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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

Overexpression of Hiwi by adenovirus vector at a series of MOI. (a) and (b): the expression of Hiwi at mRNA (a) or protein (b) level in the Caco-2 cells, which were infected with 0, 1, or 10 MOI Ad-Hiwi or Ad-RFP. The protein level of Hiwi was expressed as the percent level to GAPDH (b). (c) The expression of Hiwi in the Caco-2 cells, which were infected with 0, 1, or 10 MOI Ad-Hiwi by immunohistochemistry. (d) The RFP fluorescence microscopy of Caco-2 cells infected by Ad-RFP at 0, 1, or 10 MOI. Statistical significance was shown as ∗p < 0.05, ∗∗p < 0.01, and ∗∗∗p < 0.001; ns, no significance.
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fig2: Overexpression of Hiwi by adenovirus vector at a series of MOI. (a) and (b): the expression of Hiwi at mRNA (a) or protein (b) level in the Caco-2 cells, which were infected with 0, 1, or 10 MOI Ad-Hiwi or Ad-RFP. The protein level of Hiwi was expressed as the percent level to GAPDH (b). (c) The expression of Hiwi in the Caco-2 cells, which were infected with 0, 1, or 10 MOI Ad-Hiwi by immunohistochemistry. (d) The RFP fluorescence microscopy of Caco-2 cells infected by Ad-RFP at 0, 1, or 10 MOI. Statistical significance was shown as ∗p < 0.05, ∗∗p < 0.01, and ∗∗∗p < 0.001; ns, no significance.

Mentions: To investigate the role of Hiwi overexpression in CRC cells, we used an adenovirus vector carrying Hiwi gene to overexpress the Hiwi in Caco-2 or HT-29 cells. Whole coding sequence of Hiwi was cloned into adenovirus vector, and the recombinant adenovirus harboring Hiwi coding sequence (named as Ad-Hiwi) was rescued and determined. Red fluorescence protein gene (RFP) was also cloned into adenovirus vector as control (Ad-RFP). As shown in Figure 2(a), the Hiwi mRNA level in the Ad-Hiwi-infected Caco-2 cells was significantly higher than in the Ad-RFP-infected Caco-2 cells at 1 or 10 MOI. The protein level was also significantly higher in the Caco-2 cells which were infected with the Ad-Hiwi virus than in those infected with the Ad-RFP virus, by western blot analysis (Figure 2(b)). In addition, the immunohistochemistry analysis also indicated a high Hiwi expression in Caco-2 cells which were infected with 1 or 10 MOI Ad-Hiwi; there was a higher level of green fluorescence in the Ad-Hiwi-infected Caco-2 cells (Figure 2(c)), whereas the Ad-RFP infection caused a higher level of red fluorescence in Caco-2 cells at 1 or 10 MOI (Figure 2(c)).


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)

Overexpression of Hiwi by adenovirus vector at a series of MOI. (a) and (b): the expression of Hiwi at mRNA (a) or protein (b) level in the Caco-2 cells, which were infected with 0, 1, or 10 MOI Ad-Hiwi or Ad-RFP. The protein level of Hiwi was expressed as the percent level to GAPDH (b). (c) The expression of Hiwi in the Caco-2 cells, which were infected with 0, 1, or 10 MOI Ad-Hiwi by immunohistochemistry. (d) The RFP fluorescence microscopy of Caco-2 cells infected by Ad-RFP at 0, 1, or 10 MOI. 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

Show All Figures
getmorefigures.php?uid=PMC4556077&req=5

fig2: Overexpression of Hiwi by adenovirus vector at a series of MOI. (a) and (b): the expression of Hiwi at mRNA (a) or protein (b) level in the Caco-2 cells, which were infected with 0, 1, or 10 MOI Ad-Hiwi or Ad-RFP. The protein level of Hiwi was expressed as the percent level to GAPDH (b). (c) The expression of Hiwi in the Caco-2 cells, which were infected with 0, 1, or 10 MOI Ad-Hiwi by immunohistochemistry. (d) The RFP fluorescence microscopy of Caco-2 cells infected by Ad-RFP at 0, 1, or 10 MOI. Statistical significance was shown as ∗p < 0.05, ∗∗p < 0.01, and ∗∗∗p < 0.001; ns, no significance.
Mentions: To investigate the role of Hiwi overexpression in CRC cells, we used an adenovirus vector carrying Hiwi gene to overexpress the Hiwi in Caco-2 or HT-29 cells. Whole coding sequence of Hiwi was cloned into adenovirus vector, and the recombinant adenovirus harboring Hiwi coding sequence (named as Ad-Hiwi) was rescued and determined. Red fluorescence protein gene (RFP) was also cloned into adenovirus vector as control (Ad-RFP). As shown in Figure 2(a), the Hiwi mRNA level in the Ad-Hiwi-infected Caco-2 cells was significantly higher than in the Ad-RFP-infected Caco-2 cells at 1 or 10 MOI. The protein level was also significantly higher in the Caco-2 cells which were infected with the Ad-Hiwi virus than in those infected with the Ad-RFP virus, by western blot analysis (Figure 2(b)). In addition, the immunohistochemistry analysis also indicated a high Hiwi expression in Caco-2 cells which were infected with 1 or 10 MOI Ad-Hiwi; there was a higher level of green fluorescence in the Ad-Hiwi-infected Caco-2 cells (Figure 2(c)), whereas the Ad-RFP infection caused a higher level of red fluorescence in Caco-2 cells at 1 or 10 MOI (Figure 2(c)).

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