Limits...
Ndrg2 promoter hypermethylation triggered by helicobacter pylori infection correlates with poor patients survival in human gastric carcinoma.

Ling ZQ, Ge MH, Lu XX, Han J, Wu YC, Liu X, Zhu X, Hong LL - Oncotarget (2015)

Bottom Line: We found that Ndrg2 promoter was frequently hypermethylated in gastric cancer cell lines and in 292 gastric tumor tissues.This resulted in down-regulation of Ndrg2 mRNA and protein.We found that H. pylori silenced Ndrg2 by activating the NF-κB pathway and up-regulating DNMT3b, promoting gastric cancer progression.

View Article: PubMed Central - PubMed

Affiliation: Zhejiang Cancer Research Institute, Zhejiang Province Cancer Hospital, Zhejiang Cancer Center, Hangzhou, China.

ABSTRACT
N-myc downstream regulated gene 2 (Ndrg2) is a candidate suppressor of cancer metastasis. We found that Ndrg2 promoter was frequently hypermethylated in gastric cancer cell lines and in 292 gastric tumor tissues. This resulted in down-regulation of Ndrg2 mRNA and protein. Ndrg2 promoter methylation was associated with H. pylori infection and worse prognosis of gastric cancer patients, which is an independent prognostic factor for the disease-free survival (DFS). We found that H. pylori silenced Ndrg2 by activating the NF-κB pathway and up-regulating DNMT3b, promoting gastric cancer progression. These findings uncover a previously unrecognized role for H. pylori infection in gastric cancer.

No MeSH data available.


Related in: MedlinePlus

H. pylori infection enhances DNMT activity and NDMT3b expression in vitroResults from triplicate determination of total DNMT activity and individual DNMT protein assays are shown. Error bars represent S.E.M. A: Fold change of DNMT activity in HP-treated SGC-7901 cells corresponds to the equivalent amount of untreated control cells with a set value of 1.0 for each point in time. * P < 0.05 vs. untreated controls. B: The total DNMT activity was measured using the EpiQuik™ DNA Methyltransferase Activity/Inhibition Assay (Epigentek), and the individual DNMT proteins of interest (DNMT1, DNMT3a, or DNMT3b) was measured using the Epiquik DNMT1, -3a, and -3b assay kits, respectively. Fold change of DNMT protein level in HP-treated SGC-7901 cells corresponds to the equivalent amount of untreated control cells at each point in time using the set value 1.0. * P < 0.05 vs. untreated controls. C: Results were normalized to the Gapdh level of each sample and are expressed fold change of DNMT mRNA level in HP-treated SGC-7901 cells corresponds to the equivalent amount of untreated control cells at each point in time using the set value 1.0. * P < 0.05 vs. untreated controls. D: Western blot analysis of DNMT expression change in HP-treated SGC-7901 cells. Human β-actin is used as internal reference. E: Effects of HP infection on Ndrg2 methylation in SGC-7901 cells at each point in time in vitro. * P < 0.05 vs. untreated controls (0 h). F: Effects of HP infection on Ndrg2 mRNA expression in SGC-7901 cells at each point in time in vitro. Ndrg2 mRNA level was determined by RT-PCR and adjusted for Gapdh. * P < 0.05 vs. untreated controls (0 h). G: Representative western blots of SGC-7901 cells treated with H. pylori or cells pretreated with H. pylori and then cocultured with DNTM3b siRNA were shown. Human β-actin is used as internal reference in each group of western blot analysis
© Copyright Policy - open-access
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC4480746&req=5

