miR-362-5p targets DKK1 and promotes the proliferation and invasion in tongue squamous cell carcinoma

doi:10.3877/cma.j.issn.1674-1366.2017.02.004

Abstract

Abstract:

Objective

To investigate the expression level of miR-362-5p in tongue squamous cell carcinoma (TSCC) tissues and cell lines, and to explore the effect of miR-362-5p on TSCC cells proliferation and invasion, and their related mechanisms.

Methods

Real-time quantitative PCR was performed to detect the relative expression of miR-362-5p in TSCC tissues and cell lines. SCC-9 and UM1 cells were transfected with miRNA mimic or antagomiR, cell counting kit-8 (CCK-8) assay was used to evaluate cell proliferation, and transwell assay was carried out to assess cell invasion. TOP/FOP luciferase ratio assay was used to test the Wnt/β-catenin pathway activity. Western blotting was carried out to measure the accumulation of β-catenin in nuclear. Bioinformatics, Western blotting and dual luciferase assay were performed to investigate the target genes of miR-362-5p. Statistical differences were determined by the two-tailed Student′s t-test between two groups of data; a P value<0.05 was considered statistically significant.

Results

miR-362-5p was highly expressed both in TSCC samples and cell lines, P values were less than 0.001. SCC-9 and UM1 cells transfected with antagomiR-362-5p or miR-362-5p mimic significantly inhibited (t_SCC-9= 26.53, P_SCC-9= 0.0083; t_UM1= 24.45, P_UM1= 0.0094) or upregulated (t_SCC-9=-257.87, P_SCC-9= 0.0008; t_UM1=-270.44, P_UM1= 0.0007) miR-362-5p expression levels. MiR-362-5p inhibition markedly decreased SCC-9 and UM1 cells proliferation (t_SCC-9=32.44, P_SCC-9= 0.0009; t_UM1= 22.32, P_UM1= 0.0013) and invasion (t_SCC-9= 39.55, P_SCC-9= 0.0008; t_UM1= 23.78, P_UM1= 0.0092) , while the cell proliferation (t_SCC-9=-23.35, P_SCC-9= 0.0084; t_UM1=-17.47, P_UM1=0.0138) and invasion (t_SCC-9=-26.23, P_SCC-9= 0.0072; t_UM1=-18.69, P_UM1= 0.0145) was apparently enhanced by ectopic expression of miR-362-5p. Compared with that in control cells, miR-362-5p overexpression enhanced the Wnt/β-catenin signaling pathway by down-regulating DKK1 expression.

Conclusion

miR-362-5p activated Wnt/β-catenin signaling by directly targeting DKK1, which enhanced TSCC cells proliferation and invasion.

Key words: Carcinoma, squamous cell, Tongue, Neoplasm invasiveness, MiR-362-5p, DKK1, Wnt/β-catenin signaling pathway

Yuan Zhou, Haikuo Tang, Biao Hu. miR-362-5p targets DKK1 and promotes the proliferation and invasion in tongue squamous cell carcinoma[J]. Chinese Journal of Stomatological Research(Electronic Edition), 2017, 11(02): 86-92.

