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MicroRNA‑197 reverses the drug resistance of fluorouracil‑induced SGC7901 cells by targeting mitogen‑activated protein kinase 1.

Xiong HL, Zhou SW, Sun AH, He Y, Li J, Yuan X - Mol Med Rep (2015)

Bottom Line: A luciferase reporter assay confirmed that miR‑197 led to silencing of the MAPK1 gene by recognizing and then specifically binding to the predicted site of the MAPK1 mRNA 3'‑untranslated region.When miR‑197 was overexpressed in SGC7901 cells, the protein levels of MAPK1 were downregulated.Furthermore, MAPK1 knockdown significantly increased the growth inhibition rate of the SGC7901/5‑FU cells compared with those in the control group.

View Article: PubMed Central - PubMed

Affiliation: Department of Medical Oncology, Huizhou Municipal Central Hospital of Guangdong Province, Huizhou, Guangdong 516000, P.R. China.

ABSTRACT
MicroRNAs (miRNAs) are a group of small non‑coding RNA molecules, which serve an important function in the development of multidrug resistance in cancer through the post‑transcriptional regulation of gene expression and RNA silencing. In the present study, the functional effects of miR‑197 were analyzed in chemo‑resistant gastric cancer cells. Low expression levels of miR‑197 were observed in the fluorouracil (5‑FU)‑resistant gastric cell line SGC7901/5‑FU when compared with those in the parental gastric cell line SGC7901. Overexpression of miR‑197 in SGC7901/5‑FU cells was identified to partially restore 5‑FU sensitivity. miRNA target prediction algorithms suggested that mitogen‑activated protein kinase 1 (MAPK1) is a candidate target gene for miR‑197. A luciferase reporter assay confirmed that miR‑197 led to silencing of the MAPK1 gene by recognizing and then specifically binding to the predicted site of the MAPK1 mRNA 3'‑untranslated region. When miR‑197 was overexpressed in SGC7901 cells, the protein levels of MAPK1 were downregulated. Furthermore, MAPK1 knockdown significantly increased the growth inhibition rate of the SGC7901/5‑FU cells compared with those in the control group. These results indicated that miR‑197 may influence the sensitivity of 5‑FU treatment in a gastric cancer cell line by targeting MAPK1.

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Expression levels of miR-197 affect the sensitivity of SGC7901 cells to 5-FU. (A) Following transfection with anti-miR-197, the expression levels of miR-197 were significantly reduced in SGC7901 cells. (B) Following transfection with anti-miR-197 or control, SGC7901 cells were treated with various doses of 5-FU (0.5, 1.0, 2.0, 4.0 or 8.0 µmol/l) for 48 h. An MTT assay was performed to determine the cell growth inhibition. (C) Following transfection of the miR-197 mimics, the expression levels of miR-197 were significantly increased in SGC7901/5-FU cells. (D) Following transfection with the miR-197 mimics or control, the SGC7901/5-FU cells were treated with various doses of 5-FU (0.5, 1.0, 2.0, 4.0 and 8.0 µmol/l) for 48 h. The MTT assay was performed to determine the cell growth inhibition. Values are expressed as the mean ± standard deviation (n=3). *P<0.05 vs. control. miR-197, microRNA-197; 5-FU, fluorouracil; SGC7901/5-FU, SGC7901 cell line resistant to 5-FU.
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f2-mmr-12-04-5019: Expression levels of miR-197 affect the sensitivity of SGC7901 cells to 5-FU. (A) Following transfection with anti-miR-197, the expression levels of miR-197 were significantly reduced in SGC7901 cells. (B) Following transfection with anti-miR-197 or control, SGC7901 cells were treated with various doses of 5-FU (0.5, 1.0, 2.0, 4.0 or 8.0 µmol/l) for 48 h. An MTT assay was performed to determine the cell growth inhibition. (C) Following transfection of the miR-197 mimics, the expression levels of miR-197 were significantly increased in SGC7901/5-FU cells. (D) Following transfection with the miR-197 mimics or control, the SGC7901/5-FU cells were treated with various doses of 5-FU (0.5, 1.0, 2.0, 4.0 and 8.0 µmol/l) for 48 h. The MTT assay was performed to determine the cell growth inhibition. Values are expressed as the mean ± standard deviation (n=3). *P<0.05 vs. control. miR-197, microRNA-197; 5-FU, fluorouracil; SGC7901/5-FU, SGC7901 cell line resistant to 5-FU.

Mentions: To investigate the association between miR-197 and 5-FU resistance in SGC7901 cells, the effect of downregulation of miR-197 in 5-FU-sensitive SGC7901 cells was assessed. A commercially synthesized miR-197 inhibitor (anti-miR-197) was used to alter the miR-197 expression levels in SGC7901 cells. The validity of miR-197 ectopic expression was confirmed by RT-qPCR following transfection with anti-miR-197 or the control vector into SGC7901 cells. The results indicated that the commercially synthe-sized anti-miR-197 was able to reduce miR-197 expression in SGC7901 cells compared with that in the control group (P<0.05; Fig. 2A). The group transfected with anti-miR-197 was observed to have a significantly higher survival rate than the control group (Fig. 2B). These results suggested that down-regulation of miR-197 led to 5-FU resistance of SGC7901 cells.