Figure 5: H. pylori infection enhances DNMT activity and NDMT3b expression in vitroResults from triplicate determination of total DNMT activity and individual DNMT protein assays are shown. Error bars represent S.E.M. A: Fold change of DNMT activity in HP-treated SGC-7901 cells corresponds to the equivalent amount of untreated control cells with a set value of 1.0 for each point in time. * P < 0.05 vs. untreated controls. B: The total DNMT activity was measured using the EpiQuik™ DNA Methyltransferase Activity/Inhibition Assay (Epigentek), and the individual DNMT proteins of interest (DNMT1, DNMT3a, or DNMT3b) was measured using the Epiquik DNMT1, -3a, and -3b assay kits, respectively. Fold change of DNMT protein level in HP-treated SGC-7901 cells corresponds to the equivalent amount of untreated control cells at each point in time using the set value 1.0. * P < 0.05 vs. untreated controls. C: Results were normalized to the Gapdh level of each sample and are expressed fold change of DNMT mRNA level in HP-treated SGC-7901 cells corresponds to the equivalent amount of untreated control cells at each point in time using the set value 1.0. * P < 0.05 vs. untreated controls. D: Western blot analysis of DNMT expression change in HP-treated SGC-7901 cells. Human β-actin is used as internal reference. E: Effects of HP infection on Ndrg2 methylation in SGC-7901 cells at each point in time in vitro. * P < 0.05 vs. untreated controls (0 h). F: Effects of HP infection on Ndrg2 mRNA expression in SGC-7901 cells at each point in time in vitro. Ndrg2 mRNA level was determined by RT-PCR and adjusted for Gapdh. * P < 0.05 vs. untreated controls (0 h). G: Representative western blots of SGC-7901 cells treated with H. pylori or cells pretreated with H. pylori and then cocultured with DNTM3b siRNA were shown. Human β-actin is used as internal reference in each group of western blot analysis

Mentions: After 6 h of H. pylori treatment, incubation of SGC-7901 cells led to a rapid and significant increase in total DNMT activity by 29.8% as compared to the controls. At later points in time, DNMT activity was stably increased by approximately 10% for 12h, 24 h, 48 h time point, and 5% for 72 h time point, respectively, where a total increase of more than 60% was observed (Figure 5A). No significant differences were detected for DNMT1 or DNMT3a protein levels between SGC-7901 cells with and those without H. pylori treatment at every time point (Figure 5B). The average DNMT3b protein level for the SGC-7901 cells with H. pylori treated was 1.3-fold at 6 h and reached a maximum of 1.8-fold at 48 h, compared to the controls (Figure 5B).


Ndrg2 promoter hypermethylation triggered by helicobacter pylori infection correlates with poor patients survival in human gastric carcinoma.

Ling ZQ, Ge MH, Lu XX, Han J, Wu YC, Liu X, Zhu X, Hong LL - Oncotarget (2015)

H. pylori infection enhances DNMT activity and NDMT3b expression in vitroResults from triplicate determination of total DNMT activity and individual DNMT protein assays are shown. Error bars represent S.E.M. A: Fold change of DNMT activity in HP-treated SGC-7901 cells corresponds to the equivalent amount of untreated control cells with a set value of 1.0 for each point in time. * P < 0.05 vs. untreated controls. B: The total DNMT activity was measured using the EpiQuik™ DNA Methyltransferase Activity/Inhibition Assay (Epigentek), and the individual DNMT proteins of interest (DNMT1, DNMT3a, or DNMT3b) was measured using the Epiquik DNMT1, -3a, and -3b assay kits, respectively. Fold change of DNMT protein level in HP-treated SGC-7901 cells corresponds to the equivalent amount of untreated control cells at each point in time using the set value 1.0. * P < 0.05 vs. untreated controls. C: Results were normalized to the Gapdh level of each sample and are expressed fold change of DNMT mRNA level in HP-treated SGC-7901 cells corresponds to the equivalent amount of untreated control cells at each point in time using the set value 1.0. * P < 0.05 vs. untreated controls. D: Western blot analysis of DNMT expression change in HP-treated SGC-7901 cells. Human β-actin is used as internal reference. E: Effects of HP infection on Ndrg2 methylation in SGC-7901 cells at each point in time in vitro. * P < 0.05 vs. untreated controls (0 h). F: Effects of HP infection on Ndrg2 mRNA expression in SGC-7901 cells at each point in time in vitro. Ndrg2 mRNA level was determined by RT-PCR and adjusted for Gapdh. * P < 0.05 vs. untreated controls (0 h). G: Representative western blots of SGC-7901 cells treated with H. pylori or cells pretreated with H. pylori and then cocultured with DNTM3b siRNA were shown. Human β-actin is used as internal reference in each group of western blot analysis
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4480746&req=5

