Fig 2: miR-340-5p downregulates FHL2 expression in colon cancer cells. FHL2 expression was evaluated using (A and B) RT-qPCR and (C-F) confocal microscopy (scale bars, 10 µm). Cells were transfected with miR-340-5p mimic, mimic control, TSB control and TSB for 24 h in low-serum conditions. Colon cancer cells were then stimulated using 10% FBS for 30 min. Relative expressions of FHL2 mRNA was quantified using RT-qPCR where β-actin was used as an internal control for FHL2 mRNA. Expressions were determined using 2−ΔΔCq method. Data are presented as the mean ± SEM (n=4). miR, microRNA; FHL2, four and a half LIM domains protein 2; RT-qPCR, reverse transcription-quantitative PCR; miR, microRNA; Ctrl, control.

Fig 3: Effect of short‐ and long‐term treatments with CoCl2 and PM on VEGFA, FHL2 and HIF1A‐AS2 RNA accumulation. (A) Depletions of glucose, pyruvate and glutamine, in conjunction or not with long‐term treatment with CoCl2. (B) Short‐term treatment with CoCl2 with or without PM. Columns with different superscripts differ significantly (p < .05).

Fig 4: Expression of miR-340-5p and FHL2 in HT-29 and AZ-97 colon cancer cell lines. Expression of (A) miR-340-5p and (B) FHL2 mRNA in low-serum and serum-grown HT-29 and AZ-97 cells. Relative expressions were determined using reverse transcription-quantitative PCR with U6 as a control for miR-340-5p and β-actin as a control for FHL2 mRNA. Relative expressions were determined using the 2−ΔΔCq method. Data are presented as the mean ± SEM (n=4). miR, microRNA; FHL2, four and a half LIM domains protein 2.

Fig 5: Clinical associations of miR-340 with FHL2 expression in human OC tissues.a Levels of total β-catenin, phosphorylated β-catenin (Ser33/37/Thr41), cyclin D1, p53, PUMA, E-cadherin, and N-cadherin in miR-340-overexpressing tumor samples from nude mice by immunoblot analysis. b miR-340 was significantly associated with FHL2 in 481 ovarian serous cystadenocarcinoma tissues by CancerMiner. c FHL2 expression was significantly upregulated in OC tissues with lower miR-340 levels. Means ± SEM are shown. *P < 0.05 by Student’s t-test. d, e Kaplan–Meier survival curves for OC patients according to miR-340 (d) and FHL2 (e) expression levels in tumor tissues; significance was calculated using log-rank test. f Schematic diagram of roles of miR-340-FHL2 axis on Wnt/β-catenin signaling and its function in ovarian tumorigenesis