Fig 1: MiR-373-3p mimics alleviate the effects of lncRNA-LET overexpression on cell cycle and apoptosis. a Western blot analysis was used to measure the DKK1 and TIMP2 protein levels. Cell cycle phase distribution (b) and cell apoptosis (c) were analyzed through flow cytometry. ##p < 0.01, ###p < 0.001. ns, ccRCC clear cell renal cell carcinoma, mimics NC negative control mimics
Fig 2: LncRNA-LET is involved in cell growth of ccRCC cells by targeting miR-373-3p. a MiR-373-3p level was assayed in ccRCC cells with miR-373-3p mimics or inhibitor via quantitative real-time PCR. b LncRNA-LET level was assayed in miR-373-3p mimics or inhibitor ccRCC cells via quantitative real-time PCR. c Schematic diagram: lncRNA-LET (black) function as a target of miR-373-3p (green). MiR-373-3p was indicated to bind with DKK1 (red) and TIMP2 (burgandy). ORF was filled with rectangles. d The binding sites between lncRNA-LET and miR-373-3p, miR-373-3p and DKK1, miR-373-3p and TIMP2 were predicted. e The relationship between lncRNA-LET and miR-373-3p was demonstrated through luciferase reporter assay. f DKK1 and TIMP2 protein levels were examined with western blot analysis. ##p < 0.01, ###p < 0.001. ccRCC clear cell renal cell carcinoma, DKK1 Dickkopf-1, TIMP2 tissue inhibitor of metalloproteinase-2, mimics NC negative control mimics
Fig 3: ZBTB38 suppresses prostate cancer cell proliferation and migration through promoting DKK1 expression.A RT-qPCR showed the mRNA levels of DKK1 gene in the DU145 cells with a control vector or DKK1 knockdown SiRNA. B DKK1 knockdown rescued ZBTB38-mediated reduced cell migration. One-way ANOVA analysis was used. C DKK1 knockdown rescued ZBTB38-mediated reduced cell proliferation (**P < 0.01, ***P < 0.001). D Expression correlation analyses of ZBTB38 and DKK1 mRNA levels in prostate cancer samples from GSE35988.
Fig 4: BPI-9016M increases miR203 and decreases DKK1. (A) miR103, miR203, miR200c, miR30b and miR30c expression levels by qPCR in A549 cells treated with BPI-9016M. U6 was used as the internal control. (B) Wound healing analyses of spreading miR203 inhibitor-transfected A549 and 1299 cells with BPI-9016M treatment. (C) Transwell assays to assess cell migration and invasion in miR203 inhibitor-transfected A549 and H1299 cells with or without BPI-9016M treatment. (D) Western blotting analyses demonstrating DKK1 expression in A549 and H1299 cells with miR103 or mi203 mimics (50 nM) and miR103 or miR203 inhibitors (50 nM). (E) The binding site of miR203 in the DKK1 3'-UTR is shown. 3'-UTR wild-type; 3'-UTR mutation and deletion sequences of the DKK1 3'-UTR. (F) Firefly luciferase activity between miR203 and wild-type, mutated, and deleted constructs of DKK1 3'-UTR. (G) Western blotting analysis showing DKK1 expression in A549 and H1299 cells with or without BPI-9016M/miR203 inhibitor. (H) A schematic illustration showing that BPI-9016M-inhibited proliferation, migration and invasion of lung adenocarcinoma is associated with ERK activation and miR203-DKK1-AKT signaling. (B-C, F) * p<0.05, ** p<0.001, *** p<0.0001.
Fig 5: Clinical significance of BPI-9016M targeted to c-Met, DKK1 and miR203. (A-B) QPCR analyses showing DKK1 (A) and miR203 (B) expressions in the four cases of PDX tissues of lung adenocarcinoma. (C-E) Correlations between c-Met and DKK1 (C), c-Met and miR203 (D) and DKK1 and miR203 (E). (F-G) Kaplan-Meier plots showing overall survival of lung adenocarcinoma patients with high or low expression of c-Met (G) and DKK1 (H) from the database (http://www.kmplot.com). (H-J) Kaplan-Meier plots showing overall survival of lung adenocarcinoma patients from Peking University Cancer Hospital & Institute with high or low expression of c-Met (H), DKK1 (I) and miR203 (J). (K) Tumor cell growth inhibition (%) of DKK1 siRNA-transfected cells treated with variable doses of BPI-9016M. (L) Tumor cell growth inhibition (%) of miR203 mimics-transfected cells treated with variable doses of BPI-9016M. (K-L) n.s.: no statistically significant difference; *p<0.05, ** p<0.001, *** p<0.0001; HR: hazard ratio.
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