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Overexpression of miR-506 suppresses proliferation and promotes apoptosis of osteosarcoma cells by targeting astrocyte elevated gene-1

View Article: PubMed Central - PubMed

ABSTRACT

There is increasing evidence that microRNAs (miRs) are implicated in tumor development and progression; however, their specific roles in osteosarcoma are not well understood. The aim of the present study was to investigate the role of miR-506 in the pathogenesis of osteosarcoma. The expression levels of miR-506 and astrocyte elevated gene-1 (AEG-1) mRNA were detected using quantitative polymerase chain reaction, and the protein levels of AEG-1, β-catenin, c-myc and cyclin D1 were determined using western blot analysis. The effects of miR-506 and AEG-1 on cell viability, colony forming ability and apoptosis were assessed using MTT assay, colony formation assay, and flow cytometry, respectively. Lucifer reporter assays were used to demonstrate whether AEG-1 is a direct target of miR-506. The present study identified that miR-506 was downregulated in osteosarcoma tissues and cells. Overexpression of miR-506 suppressed the proliferation and induced apoptosis in osteosarcoma cells in vitro and inhibited tumor formation in vivo. Overexpression of miR-506 significantly inhibited the luciferase activity of AEG-1 with a wild-type 3′-untranslated region, providing clear evidence that AEG-1 was a direct and functional downstream target of miR-506. Similar to the overexpression of miR-506, downregulation of AEG-1 lead to an inhibitory effect on osteosarcoma in vitro. Furthermore, overexpression of miR-506 or downregulation of AEG-1 inhibited the Wnt/β-catenin signaling pathway, and inhibition of this pathway by β-catenin small interfering RNA or CGP049090, a small molecule inhibitor, suppressed cell proliferation and induced apoptosis in vitro. Overall, the present data indicated that miR-506 functions as a tumor suppressor by targeting AEG-1 in osteosarcoma via the regulation of the Wnt/β-catenin signaling pathway.

No MeSH data available.


Related in: MedlinePlus

Upregulation of miR-506 and downregulation of AEG-1 inhibited the growth of human osteosarcoma MG63 cells. (A) qPCR revealed that the relative level of miR-506 was significantly increased in MG63 cells transfected with miR-506 mimics. **P<0.01 vs. miR-control. (B) qPCR showed that the relative expression of AEG-1 mRNA was decreased in MG63 cells transfected with si-AEG-1. ***P<0.001 vs. si-control. (C) Western blot analysis showed that the relative level of AEG-1 protein (64 kDa) was decreased in MG63 cells transfected with si-AEG-1. **P<0.01 vs. si-control. (D and E) Upregulation of miR-506 (D) inhibited MG63 cell proliferation and (E) decreased the number of colonies of MG63 cells. *P<0.05, **P<0.01 vs. miR-control. (F and G) Downregulation of AEG-1 (F) inhibited MG63 cell proliferation and (G) decreased the number of colonies of MG63 cells. *P<0.05, **P<0.01 vs. si-control. miR, microRNA; AEG-1, astrocyte elevated gene-1; qPCR, quantitative polymerase chain reaction; si, small interfering RNA.
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f2-ol-0-0-4827: Upregulation of miR-506 and downregulation of AEG-1 inhibited the growth of human osteosarcoma MG63 cells. (A) qPCR revealed that the relative level of miR-506 was significantly increased in MG63 cells transfected with miR-506 mimics. **P<0.01 vs. miR-control. (B) qPCR showed that the relative expression of AEG-1 mRNA was decreased in MG63 cells transfected with si-AEG-1. ***P<0.001 vs. si-control. (C) Western blot analysis showed that the relative level of AEG-1 protein (64 kDa) was decreased in MG63 cells transfected with si-AEG-1. **P<0.01 vs. si-control. (D and E) Upregulation of miR-506 (D) inhibited MG63 cell proliferation and (E) decreased the number of colonies of MG63 cells. *P<0.05, **P<0.01 vs. miR-control. (F and G) Downregulation of AEG-1 (F) inhibited MG63 cell proliferation and (G) decreased the number of colonies of MG63 cells. *P<0.05, **P<0.01 vs. si-control. miR, microRNA; AEG-1, astrocyte elevated gene-1; qPCR, quantitative polymerase chain reaction; si, small interfering RNA.

Mentions: To clarify the regulatory effects of miR-506 on osteosarcoma, MG63 cells were transfected with miR-506 mimics. As shown in Fig. 2A, miR-506 was overexpressed in MG63 cells (P=0.0094), as determined by qPCR. In addition, the mRNA and protein level of AEG-1 was downregulated in MG63 cells by transfection with si-AEG-1 (P=0.0003 and P=0.0013, respectively; Fig. 2B and C). Overexpression of miR-506 significantly decreased the viability of MG63 cells compared with the miR-control-transfected group of cells (P=0.0038; Fig. 2D) and inhibited the colony forming ability of the cells (P=0.0157; Fig. 2E). Similarly, downregulation of AEG-1 inhibited the viability of MG63 cells (P=0.0024; Fig. 2F), and inhibited the colony forming ability of the cells (P=0.0012; Fig. 2G). These findings suggest that miR-506 and AEG-1 are involved in the regulation of MG63 cell proliferation.


