Fig 1: CircORC2 and TRIM2 were upregulated, but miR‐485‐3p was downregulated in OS cells. (A). Quantitative reverse transcriptase‐polymerase chain reaction (qRT‐PCR) was used to detect the circORC2 level in OS (Saos‐2, SW1353, U‐2Os, SJSA‐1, and HOS) and HFOB1.19 cells. (B). miR‐485‐3p level in OS cells assessed by qRT‐PCR. (C). QRT‐PCR was used to detect the TRIM2 mRNA levels in OS cells. (D). Western blot was employed to measure the TRIM2 protein level in OS cells. U‐2Os and SJSA‐1 cells were exposed to DDP at various concentrations (0, 1, 2, 4, and 8 μM). (E). MTS assay was used to detect the cell viability. (F). Cell proliferation was assessed using the colony formation assay. (G). Flow cytometry was used to detect cell apoptosis. Data are presented as mean ± SD of three replicate experiments (n = 3), *p < 0.05, **p < 0.01 and ***p < 0. 001.
Fig 2: CircORC2 promoted OS cell proliferation and DPP resistance by regulating the miR‐485‐3p/TRIM2 axis. OS cells were transfected with oe‐TRIM2, oe‐TRIM2 with miR‐485‐3p mimics, and/or oe‐circORC2 and then administrated with DDP. (A). Quantitative reverse transcriptase‐polymerase chain reaction was used to detect the TRIM2 level. (B). MTS assay was performed to assess cell viability. (C). Colony formation assay was used to evaluate cell proliferation. (D). Cell apoptosis was detected by flow cytometry. Data are presented as mean ± SD of three replicate experiments (n = 3), *p < 0.05, **p < 0.01, and ***p < 0. 001.
Fig 3: CircORC2 promoted OS cells proliferation and DDP resistance. (A). Quantitative reverse transcriptase‐polymerase chain reaction (qRT‐PCR) detected circORC2, miR‐485‐3p, and TRIM2 levels in the U‐2Os, U‐2Os‐DDP, SJSA‐1, and SJSA‐1‐DDP cells. (B). U‐2Os, U‐2Os‐DDP, SJSA‐1, and SJSA‐1‐DDP cells were transfected with oe‐circORC2 or sh‐circORC2, and qRT‐PCR was used to estimate the circORC2 expression levels in these cells. Subsequently, the cells were exposed to DDP (concentration, 4 μM). (C). The MTS assay was performed to assess the cell viability. (D). Cell proliferation was evaluated using the colony formation assay. (E). Flow cytometry was employed to assess cell apoptosis. Data are presented as mean ± SD of three replicate experiments (n = 3), *p < 0.05, **p < 0.01 and ***p < 0. 001.
Fig 4: TRIM2 is the target gene of miR-369-3p. (A) Binding sites between miR-369-3p and TRIM2. (B) Luciferase reporter assay for the confirmation of direct binding relationship between miR-369-3p and TRIM2. ***P<0.001 vs. mimic-NC + TRIM2 WT group. (C) The mRNA expression of TRIM2 in A549 cells transfected with miR-369-3p mimic was detected by RT-qPCR. (D) The protein expression of TRIM2 in A549 cells transfected with miR-369-3p mimic was detected by western blot analysis. **P<0.01 and ***P<0.001 vs. control group; ##P<0.01 and ###P<0.001 vs. mimic-NC group.
Fig 5: Knockdown of DLEU2 attenuates BLM-induced pulmonary fibrosis in mice. (A) The pathological changes, pulmonary fibrosis and collagen I expression in lung tissues were in turn detected by H&E staining, Masson's staining and immunohistochemistry. (B) The expression of collagen I, α-SMA and E-cadherin in lung tissues of mice with BLM-induced fibrosis following transfection was detected by western blot analysis. (C) DLEU2 expression in lung tissues of mice with BLM-induced fibrosis following transfection was detected by RT-qPCR. (D) The mRNA expression of TRIM2 and PTGFRN in lung tissues of mice with BLM-induced fibrosis following transfection was detected by RT-qPCR. (E) The protein expression of TRIM2 and PTGFRN in lung tissues of mice with BLM-induced fibrosis following transfection was detected by western blot analysis. (F) miR-369-3p expression in lung tissues of mice with BLM-induced fibrosis following transfection was detected by RT-qPCR. *P<0.05, **P<0.01 and ***P<0.001 vs. control group; #P<0.05, ##P<0.01 and ###P<0.001 vs. BLM group; $P<0.05, $$P<0.01 and $$$P<0.001 vs. BLM + shRNA-NC group. BLM, bleomycin; TGF-β1, transforming growth factor β1; α-SMA, α-smooth muscle actin; TRIM2, tripartite motif containing 2; PTGFRN, prostaglandin F2 receptor inhibitor.
Supplier Page from Abcam for Anti-TRIM2 antibody