Figures & data
Figure 1 The expression of miR-375 and TINCR in clinical GC tissues.
Notes: (A) miR-375 expression was examined by real-time PCR in 56 paired human GC tissues and adjacent noncancerous tissues; the normalized to U6 expression. (B) TINCR expression was examined by real-time PCR in 56 paired human GC tissues and adjacent noncancerous tissues; the normalized to GAPDH expression. (C) The correlation between miR-375 and TINCR expressions in GC cases was evaluated using Spearman’s correlation analysis; n=56; P<0.01. (D) miR-375 and TINCR expressions were examined by real-time PCR in human gastric cancer KATO III, NCI-N87, HGC-27, and SNU-1 cell lines. (E) miR-375 expression was examined by real-time PCR in HGC-27 and SNU-1 cells with TINCR-siRNA and pcDNA3.1-TINCR. Statistically significant differences are indicated: *P<0.05; Student’s t-test. The experiment was repeated at least three times.
Abbreviations: GAPDH, glyceraldehyde-3-phosphate dehydrogenase; GC, gastric cancer; PCR, polymerase chain reaction.
Abbreviations: GAPDH, glyceraldehyde-3-phosphate dehydrogenase; GC, gastric cancer; PCR, polymerase chain reaction.
Figure 2 TINCR was a target of miR-375 in GC cells.
Notes: (A) miR-375-binding sites on TINCR predicted by online soft RNAhybrid. (B) Relative expression of TINCR after transfecting miR-375 mimic into HGC-27 and SNU-1 cells was analyzed by using real-time PCR assays and normalized to GAPDH expression. (C and D) Luciferase activity of the indicated group in HGC-27 and SNU-1 cells by transfected miR-375 mimic + pmirGLO vector, miR-375 mimic + pmirGLO-TINCR-wt, or miR-375 mimic + pmirGLO-TINCR-mut. Statistically significant differences are indicated: *P<0.05; Student’s t-test. The experiment was repeated at least three times.
Abbreviations: GAPDH, glyceraldehyde-3-phosphate dehydrogenase; mut, mutant; GC, gastric cancer; PCR, polymerase chain reaction; wt, wild type.
Abbreviations: GAPDH, glyceraldehyde-3-phosphate dehydrogenase; mut, mutant; GC, gastric cancer; PCR, polymerase chain reaction; wt, wild type.
Figure 3 TINCR promoted the PDK1 expression by sponging miR-375.
Notes: (A) Relative mRNA levels of PDK1 were analyzed by using real-time PCR analysis by transfecting negative control, siTINCR, and siTINCR + miR-375 inhibitor in HGC-27 and SNU-1 cells. (B) Relative protein expression levels of PDK1 were analyzed by Western blot analysis by transfecting negative control, siTINCR, and siTINCR + miR-375 inhibitor in HGC-27 and SNU-1 cells. Statistically significant differences are indicated: *P<0.05; Student’s t-test. The experiment was repeated at least three times.
Abbreviations: GAPDH, glyceraldehyde-3-phosphate dehydrogenase; PCR, polymerase chain reaction.
Abbreviations: GAPDH, glyceraldehyde-3-phosphate dehydrogenase; PCR, polymerase chain reaction.
Figure 4 TINCR inhibited apoptosis and promoted cell proliferation by sponging miR-375.
Notes: (A) Flow cytometry analysis was performed using cells stained with Annexin V-FITC and PI after transfecting negative control, siTINCR, and siTINCR + miR-375 inhibitor in HGC-27 and SNU-1 cells. (B) Flow cytometry analysis was performed using cells stained with Annexin V-FITC and PI after transfecting negative control, mimic, and mimic + TINCR in HGC-27 and SNU-1 cells. (C) CCK-8 cell proliferation assays were analyzed by transfecting negative control, siTINCR, and siTINCR + miR-375 inhibitor in HGC-27 and SNU-1 cells. (D) CCK-8 cell proliferation assays were analyzed by transfecting negative control, mimic, and mimic + TINCR in HGC-27 and SNU-1 cells. Statistically significant differences are indicated: *P<0.05; Student’s t-test. The experiment was repeated at least three times.
Abbreviations: CCK-8, Cell Counting Kit-8; FITC, fluorescein isothiocyanate; PI, propidium iodide.
Abbreviations: CCK-8, Cell Counting Kit-8; FITC, fluorescein isothiocyanate; PI, propidium iodide.
Figure 5 Tumor growth suppression was retarded with miR-375 downregulated in TINCR knockdown of gastric cancer cell xenografts.
Notes: (A) HGC-27 cells were transfected with negative control, siTINCR, or siTINCR + miR-375 inhibitor and injected subcutaneously into 10 nude mice per flank. Surgical resections of HGC-27 xenograft tumors on week 4 for animals were shown. (B) Measurements of tumor volumes were taken weekly, and tumor volumes were shown. Statistically significant differences are indicated: *P<0.05; Student’s t-test. (C) Schematic model of the TINCR functions as a competing endogenous RNA to regulate PDK1 expression by sponging miR-375 in gastric cancer.
