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. 2020 Mar 27:13:2537-2550.
doi: 10.2147/OTT.S241199. eCollection 2020.

miR-26a-5p Inhibit Gastric Cancer Cell Proliferation and Invasion Through Mediated Wnt5a

Yu Li #  1 , Peng Wang #  1 , Lei-Lei Wu  2 , Jun Yan  1 , Xue-Ying Pang  1 , Song-Jiang Liu  1
Affiliations

miR-26a-5p Inhibit Gastric Cancer Cell Proliferation and Invasion Through Mediated Wnt5a

Yu Li et al. Onco Targets Ther. .

Abstract

Purpose: Gastric cancer (GC) is a malignant disease of digestive tract. Clinically, radiation therapy is widely applied in treating GC, while with undesirable outcome due to tumor re-proliferation and recurrence and metastasis after radiation. Therefore, it is crucial to explore potential molecular mechanisms to develop therapeutic strategies. The present study found that miR-26a-5p has low expression in GC patients and could regulate Wnt5a to inhibit tumor growth, which was a potential therapeutic target for GC. To explore the expression and related mechanism of miR-26a-5p and Wnt5a in GC.

Patients and methods: MiR-26a-5p and Wnt5a were extracted from the transcriptome data of GC downloaded from TCGA database for analysis. The expression levels of miR-26a-5p and Wnt5a in patients' tissues and serum were detected by qRT-PCR, and their correlation with patients' pathological data and survival was analyzed. In addition, miR-26a-5p and Wnt5a overexpression and inhibition vectors were transfected into cells to observe the effects on the proliferation, invasion and apoptosis of GC cells. The relationship between miR-26a-5p and Wnt5a was analyzed by dual luciferase report.

Results: The database and clinical samples showed that miR-26a-5p level was low while Wnt5a was high in GC. MiR-26a-5p level decreased in patients with stage III+IV, lymphatic metastasis and tumor ≥3cm, and Wnt5a was contrary to that of the miR-26a-5p, with diagnostic value. Overexpressed miR-26a-5p and inhibited Wnt5a enhanced apoptosis, decreased proliferation and invasion, reduced Bcl-2 and β-catenin proteins, and elevated Caspase 3, E-cadherin and Bax proteins, while inhibited miR-26a-5p and over-expressed Wnt5a showed the opposite results. Dual luciferase report confirmed that miR-26a-5p targeted to regulate Wnt5a, and rescue experiments found that these effects could be counteracted by reducing miR-26a-5p level.

Conclusion: Overexpressed miR-26a-5p can inhibit Wnt5a expression, promote cell apoptosis, and suppress cell proliferation and invasion in GC.

Keywords: TCGA; Wnt5a; diagnosis; gastric cancer; miR-26a-5p; survival.

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Conflict of interest statement

Yu Li and Peng Wang contributed equally to this study as co-first authors. The authors report no conflicts of interest in this work.

