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Cisplatin-induced epigenetic activation of miR-34a sensitizes bladder cancer cells to chemotherapy.

Li H, Yu G, Shi R, Lang B, Chen X, Xia D, Xiao H, Guo X, Guan W, Ye Z, Xiao W, Xu H - Mol. Cancer (2014)

Bottom Line: Synthetic short single or double stranded RNA oligonucleotides and lentiviral vector were used to regulate miR-34a expression in MIBC cells to investigate its function in vitro and in vivo. miR-34a expression was frequently decreased in MIBC tissues and cell lines through promoter hypermethylation while it was epigenetically increased in MIBC cells following cisplatin treatment.Furthermore, we identified CD44 as being targeted by miR-34a in MIBC cells following cisplatin treatment, and increased CD44 expression could efficiently reverse the effect of miR-34a on MIBC cell proliferation, colongenic potential and chemosensitivity.Cisplatin-based chemotherapy induced demethylation of miR-34a promoter and increased miR-34a expression, which in turn sensitized MIBC cells to cisplatin and decreased the tumorigenicity and proliferation of cancer cells that by reducing the production of CD44.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Urology, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430030, China. xiaowei0041@163.com.

ABSTRACT

Background: Accumulating evidence suggests a tumor suppressive role for miR-34a in human carcinogenesis. However, its precise biological role remains largely elusive. This study aimed to reveal the association of the miR-34a expression and its modulation of sensitivity to cisplatin in muscle-invasive bladder cancer (MIBC).

Methods: miR-34a expression in MIBC cell lines and patient tissues was investigated using qPCR. The methylation analysis of miR-34a promoter region was performed by MassARRAY. Synthetic short single or double stranded RNA oligonucleotides and lentiviral vector were used to regulate miR-34a expression in MIBC cells to investigate its function in vitro and in vivo.

Results: miR-34a expression was frequently decreased in MIBC tissues and cell lines through promoter hypermethylation while it was epigenetically increased in MIBC cells following cisplatin treatment. Increased miR-34a expression significantly sensitized MIBC cells to cisplatin and inhibited the tumorigenicity and proliferation of cancer cells in vitro and in vivo. Furthermore, we identified CD44 as being targeted by miR-34a in MIBC cells following cisplatin treatment, and increased CD44 expression could efficiently reverse the effect of miR-34a on MIBC cell proliferation, colongenic potential and chemosensitivity.

Conclusions: Cisplatin-based chemotherapy induced demethylation of miR-34a promoter and increased miR-34a expression, which in turn sensitized MIBC cells to cisplatin and decreased the tumorigenicity and proliferation of cancer cells that by reducing the production of CD44.

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The tumor-suppressive and chemosensitivity functions of miR-34a were mediated by reduction the production of CD44. Downregulation of CD44 by siRNA led to similar effect of miR-34a overexpression on A) cell proliferation (mean ± SEM; n = 3; *p < 0.05) and B-C) tumorigenity (mean + SEM; n = 3; *p < 0.05). Increased CD44 expression could efficiently reverse the effect of miR-34a on MIBC D) cell proliferation (mean ± SEM; n = 3; *p < 0.05), E-F) colongenic potential and G) chemosensitivity (mean + SEM; n = 3; *p < 0.05).
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Figure 6: The tumor-suppressive and chemosensitivity functions of miR-34a were mediated by reduction the production of CD44. Downregulation of CD44 by siRNA led to similar effect of miR-34a overexpression on A) cell proliferation (mean ± SEM; n = 3; *p < 0.05) and B-C) tumorigenity (mean + SEM; n = 3; *p < 0.05). Increased CD44 expression could efficiently reverse the effect of miR-34a on MIBC D) cell proliferation (mean ± SEM; n = 3; *p < 0.05), E-F) colongenic potential and G) chemosensitivity (mean + SEM; n = 3; *p < 0.05).

Mentions: We then tested whether miR-34a inhibited MIBC cell proliferation, colongenic potential and chemosensitivity through targeting CD44. Firstly, by using a CD44 siRNA, we demonstrated that downregulation of CD44 could efficiently inhibit cell proliferation (Figure 6A), decrease colony and sphere formation ability (Figure 6B and C) in all three MIBC cell lines, which correlated with the effect of overexpressed miR-34a. Subsequently, these assays were repeated when miR-34a mimics and CD44 expressing vector were co-transfected into 5637 and T24 cells. As shown in Figure 6D-F, CD44 overproduction appeared to have a dramatic positive effect on tumor cell growth and tumorigenesis, and importantly, miR-34a induced tumor suppression was largely eliminated upon the overexpression of CD44. Finally, we measured the IC50 values of cisplatin for these MIBC cell lines in differently treated group (Figure 6G), and the results showed that overexpression of CD44 could also efficiently reverse the effect of miR-34a on chemosensitivity. All these data presented here strongly suggesting that the tumor-suppressive and chemosensitivity effect of miR-34a was mediated by reducing the production of CD44 as its predominant target.


