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Hiwi Promotes the Proliferation of Colorectal Cancer Cells via Upregulating Global DNA Methylation.

Yang L, Bi L, Liu Q, Zhao M, Cao B, Li D, Xiu J - Dis. Markers (2015)

Bottom Line: And the chemical inhibition of DNA methylation significantly restrained such proliferation promotion.In summary, we confirmed that Hiwi was overexpressed in CRC tissues and that the forced Hiwi overexpression promoted the proliferation and global DNA methylation of CRC cell lines.Our results imply for the first time that Hiwi promotes the proliferation of CRC cells via promoting global DNA methylation.

View Article: PubMed Central - PubMed

Affiliation: Department of Radiology, Provincial Hospital Affiliated to Shandong University, Jinan 250021, China.

ABSTRACT
Hiwi is well known for its role in stem cell renewal, maintaining the resting stage, and downregulating cell cycle of stem cells via RNA silencing. And Hiwi overexpression has been recognized in several types of cancers. In the present study, we examined the Hiwi expression in colorectal cancer (CRC) specimens in both mRNA and protein levels via real-time quantitative PCR, western blot assay, and immunohistochemical staining. Then we explored the role of Hiwi in the cancer cell proliferation and in the DNA methylation in human CRC Caro-2 and HT-29 cell lines. Results demonstrated that both mRNA and protein levels of Hiwi were significantly higher in 38 CRC tissues than in 38 peritumor tissues. Moreover, the Hiwi overexpression with an adenovirus vector significantly promoted the proliferation of Caro-2 and HT-29 cells, associated with significant increase in the global DNA methylation levels. And the chemical inhibition of DNA methylation significantly restrained such proliferation promotion. In summary, we confirmed that Hiwi was overexpressed in CRC tissues and that the forced Hiwi overexpression promoted the proliferation and global DNA methylation of CRC cell lines. Our results imply for the first time that Hiwi promotes the proliferation of CRC cells via promoting global DNA methylation.

No MeSH data available.


Related in: MedlinePlus

Inhibition of methylation blocked proliferation effect of Ad-Hiwi to Caco-2 or HT-29 cells. (a)–(d): DNA methylation levels in CACO-2 cells ((a), (b)) or HT-29 cells ((c), (d)) infected with 1 MOI Ad-RFP ((a), (c)) or with 1 MOI Ad-Hiwi ((b), (d)) after the treatment with 0, 0.05, 0.2, 0.8, or 2 μM DAC for 24 hours, respectively. (e) and (f): 2 μM DAC inhibited the Ad-Hiwi-promoted proliferation of Caco-2 or HT-29 cells by CCK-8 assay. Statistical significance was shown as ∗p < 0.05, ∗∗p < 0.01, and ∗∗∗p < 0.001; ns, no significance.
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fig5: Inhibition of methylation blocked proliferation effect of Ad-Hiwi to Caco-2 or HT-29 cells. (a)–(d): DNA methylation levels in CACO-2 cells ((a), (b)) or HT-29 cells ((c), (d)) infected with 1 MOI Ad-RFP ((a), (c)) or with 1 MOI Ad-Hiwi ((b), (d)) after the treatment with 0, 0.05, 0.2, 0.8, or 2 μM DAC for 24 hours, respectively. (e) and (f): 2 μM DAC inhibited the Ad-Hiwi-promoted proliferation of Caco-2 or HT-29 cells by CCK-8 assay. Statistical significance was shown as ∗p < 0.05, ∗∗p < 0.01, and ∗∗∗p < 0.001; ns, no significance.

Mentions: Aiming to confirm the influence by DNA methylation on the growth of CRC cells, 5-aza-2′-deoxycytidine (DAC) was used for blocking the DNA methylation in Caco-2 or HT-29 cells which were infected with Ad-Hiwi or Ad-RFP. As shown in Figure 5(a), there was no difference in the DNA methylation level between 0 mM DAC (control) and 0.05, 0.2, or 0.8 mM DAC in Caco-2 cells infected by Ad-RFP; only the DAC of 2 mM significantly reduced the DNA methylation level in the Ad-RFP-infected Caco-2 cells (Figure 4(a); p < 0.05). However, the DAC with more than 200 nM significantly reduced the DNA methylation level in the Ad-Hiwi-infected Caco-2 cells (Figure 4(b); p < 0.05), dose dependently (p < 0.05 for 200 Nm and 2 Mm DAC). Compared to Caco-2 cells infected by Ad-RFP or Ad-Hiwi and without DAC treatment, there was a difference when DAC was more than 0.8 mM in HT-29 cells infected by Ad-RFP and when DAC was more than 0.2 mM in HT-29 cells after the infection with Ad-Hiwi (Figures 5(c) and 5(d)). Moreover, the cell proliferation was determined by CCK-8 assay after the Ad-Hiwi infection and the treatment with 0, 0.05, 0.2, 0.8, or 2 μM DAC for 24 hours. As shown in Figure 5(e), the 2 mM DAC significantly inhibited the promoted cell proliferation by the Ad-Hiwi infection (p < 0.05 for 24 or 72 H.P.I., p < 0.01 for 48 H.P.I.). And such inhibition by DAC was reconfirmed in HT-29 cells. The proliferation curve of the Ad-Hiwi-infected HT-29 cells treated with 2 mM DAC was also lower than the Ad-Hiwi-infected HT-29 cells without DAC treatment (Figure 5(f)). These findings indicated that DNA methylation knockdown blocked the proliferation promotion by the Hiwi overexpression in CRC cells.