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References 15

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编号	样本量	舌鳞状细胞癌组织	癌旁正常组织	t值	P值
1	3	5.04 ± 1.27	0.50 ± 0.09	-115.34	0.0001
2	3	11.54 ± 3.44	0.49 ± 0.12	-298.89	0.0001
3	3	4.54 ± 0.98	0.40 ± 0.10	-278.55	0.0001
4	3	2.36 ± 0.88	0.28 ± 0.03	-90.67	0.0003
5	3	18.45 ± 1.00	0.75 ± 0.06	-114.87	0.0001
6	3	14.87 ± 3.74	0.47 ± 0.13	-220.65	0.0001
7	3	12.78 ± 2.88	0.58 ± 0.14	-269.85	0.0001
8	3	2.55 ± 0.89	0.32 ± 0.12	-88.56	0.0003
9	3	16.60 ± 1.46	0.67 ± 0.11	-170.36	0.0001
10	3	7.98 ± 1.98	0.45 ± 0.08	-159.88	0.0001
11	3	17.12 ± 4.00	0.57 ± 0.09	-240.44	0.0001
12	3	8.56 ± 2.07	0.68 ± 0.10	-115.78	0.0001
13	3	20.17 ± 3.12	0.75 ± 0.10	-157.87	0.0001
14	3	8.23 ± 2.82	0.50 ± 0.06	-263.68	0.0001
15	3	3.44 ± 0.83	0.58 ± 0.08	-98.99	0.0002
16	3	21.44 ± 3.63	0.61 ± 0.10	-227.76	0.0001
17	3	10.81 ± 1.42	0.44 ± 0.05	-233.34	0.0001
18	3	8.65 ± 1.10	0.57 ± 0.07	-189.57	0.0001
19	3	6.46 ± 1.55	0.64 ± 0.07	-159.98	0.0001
20	3	12.00 ± 1.43	0.55 ± 0.06	-210.68	0.0001

编号	样本量	舌鳞状细胞癌组织	癌旁正常组织	t值	P值
1	3	5.04 ± 1.27	0.50 ± 0.09	-115.34	0.0001
2	3	11.54 ± 3.44	0.49 ± 0.12	-298.89	0.0001
3	3	4.54 ± 0.98	0.40 ± 0.10	-278.55	0.0001
4	3	2.36 ± 0.88	0.28 ± 0.03	-90.67	0.0003
5	3	18.45 ± 1.00	0.75 ± 0.06	-114.87	0.0001
6	3	14.87 ± 3.74	0.47 ± 0.13	-220.65	0.0001
7	3	12.78 ± 2.88	0.58 ± 0.14	-269.85	0.0001
8	3	2.55 ± 0.89	0.32 ± 0.12	-88.56	0.0003
9	3	16.60 ± 1.46	0.67 ± 0.11	-170.36	0.0001
10	3	7.98 ± 1.98	0.45 ± 0.08	-159.88	0.0001
11	3	17.12 ± 4.00	0.57 ± 0.09	-240.44	0.0001
12	3	8.56 ± 2.07	0.68 ± 0.10	-115.78	0.0001
13	3	20.17 ± 3.12	0.75 ± 0.10	-157.87	0.0001
14	3	8.23 ± 2.82	0.50 ± 0.06	-263.68	0.0001
15	3	3.44 ± 0.83	0.58 ± 0.08	-98.99	0.0002
16	3	21.44 ± 3.63	0.61 ± 0.10	-227.76	0.0001
17	3	10.81 ± 1.42	0.44 ± 0.05	-233.34	0.0001
18	3	8.65 ± 1.10	0.57 ± 0.07	-189.57	0.0001
19	3	6.46 ± 1.55	0.64 ± 0.07	-159.98	0.0001
20	3	12.00 ± 1.43	0.55 ± 0.06	-210.68	0.0001

组别	样本量(n)	miR-362-5p	t值	P值
NTSC组	3	0.072 ± 0.015	-	-
SCC-9组	3	0.146 ± 0.026	-64.34	0.0014
SCC-15组	3	0.141 ± 0.029	-62.89	0.0014
SCC-25组	3	0.152 ± 0.032	-66.59	0.0013
UM1组	3	0.195 ± 0.055	-73.87	0.0005
UM2组	3	0.124 ± 0.021	-25.97	0.0084

组别	样本量(n)	miR-362-5p	t值	P值
NTSC组	3	0.072 ± 0.015	-	-
SCC-9组	3	0.146 ± 0.026	-64.34	0.0014
SCC-15组	3	0.141 ± 0.029	-62.89	0.0014
SCC-25组	3	0.152 ± 0.032	-66.59	0.0013
UM1组	3	0.195 ± 0.055	-73.87	0.0005
UM2组	3	0.124 ± 0.021	-25.97	0.0084

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