Cisplatin-induced epigenetic activation of miR-34a sensitizes bladder cancer cells to chemotherapy.

Li H, Yu G, Shi R, Lang B, Chen X, Xia D, Xiao H, Guo X, Guan W, Ye Z, Xiao W, Xu H - Mol. Cancer (2014)

The tumor-suppressive and chemosensitivity functions of miR-34a were mediated by reduction the production of CD44. Downregulation of CD44 by siRNA led to similar effect of miR-34a overexpression on A) cell proliferation (mean ± SEM; n = 3; *p < 0.05) and B-C) tumorigenity (mean + SEM; n = 3; *p < 0.05). Increased CD44 expression could efficiently reverse the effect of miR-34a on MIBC D) cell proliferation (mean ± SEM; n = 3; *p < 0.05), E-F) colongenic potential and G) chemosensitivity (mean + SEM; n = 3; *p < 0.05).
© Copyright Policy - open-access
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC4022035&req=5

Figure 6: The tumor-suppressive and chemosensitivity functions of miR-34a were mediated by reduction the production of CD44. Downregulation of CD44 by siRNA led to similar effect of miR-34a overexpression on A) cell proliferation (mean ± SEM; n = 3; *p < 0.05) and B-C) tumorigenity (mean + SEM; n = 3; *p < 0.05). Increased CD44 expression could efficiently reverse the effect of miR-34a on MIBC D) cell proliferation (mean ± SEM; n = 3; *p < 0.05), E-F) colongenic potential and G) chemosensitivity (mean + SEM; n = 3; *p < 0.05).
Mentions: We then tested whether miR-34a inhibited MIBC cell proliferation, colongenic potential and chemosensitivity through targeting CD44. Firstly, by using a CD44 siRNA, we demonstrated that downregulation of CD44 could efficiently inhibit cell proliferation (Figure 6A), decrease colony and sphere formation ability (Figure 6B and C) in all three MIBC cell lines, which correlated with the effect of overexpressed miR-34a. Subsequently, these assays were repeated when miR-34a mimics and CD44 expressing vector were co-transfected into 5637 and T24 cells. As shown in Figure 6D-F, CD44 overproduction appeared to have a dramatic positive effect on tumor cell growth and tumorigenesis, and importantly, miR-34a induced tumor suppression was largely eliminated upon the overexpression of CD44. Finally, we measured the IC50 values of cisplatin for these MIBC cell lines in differently treated group (Figure 6G), and the results showed that overexpression of CD44 could also efficiently reverse the effect of miR-34a on chemosensitivity. All these data presented here strongly suggesting that the tumor-suppressive and chemosensitivity effect of miR-34a was mediated by reducing the production of CD44 as its predominant target.

Bottom Line: Synthetic short single or double stranded RNA oligonucleotides and lentiviral vector were used to regulate miR-34a expression in MIBC cells to investigate its function in vitro and in vivo. miR-34a expression was frequently decreased in MIBC tissues and cell lines through promoter hypermethylation while it was epigenetically increased in MIBC cells following cisplatin treatment.Furthermore, we identified CD44 as being targeted by miR-34a in MIBC cells following cisplatin treatment, and increased CD44 expression could efficiently reverse the effect of miR-34a on MIBC cell proliferation, colongenic potential and chemosensitivity.Cisplatin-based chemotherapy induced demethylation of miR-34a promoter and increased miR-34a expression, which in turn sensitized MIBC cells to cisplatin and decreased the tumorigenicity and proliferation of cancer cells that by reducing the production of CD44.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Urology, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430030, China. xiaowei0041@163.com.

ABSTRACT

Background: Accumulating evidence suggests a tumor suppressive role for miR-34a in human carcinogenesis. However, its precise biological role remains largely elusive. This study aimed to reveal the association of the miR-34a expression and its modulation of sensitivity to cisplatin in muscle-invasive bladder cancer (MIBC).

Methods: miR-34a expression in MIBC cell lines and patient tissues was investigated using qPCR. The methylation analysis of miR-34a promoter region was performed by MassARRAY. Synthetic short single or double stranded RNA oligonucleotides and lentiviral vector were used to regulate miR-34a expression in MIBC cells to investigate its function in vitro and in vivo.

Results: miR-34a expression was frequently decreased in MIBC tissues and cell lines through promoter hypermethylation while it was epigenetically increased in MIBC cells following cisplatin treatment. Increased miR-34a expression significantly sensitized MIBC cells to cisplatin and inhibited the tumorigenicity and proliferation of cancer cells in vitro and in vivo. Furthermore, we identified CD44 as being targeted by miR-34a in MIBC cells following cisplatin treatment, and increased CD44 expression could efficiently reverse the effect of miR-34a on MIBC cell proliferation, colongenic potential and chemosensitivity.

Conclusions: Cisplatin-based chemotherapy induced demethylation of miR-34a promoter and increased miR-34a expression, which in turn sensitized MIBC cells to cisplatin and decreased the tumorigenicity and proliferation of cancer cells that by reducing the production of CD44.

Show MeSH
Related in: MedlinePlus