Hiwi Promotes the Proliferation of Colorectal Cancer Cells via Upregulating Global DNA Methylation.

Yang L, Bi L, Liu Q, Zhao M, Cao B, Li D, Xiu J - Dis. Markers (2015)

Inhibition of methylation blocked proliferation effect of Ad-Hiwi to Caco-2 or HT-29 cells. (a)–(d): DNA methylation levels in CACO-2 cells ((a), (b)) or HT-29 cells ((c), (d)) infected with 1 MOI Ad-RFP ((a), (c)) or with 1 MOI Ad-Hiwi ((b), (d)) after the treatment with 0, 0.05, 0.2, 0.8, or 2 μM DAC for 24 hours, respectively. (e) and (f): 2 μM DAC inhibited the Ad-Hiwi-promoted proliferation of Caco-2 or HT-29 cells by CCK-8 assay. Statistical significance was shown as ∗p < 0.05, ∗∗p < 0.01, and ∗∗∗p < 0.001; ns, no significance.
© Copyright Policy - open-access
Related In: Results  -  Collection

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fig5: Inhibition of methylation blocked proliferation effect of Ad-Hiwi to Caco-2 or HT-29 cells. (a)–(d): DNA methylation levels in CACO-2 cells ((a), (b)) or HT-29 cells ((c), (d)) infected with 1 MOI Ad-RFP ((a), (c)) or with 1 MOI Ad-Hiwi ((b), (d)) after the treatment with 0, 0.05, 0.2, 0.8, or 2 μM DAC for 24 hours, respectively. (e) and (f): 2 μM DAC inhibited the Ad-Hiwi-promoted proliferation of Caco-2 or HT-29 cells by CCK-8 assay. Statistical significance was shown as ∗p < 0.05, ∗∗p < 0.01, and ∗∗∗p < 0.001; ns, no significance.
Mentions: Aiming to confirm the influence by DNA methylation on the growth of CRC cells, 5-aza-2′-deoxycytidine (DAC) was used for blocking the DNA methylation in Caco-2 or HT-29 cells which were infected with Ad-Hiwi or Ad-RFP. As shown in Figure 5(a), there was no difference in the DNA methylation level between 0 mM DAC (control) and 0.05, 0.2, or 0.8 mM DAC in Caco-2 cells infected by Ad-RFP; only the DAC of 2 mM significantly reduced the DNA methylation level in the Ad-RFP-infected Caco-2 cells (Figure 4(a); p < 0.05). However, the DAC with more than 200 nM significantly reduced the DNA methylation level in the Ad-Hiwi-infected Caco-2 cells (Figure 4(b); p < 0.05), dose dependently (p < 0.05 for 200 Nm and 2 Mm DAC). Compared to Caco-2 cells infected by Ad-RFP or Ad-Hiwi and without DAC treatment, there was a difference when DAC was more than 0.8 mM in HT-29 cells infected by Ad-RFP and when DAC was more than 0.2 mM in HT-29 cells after the infection with Ad-Hiwi (Figures 5(c) and 5(d)). Moreover, the cell proliferation was determined by CCK-8 assay after the Ad-Hiwi infection and the treatment with 0, 0.05, 0.2, 0.8, or 2 μM DAC for 24 hours. As shown in Figure 5(e), the 2 mM DAC significantly inhibited the promoted cell proliferation by the Ad-Hiwi infection (p < 0.05 for 24 or 72 H.P.I., p < 0.01 for 48 H.P.I.). And such inhibition by DAC was reconfirmed in HT-29 cells. The proliferation curve of the Ad-Hiwi-infected HT-29 cells treated with 2 mM DAC was also lower than the Ad-Hiwi-infected HT-29 cells without DAC treatment (Figure 5(f)). These findings indicated that DNA methylation knockdown blocked the proliferation promotion by the Hiwi overexpression in CRC cells.

Bottom Line: And the chemical inhibition of DNA methylation significantly restrained such proliferation promotion.In summary, we confirmed that Hiwi was overexpressed in CRC tissues and that the forced Hiwi overexpression promoted the proliferation and global DNA methylation of CRC cell lines.Our results imply for the first time that Hiwi promotes the proliferation of CRC cells via promoting global DNA methylation.

View Article: PubMed Central - PubMed

Affiliation: Department of Radiology, Provincial Hospital Affiliated to Shandong University, Jinan 250021, China.

ABSTRACT
Hiwi is well known for its role in stem cell renewal, maintaining the resting stage, and downregulating cell cycle of stem cells via RNA silencing. And Hiwi overexpression has been recognized in several types of cancers. In the present study, we examined the Hiwi expression in colorectal cancer (CRC) specimens in both mRNA and protein levels via real-time quantitative PCR, western blot assay, and immunohistochemical staining. Then we explored the role of Hiwi in the cancer cell proliferation and in the DNA methylation in human CRC Caro-2 and HT-29 cell lines. Results demonstrated that both mRNA and protein levels of Hiwi were significantly higher in 38 CRC tissues than in 38 peritumor tissues. Moreover, the Hiwi overexpression with an adenovirus vector significantly promoted the proliferation of Caro-2 and HT-29 cells, associated with significant increase in the global DNA methylation levels. And the chemical inhibition of DNA methylation significantly restrained such proliferation promotion. In summary, we confirmed that Hiwi was overexpressed in CRC tissues and that the forced Hiwi overexpression promoted the proliferation and global DNA methylation of CRC cell lines. Our results imply for the first time that Hiwi promotes the proliferation of CRC cells via promoting global DNA methylation.

No MeSH data available.


Related in: MedlinePlus