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MicroRNA-7 inhibits tumor metastasis and reverses epithelial-mesenchymal transition through AKT/ERK1/2 inactivation by targeting EGFR in epithelial ovarian cancer.

Zhou X, Hu Y, Dai L, Wang Y, Zhou J, Wang W, Di W, Qiu L - PLoS ONE (2014)

Bottom Line: Overexpression of miR-7 markedly suppressed the capacities of cell invasion and migration and resulted in morphological changes from a mesenchymal phenotype to an epithelial-like phenotype in EOC.The pharmacological inhibition of PI3K-AKT and ERK1/2 both significantly enhanced CK-18 and β-catenin expression and suppressed vimentin expression, indicating that AKT and ERK1/2 pathways are required for miR-7 mediating EMT.Taken together, our results suggested that miR-7 inhibited tumor metastasis and reversed EMT through AKT and ERK1/2 pathway inactivation by reducing EGFR expression in EOC cell lines.

View Article: PubMed Central - PubMed

Affiliation: Department of Gynecology and Obstetrics, Renji Hospital, School of Medicine, Shanghai Jiao Tong University, Shanghai, China; Shanghai Key Laboratory of Gynecologic Oncology, Shanghai, China.

ABSTRACT
Epidermal growth factor receptor (EGFR) overexpression and activation result in increased proliferation and migration of solid tumors including ovarian cancer. In recent years, mounting evidence indicates that EGFR is a direct and functional target of miR-7. In this study, we found that miR-7 expression was significantly downregulated in highly metastatic epithelial ovarian cancer (EOC) cell lines and metastatic tissues, whereas the expression of, EGFR correlated positively with metastasis in both EOC patients and cell lines. Overexpression of miR-7 markedly suppressed the capacities of cell invasion and migration and resulted in morphological changes from a mesenchymal phenotype to an epithelial-like phenotype in EOC. In addition, overexpression of miR-7 upregulated CK-18 and β-catenin expression and downregulated Vimentin expression, accompanied with EGFR inhibition and AKT/ERK1/2 inactivation. Similar to miR-7 transfection, silencing of EGFR with this siRNA in EOC cells also upregulated CK-18 and β-catenin expression and downregulated Vimentin expression, and decreased phosphorylation of both Akt and ERK1/2, confirming that EGFR is a target of miR-7 in reversing EMT. The pharmacological inhibition of PI3K-AKT and ERK1/2 both significantly enhanced CK-18 and β-catenin expression and suppressed vimentin expression, indicating that AKT and ERK1/2 pathways are required for miR-7 mediating EMT. Finally, the expression of miR-7 and EGFR in primary EOC with matched metastasis tissues was explored. It was showed that miR-7 is inversely correlated with EGFR. Taken together, our results suggested that miR-7 inhibited tumor metastasis and reversed EMT through AKT and ERK1/2 pathway inactivation by reducing EGFR expression in EOC cell lines. Thus, miR-7 might be a potential prognostic marker and therapeutic target for ovarian cancer metastasis intervention.

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miR-7 expression is inversely correlated with EOC metastasis.(A) The relative expression of miR-7 in 17-paired EOC tissues from omentum or peritoneum metastases and primary EOC tissues was detected by qRT-PCR. The U6 small nuclear RNA was used as an internal control and the fold change was calculated by the ΔΔCt method (B) The expression level of miR-7 in one pair of low and high metastatic EOC cell lines. (C) The expression level of miR-7 in seven EOC cell lines. (*P<0.05. **P<0.01).
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pone-0096718-g001: miR-7 expression is inversely correlated with EOC metastasis.(A) The relative expression of miR-7 in 17-paired EOC tissues from omentum or peritoneum metastases and primary EOC tissues was detected by qRT-PCR. The U6 small nuclear RNA was used as an internal control and the fold change was calculated by the ΔΔCt method (B) The expression level of miR-7 in one pair of low and high metastatic EOC cell lines. (C) The expression level of miR-7 in seven EOC cell lines. (*P<0.05. **P<0.01).

Mentions: To explore the expression and significance of miR-7 in EOC metastasis, we detected miR-7 expression in 17-paired metastatic EOC tissues and primary EOC tissues. Quantitative real-time PCR (qRT-PCR) showed that tissues from omentum or peritoneum metastases expressed lower levels of miR-7 compared with primary EOC tissues, indicating an inverse relationship between the expression of miR-7 and the metastatic status of EOC tissues (Figure 1A). Furthermore, we selected HO-8910 and its highly metastatic clone HO-8910pm. HO-8910pm was established and characterized in Cell Bank, Chinese Academy of Sciences [19], [20], [21]. We found that miR-7 expression was significantly decreased in highly metastatic clone HO-8910pm compared with HO-8910 (Figure 1B). Taken together, our results suggest that down-regulation of miR-7 is correlated with increased EOC metastasis and that miR-7 might suppress EOC progression. To determine the optimal cell lines for further study, we measured the expression of miR-7 in seven EOC cell lines (HO-8910pm, HO-8910, A2780, A2780/DDP, SKOV-3, CAOV-3 and ES-2). Our data showed that the expression of miR-7 was lowest in ES-2 cell line (P<0.05) (Figure 1C), which is another EOC cell with highly metastatic potential. Therefore, we choose HO-8910PM and ES-2 to study the mechanisms of miR-7 inhibiting metastasis in EOC in the following experiments.


