Limits...
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

miR-506 inhibited osteosarcoma by targeting AEG-1 via the Wnt/β-catenin pathway signaling. (A) Western blot analysis showed that overexpression of miR-506 and knockdown of AEG-1 decreased the expression levels of β-catenin (94 kDa), c-myc (49 kDa) and cyclin D1 (34 kDa) in human osteosarcoma MG63 cells. *P<0.05 vs. miR-control or nontransfected; #P<0.05 vs. si-control. (B and C) Relative levels of β-catenin (B) mRNA and (C) protein were significantly decreased in MG63 cells transfected with si-β-catenin. *P<0.05, **P<0.01 vs. si-control. (D and E) Blocking the Wnt/β-catenin pathway using si-β-catenin (D) inhibited MG63 cell proliferation and (E) induced apoptosis of MG63 cells. *P<0.05, **P<0.01 vs. si-control. (F-H) CGP049090, a small molecule inhibitor of Wnt/β-catenin, (F) inhibited the expression of β-catenin, c-myc and cyclin D1 in MG63 cells in a concentration-dependent manner, (G) inhibited MG63 cell proliferation in a concentration-dependent manner and (H) induced apoptosis of MG63 cells (10µM CGP049090). *P<0.05, **P<0.01, ***P<0.001 vs. control (0 µM CGP049090). miR, microRNA; AEG-1, astrocyte elevated gene-1; si, small interfering RNA.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC4998420&req=5

f6-ol-0-0-4827: miR-506 inhibited osteosarcoma by targeting AEG-1 via the Wnt/β-catenin pathway signaling. (A) Western blot analysis showed that overexpression of miR-506 and knockdown of AEG-1 decreased the expression levels of β-catenin (94 kDa), c-myc (49 kDa) and cyclin D1 (34 kDa) in human osteosarcoma MG63 cells. *P<0.05 vs. miR-control or nontransfected; #P<0.05 vs. si-control. (B and C) Relative levels of β-catenin (B) mRNA and (C) protein were significantly decreased in MG63 cells transfected with si-β-catenin. *P<0.05, **P<0.01 vs. si-control. (D and E) Blocking the Wnt/β-catenin pathway using si-β-catenin (D) inhibited MG63 cell proliferation and (E) induced apoptosis of MG63 cells. *P<0.05, **P<0.01 vs. si-control. (F-H) CGP049090, a small molecule inhibitor of Wnt/β-catenin, (F) inhibited the expression of β-catenin, c-myc and cyclin D1 in MG63 cells in a concentration-dependent manner, (G) inhibited MG63 cell proliferation in a concentration-dependent manner and (H) induced apoptosis of MG63 cells (10µM CGP049090). *P<0.05, **P<0.01, ***P<0.001 vs. control (0 µM CGP049090). miR, microRNA; AEG-1, astrocyte elevated gene-1; si, small interfering RNA.

Mentions: The Wnt/β-catenin signaling pathway has been widely implicated in the development of multiple tumors, including osteosarcoma (26). The present study detected the expression levels of β-catenin, c-myc, and cyclin D1 in MG63 cells to determine whether miR-506 inhibits osteosarcoma via the Wnt/β-catenin pathway. As shown in Fig. 6A, upregulation of miR-506 and downregulation of AEG-1 clearly decreased the expression levels of β-catenin (P=0.0268 and P=0.0134, respectively), c-myc (P=0.0166 and 0.0129, respectively) and cyclin D1 (P=0.0288 and P=0.0260, respectively) in MG63 cells. In addition, the Wnt/β-catenin signaling pathway was inhibited by the present study using si-β-catenin. The levels of β-catenin mRNA and protein were significantly decreased in MG63 cells following transfection (P=0.0011 and P=0.0103, respectively; Fig. 6B and C). Blocking of the Wnt/β-catenin signaling pathway suppressed proliferation (P=0.0236) and induced apoptosis (P=0.0046) of MG63 cells (Fig. 6D and E). Furthermore, CGP049090, a small molecule inhibitor of Wnt/β-catenin, inhibited the expression of β-catenin (5 µM, P=0.0227; 10 µM, P=0.0086), c-myc (5 µM, P=0.0213; 10 µM, P=0.0017) and cyclin D1 (5 µM, P=0.0243; 10 µM, P=0.0033) in MG63 cells in a concentration-dependent manner (Fig. 6F). CGP049090 clearly inhibited proliferation (5 µM, P=0.0373; 10 µM, P=0.0088) and induced apoptosis (P=0.0008) of MG63 cells (Fig. 6G and H). These data demonstrate that miR-506 suppresses osteosarcoma by regulation of AEG-1 through inhibition of the Wnt/β-catenin signaling pathway.


