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Promoter hypermethylation mediated downregulation of FBP1 in human hepatocellular carcinoma and colon cancer.

Chen M, Zhang J, Li N, Qian Z, Zhu M, Li Q, Zheng J, Wang X, Shi G - PLoS ONE (2011)

Bottom Line: Restoring FBP1 expression in low expressed cells significantly inhibited cell growth and colony formation ability through the induction of G2-M phase cell cycle arrest.Moreover, the observed effects coincided with an increase in reactive oxygen species (ROS) generation.In summary, epigenetic inactivation of FBP1 is also common in human liver and colon cancer.

View Article: PubMed Central - PubMed

Affiliation: Department of Infectious Diseases, Huashan Hospital, Fudan University, Shanghai, China.

ABSTRACT
FBP1, fructose-1,6-bisphosphatase-1, a gluconeogenesis regulatory enzyme, catalyzes the hydrolysis of fructose 1,6-bisphosphate to fructose 6-phosphate and inorganic phosphate. The mechanism that it functions to antagonize glycolysis and was epigenetically inactivated through NF-kappaB pathway in gastric cancer has been reported. However, its role in the liver carcinogenesis still remains unknown. Here, we investigated the expression and DNA methylation of FBP1 in primary HCC and colon tumor. FBP1 was lowly expressed in 80% (8/10) human hepatocellular carcinoma, 66.7% (6/9) liver cancer cell lines and 100% (6/6) colon cancer cell lines, but was higher in paired adjacent non-tumor tissues and immortalized normal cell lines, which was well correlated with its promoter methylation status. Methylation was further detected in primary HCCs, gastric and colon tumor tissues, but none or occasionally in paired adjacent non-tumor tissues. Detailed methylation analysis of 29 CpG sites at a 327-bp promoter region by bisulfite genomic sequencing confirmed its methylation. FBP1 silencing could be reversed by chemical demethylation treatment with 5-aza-2'-deoxycytidine (Aza), indicating direct epigenetic silencing. Restoring FBP1 expression in low expressed cells significantly inhibited cell growth and colony formation ability through the induction of G2-M phase cell cycle arrest. Moreover, the observed effects coincided with an increase in reactive oxygen species (ROS) generation. In summary, epigenetic inactivation of FBP1 is also common in human liver and colon cancer. FBP1 appears to be a functional tumor suppressor involved in the liver and colon carcinogenesis.

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Pharmacological demethylation reversed FBP1 downregulation in human liver and other digestive cancers.(A) FBP1 expression in human liver and colon cancer cell lines and normal controls were determined by Real-Time RT-PCR. GAPDH was used to normalize the template amount. (B) Relative FBP1 expression before and after Aza treatment were determined by Real-Time RT-PCR. GAPDH was used to normalize the template amount. It was shown as average fold changes ± SD.
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pone-0025564-g001: Pharmacological demethylation reversed FBP1 downregulation in human liver and other digestive cancers.(A) FBP1 expression in human liver and colon cancer cell lines and normal controls were determined by Real-Time RT-PCR. GAPDH was used to normalize the template amount. (B) Relative FBP1 expression before and after Aza treatment were determined by Real-Time RT-PCR. GAPDH was used to normalize the template amount. It was shown as average fold changes ± SD.

Mentions: The expression of FBP1 was dramatically downregulated in 66.7% (6/9) human liver cancer cell lines including HepG2, BEL-7402, SMMC-7721, Sk-Hep1, MHCC-97H and MHCC-97L (Figure 1A) when compared to human normal liver cells Lo2. Similarly, downregualtion of FBP1 was also found in 100% (6/6) human colon cancer cell lines including HT29, SW480, SW620, HCT116, LoVo and RKO (Figure 1A) when compared to human normal adult colon tissue. In order to know whether downregulation of FBP1 should be attributed to DNA methylation, the expression of FBP1 in human liver and colon cancer cell lines before and after Aza treatment were determined by Real-Time RT-PCR. The expression of FBP1 was significantly upregulated in liver cancer cell lines HepG2, BEL-7402, SMMC-7721, Sk-Hep1, MHCC-97H and MHCC-97L (6/9, 66.7%) after Aza treatment (Figure 1B) and also in colon cancer cell lines HT29, SW620, LoVo and RKO (4/6, 66.7%) (Figure 1B), indicating that FBP1 is likely downregulated through promoter hypermethylation in human liver and colon cancer.