Figures

Figure 1
Figure 1
The expression of miR-26a-5p and Wnt5a in TCGA database. (A) MiR-26a-5p expression in hepatocellular carcinoma; (B) Wnt5a expression in hepatocellular carcinoma. ***Represented P<0.001.
Figure 2
Figure 2
The expression and clinical value of miR-26a-5p and Wnt5a in gastric cancer. (A) Expression and diagnostic value of miR-26a-5p in serum; (B) Expression and diagnostic value of Wnt5a in serum; (C) Expression of miR-26a-5p and Wnt5a in tissues; (D) Correlation of miR-26a-5p and Wnt5a in tissue and serum; (E) Expression and diagnostic value of miR-26a-5p in differentiating tumor size from ≥3 cm to <3 cm; (F) Expression and diagnostic value of miR-26a-5p in differentiating I+II from III+IV patients; (G) Expression and diagnostic value of miR-26a-5p in differentiating lymphatic metastasis; (H) Expression and diagnostic value of Wnt5a in differentiating the size of tumors (> 3cm) from < 3cm; (I) Expression and diagnostic value of Wnt5a in differentiating stage I+II from stage III+IV patients; (J) Wnt5a expression and diagnostic value in differentiating lymphatic metastasis patients. **Represented P<0.01, and ***Represented P<0.001.
Figure 3
Figure 3
The relationship between miR-26a-5p, Wnt5a expressions and the survival of patients with gastric cancer. (A) The five-year survival rate of gastric cancer patients; (B) Comparison of five-year survival of patients with high and low expression of miR-26a-5p; (C) Comparison of five-year survival of patients with high and low expression of Wnt5a.
Figure 4
Figure 4
The expression of miR-26a-5p in gastric cancer cells and its effect on cell biological function. (A) Expression of miR-26a-5p in different groups; (B) Expression of miR-26a-5p in transfected SGC-7901 and NCI-N87 cells; (C) Comparison of proliferation between transfected SGC-7901 and NCI-N87 cells; (D) Comparison of invasive ability between transfected SGC-7901 and NCI-N87 cells; (E) Comparison of apoptosis rate between transfected SGC-7901 and NCI-N87; (F) Comparison of Caspase 3, Bcl-2, Bax, E-cadherin and β-catenin proteins in transfected SGC-7901 and NCI-N87 cells. *Represented the comparison with the miR-NC group, P<0.05. **Represented the comparison with the miR-NC group, P<0.01.
Figure 5
Figure 5
The expression of Wnt5a in gastric cancer cells and its effect on cell biological function. (A) Expression of Wnt5a in all groups of cells; (B) Expression of Wnt5a in transfected SGC-7901 and NCI-N87 cells; (C) Comparison of proliferation between transfected SGC-7901 and NCI-N87 cells; (D) Comparison of invasive ability between transfected SGC-7901 and NCI-N87 cells; (E) Comparison of apoptosis rate between transfected SGC-7901 and NCI-N87; (F) Comparison of Caspase 3, Bcl-2, Bax, E-cadherin and β-catenin proteins in transfected SGC-7901 and NCI-N87 cells. *Represented the comparison with the miR-NC group, P<0.05. **Represented the comparison with the miR-NC group, P<0.01.
Figure 6
Figure 6
Target gene identification of miR-26a-5p. (A) miR-26a-5p binds to Wnt5a and mutates; (B) Dual-Luciferase reporter assay confirms targeted binding of miR-26a-5p to Wnt5a; (C) Expression of Wnt5a protein in SGC-7901 and NCI-N87 cells after transfection of miR-26a-5pmimics and miR-26a-5p-inhibit; (D) Analysis of the correlation between miR-26a-5p and Wnt5a in patients’ serum; (E) Analysis of the correlation between miR-26a-5p and Wnt5a in patients’ tissues. *Represented P<0.05, and **Represented P<0.001.
Figure 7
Figure 7
Rescue experiment. (A) Effects of transfection with miR-26a-5p-mimics+sh-Wnt5a and miR-26a-5p-inhibitor+si-Wnt5a on proliferation of SGC-7901 and NCI-N87 cells; (B) Effects of transfection with miR-26a-5p-mimics+sh-Wnt5a and miR-26a-5p-inhibitor+si-Wnt5a on the invasive function of SGC-7901 and NCI-N87 cells; (C) Effect of transfection with miR-26a-5p-mimics+sh-Wnt5a and miR-26a-5p-inhibitor+si-Wnt5a on the apoptosis rate of SGC-7901 and NCI-N87 cells; (D) Effects of transfection with miR-26a-5p-mimics+sh-Wnt5a and miR-26a-5p-inhibitor+si-Wnt5a on apoptosis proteins of SGC-7901 and NCI-N87 cells. *Represented the comparison with the miR-NC group, P<0.05. **Represented the comparison with the miR-NC group, P<0.01.
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