Overexpression of miR-506 suppresses proliferation and promotes apoptosis of osteosarcoma cells by targeting astrocyte elevated gene-1
miR-506 inhibited osteosarcoma by targeting AEG-1 via the Wnt/β-catenin pathway signaling. (A) Western blot analysis showed that overexpression of miR-506 and knockdown of AEG-1 decreased the expression levels of β-catenin (94 kDa), c-myc (49 kDa) and cyclin D1 (34 kDa) in human osteosarcoma MG63 cells. *P<0.05 vs. miR-control or nontransfected; #P<0.05 vs. si-control. (B and C) Relative levels of β-catenin (B) mRNA and (C) protein were significantly decreased in MG63 cells transfected with si-β-catenin. *P<0.05, **P<0.01 vs. si-control. (D and E) Blocking the Wnt/β-catenin pathway using si-β-catenin (D) inhibited MG63 cell proliferation and (E) induced apoptosis of MG63 cells. *P<0.05, **P<0.01 vs. si-control. (F-H) CGP049090, a small molecule inhibitor of Wnt/β-catenin, (F) inhibited the expression of β-catenin, c-myc and cyclin D1 in MG63 cells in a concentration-dependent manner, (G) inhibited MG63 cell proliferation in a concentration-dependent manner and (H) induced apoptosis of MG63 cells (10µM CGP049090). *P<0.05, **P<0.01, ***P<0.001 vs. control (0 µM CGP049090). miR, microRNA; AEG-1, astrocyte elevated gene-1; si, small interfering RNA.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4998420&req=5

f6-ol-0-0-4827: miR-506 inhibited osteosarcoma by targeting AEG-1 via the Wnt/β-catenin pathway signaling. (A) Western blot analysis showed that overexpression of miR-506 and knockdown of AEG-1 decreased the expression levels of β-catenin (94 kDa), c-myc (49 kDa) and cyclin D1 (34 kDa) in human osteosarcoma MG63 cells. *P<0.05 vs. miR-control or nontransfected; #P<0.05 vs. si-control. (B and C) Relative levels of β-catenin (B) mRNA and (C) protein were significantly decreased in MG63 cells transfected with si-β-catenin. *P<0.05, **P<0.01 vs. si-control. (D and E) Blocking the Wnt/β-catenin pathway using si-β-catenin (D) inhibited MG63 cell proliferation and (E) induced apoptosis of MG63 cells. *P<0.05, **P<0.01 vs. si-control. (F-H) CGP049090, a small molecule inhibitor of Wnt/β-catenin, (F) inhibited the expression of β-catenin, c-myc and cyclin D1 in MG63 cells in a concentration-dependent manner, (G) inhibited MG63 cell proliferation in a concentration-dependent manner and (H) induced apoptosis of MG63 cells (10µM CGP049090). *P<0.05, **P<0.01, ***P<0.001 vs. control (0 µM CGP049090). miR, microRNA; AEG-1, astrocyte elevated gene-1; si, small interfering RNA.
Mentions: The Wnt/β-catenin signaling pathway has been widely implicated in the development of multiple tumors, including osteosarcoma (26). The present study detected the expression levels of β-catenin, c-myc, and cyclin D1 in MG63 cells to determine whether miR-506 inhibits osteosarcoma via the Wnt/β-catenin pathway. As shown in Fig. 6A, upregulation of miR-506 and downregulation of AEG-1 clearly decreased the expression levels of β-catenin (P=0.0268 and P=0.0134, respectively), c-myc (P=0.0166 and 0.0129, respectively) and cyclin D1 (P=0.0288 and P=0.0260, respectively) in MG63 cells. In addition, the Wnt/β-catenin signaling pathway was inhibited by the present study using si-β-catenin. The levels of β-catenin mRNA and protein were significantly decreased in MG63 cells following transfection (P=0.0011 and P=0.0103, respectively; Fig. 6B and C). Blocking of the Wnt/β-catenin signaling pathway suppressed proliferation (P=0.0236) and induced apoptosis (P=0.0046) of MG63 cells (Fig. 6D and E). Furthermore, CGP049090, a small molecule inhibitor of Wnt/β-catenin, inhibited the expression of β-catenin (5 µM, P=0.0227; 10 µM, P=0.0086), c-myc (5 µM, P=0.0213; 10 µM, P=0.0017) and cyclin D1 (5 µM, P=0.0243; 10 µM, P=0.0033) in MG63 cells in a concentration-dependent manner (Fig. 6F). CGP049090 clearly inhibited proliferation (5 µM, P=0.0373; 10 µM, P=0.0088) and induced apoptosis (P=0.0008) of MG63 cells (Fig. 6G and H). These data demonstrate that miR-506 suppresses osteosarcoma by regulation of AEG-1 through inhibition of the Wnt/β-catenin signaling pathway.

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