miR-106b靶向调控TGF-β/Smad通路对结肠癌细胞侵袭和迁移的促进作用

doi:10.13481/j.1671-587X.20210312

Abstract

Abstract: Objective

To investigate the effect of miR-106b targeting transforming growth factor-β receptor1（TGF-βR1）on the invasion and migration of colon cancer cells， and to clarify the targeting relationship between miR-106b and TGF-βR1 and its possible mechanism.

Methods

A total of 30 cases of colon cancer tissue and adjacent normal colon tissue，the colon cancer SW-480 cells and normal colon epithelial NCM460 cells were selected. The Real-time fluorescence quantitative PCR（RT-qPCR）method was used to detect the expression levels of miR-106b and TGF-βR1 mRNA in different kinds of tissues and cells.The colon cancer SW-480 cells were transfected and divided into empty vector group （transfected with miR-NC），miR-106b group （transfected with miR-106b-mimics），pGL3-TGF-βR1 group （transfected with pGL3-TGF-βR1）， and miR-106b-mimics+pGL3-TGF-βR1 group （transfected with miR-106b-mimics and pGL3-TGF-βR1）. The level of miR-106b in different kinds of tissues and cells，and the expression levels of TGF-βR1 mRNA in different kinds of cells were detected by RT-qPCR method；the targarc relationship between miR-106b and TGF-βR1 was predicted by bio-information analysis method.The luciferase activities of SW-480 cells in various groups were detected by luciferase reporter assay；the invasion abilities of the colon cancer SW-480 cells in various groups were detected by Transwell chamber test； the wound healing rates of the colon cancer SW-480 cells in various groups were detected by cell scratch test； the expression levels of TGF-βR1， phosphorylated Smad family member 2 （p-Smad2） and phosphorylated Smad family member 3（p-Smad3） proteins in colon cancer SW-480 cells in various groups were detected by Western blotting method.

Results

The level of miR-106b in the colon cancer tissue and colon cancer SW-480 cells were lower than those in normal colon tissue and normal colon epithelial NCM460 cells （P<0.01）.The results of double luciferase reporter gene analysis showed that TGFβR1 was the target gene of miR-106b.Compared with empty vector group，the expression levels of TGF-βR1 mRNA and protein in the colon cancer SW-480 cells in miR-106b-mimics+pGL3-TGF-βR1 group were significantly decreased（P<0.01）；the number of invasion cells and wound healing rate in the colon cancer SW-480 cells were significantly increased （P<0.01），and the expression levels of p-Smad2 and p-Smad3 proteins in the colon cancer SW-480 cells were decreased （P<0.01）.Compared with miR-106b group，the expression levels of TGF-βR1 mRNA and protein in the SW-480 cells in miR-106b-minics+PGL3-TGF-βR1 group were significantly increased（P<0.01），the number of invasion cells and the scrath healing rate were decreased（P<0.01），and the expression levels of p-Smad2 and p-Smad3 proteins in the cells were significantly increased（P<0.01）.

Conclusion

Over-expression of miR-106b may promote the invasion and migration of the colon cancer cells expression by inhibiting the TGF-β / Smad pathway.

Key words: miR-106b, transforming growth factor β receptor 1, colon neoplasms, cell invasion, cell migration

CLC Number:

R735.35

Bo MA, Jiangang LI, Jun WANG, Junli HOU, Liang LI. Promotion effect of miR-106b on invasion and migration of colon cancer cells through targeting TGF-β/Smad pathway[J].Journal of Jilin University(Medicine Edition), 2021, 47(3): 630-636.

Figures/Tables 7

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Group	Luciferase activity
Group	WT-TGF-βR1	MUT-TGF-βR1
Blank vector	1.12±0.09	1.09±0.12
miR-106b	0.46±0.05^*	0.99±0.11

Group	Number of invasion cells	p-Smad2 protein	p-Smad3 protein
Blank vector	169.2±12.3	1.22±0.01	1.34±0.02
miR-106b	265.4±20.3^*	0.30±0.01^*	0.23±0.01^*
pGL3-TGF-βR1	124.8±17.6^*△	2.63±0.02^*	2.54±0.12^*
miR-106b-mimics+pGL3-TGF-βR1	177.6±13.4^△#	1.51±0.06^△#	1.38±0.09^△#
F	24.521	174.532	207.348
P	<0.01	<0.01	<0.01

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Promotion effect of miR-106b on invasion and migration of colon cancer cells through targeting TGF-β/Smad pathway

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