Quaking 5 suppresses TGF-β-induced EMT and cell invasion in lung adenocarcinoma (2021)
Figure 5

Figure 5. QKI-5 suppresses TGF-β-stimulated LUAD cell metastasis in vivo.

(A) Schematic flowchart of LUAD cell in vivo metastasis model. QKI-5-overexpression and control vector A549 cells (3 × 106 cells/mouse) were i.v. injected into BALB/c nude mice (10 mice per group). TGF-β1 was injected i.p. at day 1, 6, 11, 16 after cell injection to achieve TGF-β-stimulated A549 cell metastasis.

(B) Before cell injection, QKI-5 was confirmed to overexpress in A549 cells using qRT-PCR and western blot. Data are shown as the mean ± SD of n = 3 technical replicates. ***P < 0.001 by unpaired Student's t-test.

(C) Representative images of lung metastatic nodules developed in mice 8 weeks after injection of QKI-5-overexpressing A549 cells or control A549 cells. The surgically resected lungs were stained as described in Materials and Methods. Red arrowheads indicate metastatic nodules established in lungs. Scale bar, 5 mm.

(D) Comparison of the number of lung metastatic nodules between QKI-5-overexpressing group and vector group (n = 10 mice per group). Data are shown as the boxplot. Central band: median; Upper and lower line of boxes: upper and lower quartile; Upper and lower line of whiskers: maximum and minimum. Data are shown as the mean ± SEM. **P < 0.01 by unpaired Student's t-test.

(E) H&E staining was performed for the evaluation of lung micrometastases. Representative images showing micrometastases of lung tissues from a pair of mice referred in Fig 5C. Blue and red arrowheads indicate lung micrometastases of vector group and QKI-5-overexpressing group, respectively. Scale bar: 100 μm.

(F) Dot plots showing the difference of the micrometastases counts in lung tissues between QKI-5-overexpressing group and vector control group (n = 10 mice per group). Data are shown as the mean ± SEM. *P < 0.05 by unpaired Student's t-test.

(G) Representative microscopic photographs of H&E staining for liver micrometastases in a pair of mice referred in Fig 5E. Blue and red arrowheads indicate liver micrometastases. Scale bar: 100 μm.

(H) Dot plots showing the difference of the micrometastases counts in liver tissues between QKI-5-overexpressing group and control vector group (n = 10 mice per group). Data are shown as the mean ± SEM. *P < 0.05 by unpaired Student's t-test.

(I) Left part: Photos of murine lungs with metastatic nodules after inoculation of sh-QKI-5-2 or sh-NC A549 cells. Scale bar, 5 mm. Right part: representative microscopic images of H&E staining for micrometastatic foci established in lungs from a pair of mice referred in left part. Scale bar, 1 mm. Red arrowheads indicate metastatic nodules and micrometastatic foci in lungs.

(J and K) Dot plots showing the difference of the metastatic nodules (J) and micrometastases counts (K) in lung tissues between QKI-5-silenced group and control group (n = 5 mice per group). In the boxplot, central band represents median, upper and lower line of boxes represents upper and lower quartile, upper and lower line of whiskers represents maximum and minimum (J). Data are shown as the mean ± SEM. ***P < 0.001 by unpaired Student's t-test.