.01 versus NC; *P<0.05, **P<0.01, ***P<0.001 versus normal tissues or NHOK.
- Fig. 6.
The effects of FOXF2 knockdown on the proliferation, invasion and EMT in OSCC cells. Tca8113 and Cal-27 cells were transfected with si-NC or si-FOXF2 for 48 h. (A) The protein expression of FOXF2 was determined by western blot. (B) Cell proliferation was assessed by a BrdU-ELISA assay. (C) The invasion of OSCC cells was assessed by Transwell assay. (D) Total secretions of MMP-2, MMP-9 and TIMP-1 in the culture supernatants were detected by ELISA assays. (E) The expression of E-cadherin, Vimentin and N-cadherin were detected by western blot assays. All data are presented as means±s.e.m., n=6. #P<0.05, ##P<0.01 versus si-NC.
- Fig. 7.
Introduction of FOXF2 promoted cell proliferation and invasion in miR-96-5p-overexpressing OSCC cells. Tca8113 cells were co-transfected with pcDNA3.1 or pcDNA-FOXF2 with or without miR-96-5p mimic. (A) The protein expression of FOXF2 was determined by western blot assay. (B) Cell proliferation was assessed by a BrdU-ELISA assay. (C) The expression of CDK4, cyclin D1 and p27 were determined by quantitative RT-PCR, respectively. (D) The invasion of OSCC cells was assessed by Transwell assay. (E) Total secretions of MMP-2, MMP-9 and TIMP-1 in the culture supernatants were detected by ELISA assays. (F) The expression of E-cadherin, Vimentin and N-cadherin were detected by western blot assays. All data are presented as means±s.e.m., n=6. *P<0.05, **P<0.01 versus NC; #P<0.05, ##P<0.01 versus miR-96-5p mimic+pcDNA3.1.
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