Promoter hypermethylation mediated downregulation of FBP1 in human hepatocellular carcinoma and colon cancer.

Chen M, Zhang J, Li N, Qian Z, Zhu M, Li Q, Zheng J, Wang X, Shi G - PLoS ONE (2011)

Pharmacological demethylation reversed FBP1 downregulation in human liver and other digestive cancers.(A) FBP1 expression in human liver and colon cancer cell lines and normal controls were determined by Real-Time RT-PCR. GAPDH was used to normalize the template amount. (B) Relative FBP1 expression before and after Aza treatment were determined by Real-Time RT-PCR. GAPDH was used to normalize the template amount. It was shown as average fold changes ± SD.
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Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3198434&req=5

pone-0025564-g001: Pharmacological demethylation reversed FBP1 downregulation in human liver and other digestive cancers.(A) FBP1 expression in human liver and colon cancer cell lines and normal controls were determined by Real-Time RT-PCR. GAPDH was used to normalize the template amount. (B) Relative FBP1 expression before and after Aza treatment were determined by Real-Time RT-PCR. GAPDH was used to normalize the template amount. It was shown as average fold changes ± SD.
Mentions: The expression of FBP1 was dramatically downregulated in 66.7% (6/9) human liver cancer cell lines including HepG2, BEL-7402, SMMC-7721, Sk-Hep1, MHCC-97H and MHCC-97L (Figure 1A) when compared to human normal liver cells Lo2. Similarly, downregualtion of FBP1 was also found in 100% (6/6) human colon cancer cell lines including HT29, SW480, SW620, HCT116, LoVo and RKO (Figure 1A) when compared to human normal adult colon tissue. In order to know whether downregulation of FBP1 should be attributed to DNA methylation, the expression of FBP1 in human liver and colon cancer cell lines before and after Aza treatment were determined by Real-Time RT-PCR. The expression of FBP1 was significantly upregulated in liver cancer cell lines HepG2, BEL-7402, SMMC-7721, Sk-Hep1, MHCC-97H and MHCC-97L (6/9, 66.7%) after Aza treatment (Figure 1B) and also in colon cancer cell lines HT29, SW620, LoVo and RKO (4/6, 66.7%) (Figure 1B), indicating that FBP1 is likely downregulated through promoter hypermethylation in human liver and colon cancer.

Bottom Line: Restoring FBP1 expression in low expressed cells significantly inhibited cell growth and colony formation ability through the induction of G2-M phase cell cycle arrest.Moreover, the observed effects coincided with an increase in reactive oxygen species (ROS) generation.In summary, epigenetic inactivation of FBP1 is also common in human liver and colon cancer.

View Article: PubMed Central - PubMed

Affiliation: Department of Infectious Diseases, Huashan Hospital, Fudan University, Shanghai, China.

ABSTRACT
FBP1, fructose-1,6-bisphosphatase-1, a gluconeogenesis regulatory enzyme, catalyzes the hydrolysis of fructose 1,6-bisphosphate to fructose 6-phosphate and inorganic phosphate. The mechanism that it functions to antagonize glycolysis and was epigenetically inactivated through NF-kappaB pathway in gastric cancer has been reported. However, its role in the liver carcinogenesis still remains unknown. Here, we investigated the expression and DNA methylation of FBP1 in primary HCC and colon tumor. FBP1 was lowly expressed in 80% (8/10) human hepatocellular carcinoma, 66.7% (6/9) liver cancer cell lines and 100% (6/6) colon cancer cell lines, but was higher in paired adjacent non-tumor tissues and immortalized normal cell lines, which was well correlated with its promoter methylation status. Methylation was further detected in primary HCCs, gastric and colon tumor tissues, but none or occasionally in paired adjacent non-tumor tissues. Detailed methylation analysis of 29 CpG sites at a 327-bp promoter region by bisulfite genomic sequencing confirmed its methylation. FBP1 silencing could be reversed by chemical demethylation treatment with 5-aza-2'-deoxycytidine (Aza), indicating direct epigenetic silencing. Restoring FBP1 expression in low expressed cells significantly inhibited cell growth and colony formation ability through the induction of G2-M phase cell cycle arrest. Moreover, the observed effects coincided with an increase in reactive oxygen species (ROS) generation. In summary, epigenetic inactivation of FBP1 is also common in human liver and colon cancer. FBP1 appears to be a functional tumor suppressor involved in the liver and colon carcinogenesis.

Show MeSH
Related in: MedlinePlus