MicroRNA‑197 reverses the drug resistance of fluorouracil‑induced SGC7901 cells by targeting mitogen‑activated protein kinase 1.

Xiong HL, Zhou SW, Sun AH, He Y, Li J, Yuan X - Mol Med Rep (2015)

Expression levels of miR-197 affect the sensitivity of SGC7901 cells to 5-FU. (A) Following transfection with anti-miR-197, the expression levels of miR-197 were significantly reduced in SGC7901 cells. (B) Following transfection with anti-miR-197 or control, SGC7901 cells were treated with various doses of 5-FU (0.5, 1.0, 2.0, 4.0 or 8.0 µmol/l) for 48 h. An MTT assay was performed to determine the cell growth inhibition. (C) Following transfection of the miR-197 mimics, the expression levels of miR-197 were significantly increased in SGC7901/5-FU cells. (D) Following transfection with the miR-197 mimics or control, the SGC7901/5-FU cells were treated with various doses of 5-FU (0.5, 1.0, 2.0, 4.0 and 8.0 µmol/l) for 48 h. The MTT assay was performed to determine the cell growth inhibition. Values are expressed as the mean ± standard deviation (n=3). *P<0.05 vs. control. miR-197, microRNA-197; 5-FU, fluorouracil; SGC7901/5-FU, SGC7901 cell line resistant to 5-FU.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4581796&req=5

f2-mmr-12-04-5019: Expression levels of miR-197 affect the sensitivity of SGC7901 cells to 5-FU. (A) Following transfection with anti-miR-197, the expression levels of miR-197 were significantly reduced in SGC7901 cells. (B) Following transfection with anti-miR-197 or control, SGC7901 cells were treated with various doses of 5-FU (0.5, 1.0, 2.0, 4.0 or 8.0 µmol/l) for 48 h. An MTT assay was performed to determine the cell growth inhibition. (C) Following transfection of the miR-197 mimics, the expression levels of miR-197 were significantly increased in SGC7901/5-FU cells. (D) Following transfection with the miR-197 mimics or control, the SGC7901/5-FU cells were treated with various doses of 5-FU (0.5, 1.0, 2.0, 4.0 and 8.0 µmol/l) for 48 h. The MTT assay was performed to determine the cell growth inhibition. Values are expressed as the mean ± standard deviation (n=3). *P<0.05 vs. control. miR-197, microRNA-197; 5-FU, fluorouracil; SGC7901/5-FU, SGC7901 cell line resistant to 5-FU.
Mentions: To investigate the association between miR-197 and 5-FU resistance in SGC7901 cells, the effect of downregulation of miR-197 in 5-FU-sensitive SGC7901 cells was assessed. A commercially synthesized miR-197 inhibitor (anti-miR-197) was used to alter the miR-197 expression levels in SGC7901 cells. The validity of miR-197 ectopic expression was confirmed by RT-qPCR following transfection with anti-miR-197 or the control vector into SGC7901 cells. The results indicated that the commercially synthe-sized anti-miR-197 was able to reduce miR-197 expression in SGC7901 cells compared with that in the control group (P<0.05; Fig. 2A). The group transfected with anti-miR-197 was observed to have a significantly higher survival rate than the control group (Fig. 2B). These results suggested that down-regulation of miR-197 led to 5-FU resistance of SGC7901 cells.

Bottom Line: A luciferase reporter assay confirmed that miR‑197 led to silencing of the MAPK1 gene by recognizing and then specifically binding to the predicted site of the MAPK1 mRNA 3'‑untranslated region.When miR‑197 was overexpressed in SGC7901 cells, the protein levels of MAPK1 were downregulated.Furthermore, MAPK1 knockdown significantly increased the growth inhibition rate of the SGC7901/5‑FU cells compared with those in the control group.

View Article: PubMed Central - PubMed

Affiliation: Department of Medical Oncology, Huizhou Municipal Central Hospital of Guangdong Province, Huizhou, Guangdong 516000, P.R. China.

ABSTRACT
MicroRNAs (miRNAs) are a group of small non‑coding RNA molecules, which serve an important function in the development of multidrug resistance in cancer through the post‑transcriptional regulation of gene expression and RNA silencing. In the present study, the functional effects of miR‑197 were analyzed in chemo‑resistant gastric cancer cells. Low expression levels of miR‑197 were observed in the fluorouracil (5‑FU)‑resistant gastric cell line SGC7901/5‑FU when compared with those in the parental gastric cell line SGC7901. Overexpression of miR‑197 in SGC7901/5‑FU cells was identified to partially restore 5‑FU sensitivity. miRNA target prediction algorithms suggested that mitogen‑activated protein kinase 1 (MAPK1) is a candidate target gene for miR‑197. A luciferase reporter assay confirmed that miR‑197 led to silencing of the MAPK1 gene by recognizing and then specifically binding to the predicted site of the MAPK1 mRNA 3'‑untranslated region. When miR‑197 was overexpressed in SGC7901 cells, the protein levels of MAPK1 were downregulated. Furthermore, MAPK1 knockdown significantly increased the growth inhibition rate of the SGC7901/5‑FU cells compared with those in the control group. These results indicated that miR‑197 may influence the sensitivity of 5‑FU treatment in a gastric cancer cell line by targeting MAPK1.

Show MeSH
Related in: MedlinePlus