MicroRNA-7 inhibits tumor metastasis and reverses epithelial-mesenchymal transition through AKT/ERK1/2 inactivation by targeting EGFR in epithelial ovarian cancer.

Zhou X, Hu Y, Dai L, Wang Y, Zhou J, Wang W, Di W, Qiu L - PLoS ONE (2014)

miR-7 expression is inversely correlated with EOC metastasis.(A) The relative expression of miR-7 in 17-paired EOC tissues from omentum or peritoneum metastases and primary EOC tissues was detected by qRT-PCR. The U6 small nuclear RNA was used as an internal control and the fold change was calculated by the ΔΔCt method (B) The expression level of miR-7 in one pair of low and high metastatic EOC cell lines. (C) The expression level of miR-7 in seven EOC cell lines. (*P<0.05. **P<0.01).
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Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC4016102&req=5

pone-0096718-g001: miR-7 expression is inversely correlated with EOC metastasis.(A) The relative expression of miR-7 in 17-paired EOC tissues from omentum or peritoneum metastases and primary EOC tissues was detected by qRT-PCR. The U6 small nuclear RNA was used as an internal control and the fold change was calculated by the ΔΔCt method (B) The expression level of miR-7 in one pair of low and high metastatic EOC cell lines. (C) The expression level of miR-7 in seven EOC cell lines. (*P<0.05. **P<0.01).
Mentions: To explore the expression and significance of miR-7 in EOC metastasis, we detected miR-7 expression in 17-paired metastatic EOC tissues and primary EOC tissues. Quantitative real-time PCR (qRT-PCR) showed that tissues from omentum or peritoneum metastases expressed lower levels of miR-7 compared with primary EOC tissues, indicating an inverse relationship between the expression of miR-7 and the metastatic status of EOC tissues (Figure 1A). Furthermore, we selected HO-8910 and its highly metastatic clone HO-8910pm. HO-8910pm was established and characterized in Cell Bank, Chinese Academy of Sciences [19], [20], [21]. We found that miR-7 expression was significantly decreased in highly metastatic clone HO-8910pm compared with HO-8910 (Figure 1B). Taken together, our results suggest that down-regulation of miR-7 is correlated with increased EOC metastasis and that miR-7 might suppress EOC progression. To determine the optimal cell lines for further study, we measured the expression of miR-7 in seven EOC cell lines (HO-8910pm, HO-8910, A2780, A2780/DDP, SKOV-3, CAOV-3 and ES-2). Our data showed that the expression of miR-7 was lowest in ES-2 cell line (P<0.05) (Figure 1C), which is another EOC cell with highly metastatic potential. Therefore, we choose HO-8910PM and ES-2 to study the mechanisms of miR-7 inhibiting metastasis in EOC in the following experiments.

Bottom Line: Overexpression of miR-7 markedly suppressed the capacities of cell invasion and migration and resulted in morphological changes from a mesenchymal phenotype to an epithelial-like phenotype in EOC.The pharmacological inhibition of PI3K-AKT and ERK1/2 both significantly enhanced CK-18 and β-catenin expression and suppressed vimentin expression, indicating that AKT and ERK1/2 pathways are required for miR-7 mediating EMT.Taken together, our results suggested that miR-7 inhibited tumor metastasis and reversed EMT through AKT and ERK1/2 pathway inactivation by reducing EGFR expression in EOC cell lines.

View Article: PubMed Central - PubMed

Affiliation: Department of Gynecology and Obstetrics, Renji Hospital, School of Medicine, Shanghai Jiao Tong University, Shanghai, China; Shanghai Key Laboratory of Gynecologic Oncology, Shanghai, China.

ABSTRACT
Epidermal growth factor receptor (EGFR) overexpression and activation result in increased proliferation and migration of solid tumors including ovarian cancer. In recent years, mounting evidence indicates that EGFR is a direct and functional target of miR-7. In this study, we found that miR-7 expression was significantly downregulated in highly metastatic epithelial ovarian cancer (EOC) cell lines and metastatic tissues, whereas the expression of, EGFR correlated positively with metastasis in both EOC patients and cell lines. Overexpression of miR-7 markedly suppressed the capacities of cell invasion and migration and resulted in morphological changes from a mesenchymal phenotype to an epithelial-like phenotype in EOC. In addition, overexpression of miR-7 upregulated CK-18 and β-catenin expression and downregulated Vimentin expression, accompanied with EGFR inhibition and AKT/ERK1/2 inactivation. Similar to miR-7 transfection, silencing of EGFR with this siRNA in EOC cells also upregulated CK-18 and β-catenin expression and downregulated Vimentin expression, and decreased phosphorylation of both Akt and ERK1/2, confirming that EGFR is a target of miR-7 in reversing EMT. The pharmacological inhibition of PI3K-AKT and ERK1/2 both significantly enhanced CK-18 and β-catenin expression and suppressed vimentin expression, indicating that AKT and ERK1/2 pathways are required for miR-7 mediating EMT. Finally, the expression of miR-7 and EGFR in primary EOC with matched metastasis tissues was explored. It was showed that miR-7 is inversely correlated with EGFR. Taken together, our results suggested that miR-7 inhibited tumor metastasis and reversed EMT through AKT and ERK1/2 pathway inactivation by reducing EGFR expression in EOC cell lines. Thus, miR-7 might be a potential prognostic marker and therapeutic target for ovarian cancer metastasis intervention.

Show MeSH
Related in: MedlinePlus