Overexpression of miR-506 suppresses proliferation and promotes apoptosis of osteosarcoma cells by targeting astrocyte elevated gene-1
Upregulation of miR-506 and downregulation of AEG-1 inhibited the growth of human osteosarcoma MG63 cells. (A) qPCR revealed that the relative level of miR-506 was significantly increased in MG63 cells transfected with miR-506 mimics. **P<0.01 vs. miR-control. (B) qPCR showed that the relative expression of AEG-1 mRNA was decreased in MG63 cells transfected with si-AEG-1. ***P<0.001 vs. si-control. (C) Western blot analysis showed that the relative level of AEG-1 protein (64 kDa) was decreased in MG63 cells transfected with si-AEG-1. **P<0.01 vs. si-control. (D and E) Upregulation of miR-506 (D) inhibited MG63 cell proliferation and (E) decreased the number of colonies of MG63 cells. *P<0.05, **P<0.01 vs. miR-control. (F and G) Downregulation of AEG-1 (F) inhibited MG63 cell proliferation and (G) decreased the number of colonies of MG63 cells. *P<0.05, **P<0.01 vs. si-control. miR, microRNA; AEG-1, astrocyte elevated gene-1; qPCR, quantitative polymerase chain reaction; si, small interfering RNA.
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Related In: Results  -  Collection

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f2-ol-0-0-4827: Upregulation of miR-506 and downregulation of AEG-1 inhibited the growth of human osteosarcoma MG63 cells. (A) qPCR revealed that the relative level of miR-506 was significantly increased in MG63 cells transfected with miR-506 mimics. **P<0.01 vs. miR-control. (B) qPCR showed that the relative expression of AEG-1 mRNA was decreased in MG63 cells transfected with si-AEG-1. ***P<0.001 vs. si-control. (C) Western blot analysis showed that the relative level of AEG-1 protein (64 kDa) was decreased in MG63 cells transfected with si-AEG-1. **P<0.01 vs. si-control. (D and E) Upregulation of miR-506 (D) inhibited MG63 cell proliferation and (E) decreased the number of colonies of MG63 cells. *P<0.05, **P<0.01 vs. miR-control. (F and G) Downregulation of AEG-1 (F) inhibited MG63 cell proliferation and (G) decreased the number of colonies of MG63 cells. *P<0.05, **P<0.01 vs. si-control. miR, microRNA; AEG-1, astrocyte elevated gene-1; qPCR, quantitative polymerase chain reaction; si, small interfering RNA.
Mentions: To clarify the regulatory effects of miR-506 on osteosarcoma, MG63 cells were transfected with miR-506 mimics. As shown in Fig. 2A, miR-506 was overexpressed in MG63 cells (P=0.0094), as determined by qPCR. In addition, the mRNA and protein level of AEG-1 was downregulated in MG63 cells by transfection with si-AEG-1 (P=0.0003 and P=0.0013, respectively; Fig. 2B and C). Overexpression of miR-506 significantly decreased the viability of MG63 cells compared with the miR-control-transfected group of cells (P=0.0038; Fig. 2D) and inhibited the colony forming ability of the cells (P=0.0157; Fig. 2E). Similarly, downregulation of AEG-1 inhibited the viability of MG63 cells (P=0.0024; Fig. 2F), and inhibited the colony forming ability of the cells (P=0.0012; Fig. 2G). These findings suggest that miR-506 and AEG-1 are involved in the regulation of MG63 cell proliferation.

View Article: PubMed Central - PubMed

ABSTRACT

There is increasing evidence that microRNAs (miRs) are implicated in tumor development and progression; however, their specific roles in osteosarcoma are not well understood. The aim of the present study was to investigate the role of miR-506 in the pathogenesis of osteosarcoma. The expression levels of miR-506 and astrocyte elevated gene-1 (AEG-1) mRNA were detected using quantitative polymerase chain reaction, and the protein levels of AEG-1, &beta;-catenin, c-myc and cyclin D1 were determined using western blot analysis. The effects of miR-506 and AEG-1 on cell viability, colony forming ability and apoptosis were assessed using MTT assay, colony formation assay, and flow cytometry, respectively. Lucifer reporter assays were used to demonstrate whether AEG-1 is a direct target of miR-506. The present study identified that miR-506 was downregulated in osteosarcoma tissues and cells. Overexpression of miR-506 suppressed the proliferation and induced apoptosis in osteosarcoma cells in vitro and inhibited tumor formation in vivo. Overexpression of miR-506 significantly inhibited the luciferase activity of AEG-1 with a wild-type 3&prime;-untranslated region, providing clear evidence that AEG-1 was a direct and functional downstream target of miR-506. Similar to the overexpression of miR-506, downregulation of AEG-1 lead to an inhibitory effect on osteosarcoma in vitro. Furthermore, overexpression of miR-506 or downregulation of AEG-1 inhibited the Wnt/&beta;-catenin signaling pathway, and inhibition of this pathway by &beta;-catenin small interfering RNA or CGP049090, a small molecule inhibitor, suppressed cell proliferation and induced apoptosis in vitro. Overall, the present data indicated that miR-506 functions as a tumor suppressor by targeting AEG-1 in osteosarcoma via the regulation of the Wnt/&beta;-catenin signaling pathway.

No MeSH data available.


Related in: MedlinePlus