Figure 5: H. pylori infection enhances DNMT activity and NDMT3b expression in vitroResults from triplicate determination of total DNMT activity and individual DNMT protein assays are shown. Error bars represent S.E.M. A: Fold change of DNMT activity in HP-treated SGC-7901 cells corresponds to the equivalent amount of untreated control cells with a set value of 1.0 for each point in time. * P < 0.05 vs. untreated controls. B: The total DNMT activity was measured using the EpiQuik™ DNA Methyltransferase Activity/Inhibition Assay (Epigentek), and the individual DNMT proteins of interest (DNMT1, DNMT3a, or DNMT3b) was measured using the Epiquik DNMT1, -3a, and -3b assay kits, respectively. Fold change of DNMT protein level in HP-treated SGC-7901 cells corresponds to the equivalent amount of untreated control cells at each point in time using the set value 1.0. * P < 0.05 vs. untreated controls. C: Results were normalized to the Gapdh level of each sample and are expressed fold change of DNMT mRNA level in HP-treated SGC-7901 cells corresponds to the equivalent amount of untreated control cells at each point in time using the set value 1.0. * P < 0.05 vs. untreated controls. D: Western blot analysis of DNMT expression change in HP-treated SGC-7901 cells. Human β-actin is used as internal reference. E: Effects of HP infection on Ndrg2 methylation in SGC-7901 cells at each point in time in vitro. * P < 0.05 vs. untreated controls (0 h). F: Effects of HP infection on Ndrg2 mRNA expression in SGC-7901 cells at each point in time in vitro. Ndrg2 mRNA level was determined by RT-PCR and adjusted for Gapdh. * P < 0.05 vs. untreated controls (0 h). G: Representative western blots of SGC-7901 cells treated with H. pylori or cells pretreated with H. pylori and then cocultured with DNTM3b siRNA were shown. Human β-actin is used as internal reference in each group of western blot analysis
Mentions: After 6 h of H. pylori treatment, incubation of SGC-7901 cells led to a rapid and significant increase in total DNMT activity by 29.8% as compared to the controls. At later points in time, DNMT activity was stably increased by approximately 10% for 12h, 24 h, 48 h time point, and 5% for 72 h time point, respectively, where a total increase of more than 60% was observed (Figure 5A). No significant differences were detected for DNMT1 or DNMT3a protein levels between SGC-7901 cells with and those without H. pylori treatment at every time point (Figure 5B). The average DNMT3b protein level for the SGC-7901 cells with H. pylori treated was 1.3-fold at 6 h and reached a maximum of 1.8-fold at 48 h, compared to the controls (Figure 5B).

Bottom Line: We found that Ndrg2 promoter was frequently hypermethylated in gastric cancer cell lines and in 292 gastric tumor tissues.This resulted in down-regulation of Ndrg2 mRNA and protein.We found that H. pylori silenced Ndrg2 by activating the NF-κB pathway and up-regulating DNMT3b, promoting gastric cancer progression.

View Article: PubMed Central - PubMed

Affiliation: Zhejiang Cancer Research Institute, Zhejiang Province Cancer Hospital, Zhejiang Cancer Center, Hangzhou, China.

ABSTRACT
N-myc downstream regulated gene 2 (Ndrg2) is a candidate suppressor of cancer metastasis. We found that Ndrg2 promoter was frequently hypermethylated in gastric cancer cell lines and in 292 gastric tumor tissues. This resulted in down-regulation of Ndrg2 mRNA and protein. Ndrg2 promoter methylation was associated with H. pylori infection and worse prognosis of gastric cancer patients, which is an independent prognostic factor for the disease-free survival (DFS). We found that H. pylori silenced Ndrg2 by activating the NF-κB pathway and up-regulating DNMT3b, promoting gastric cancer progression. These findings uncover a previously unrecognized role for H. pylori infection in gastric cancer.

No MeSH data available.


Related in: MedlinePlus