Limits...
Axin gene methylation status correlates with radiosensitivity of lung cancer cells.

Yang LH, Han Y, Li G, Xu HT, Jiang GY, Miao Y, Zhang XP, Zhao HY, Xu ZF, Stoecker M, Wang E, Xu K, Wang EH - BMC Cancer (2013)

Bottom Line: The mechanisms, however, were not clear.Real-time PCR was used to quantify the expression of Axin, and western blot analysis was applied to measure protein levels of Axin, β-catenin, Cyclin D1, MMP-7, DNMTS, MeCP2 and acetylated histones.Hypermethylated Axin gene was detected in 2 of 4 cell lines, and it correlated inversely with Axin expression.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Pathology, First Affiliated Hospital and College of Basic Medical Sciences, China Medical University, Shenyang, Liaoning, China.

ABSTRACT

Background: We previously reported that Axin1 (Axin) is down-regulated in many cases of lung cancer, and X-ray irradiation increased Axin expression and inhibited lung cancer cells. The mechanisms, however, were not clear.

Methods: Four lung cancer cell lines were used to detect the methylation status of Axin with or without X-ray treatment. Real-time PCR was used to quantify the expression of Axin, and western blot analysis was applied to measure protein levels of Axin, β-catenin, Cyclin D1, MMP-7, DNMTS, MeCP2 and acetylated histones. Flow cytometric analysis, colony formation assay, transwell assay and xenograft growth experiment were used to study the biological behavior of the cells with hypermethylated or unmethylated Axin gene after X-ray treatment.

Results: Hypermethylated Axin gene was detected in 2 of 4 cell lines, and it correlated inversely with Axin expression. X-ray treatment significantly up-regulated Axin expression in H446 and H157 cells, which possess intrinsic hypermethylation of the Axin gene (P<0.01), but did not show up-regulation in LTE and H460 cells, which have unmethylated Axin gene. 2Gy X-ray significantly reduced colony formation (from 71% to 10.5%) in H157 cells, while the reduction was lower in LTE cells (from 71% to 20%). After X-ray irradiation, xenograft growth was significantly decreased in H157 cells (from 1.15 g to 0.28 g) in comparison with LTE cells (from 1.06 g to 0.65 g). Significantly decreased cell invasiveness and increased apoptosis were also observed in H157 cells treated with X-ray irradiation (P<0.01). Down-regulation of DNMTs and MeCP2 and up-regulation of acetylated histones could be detected in lung cancer cells.

Conclusions: X-ray-induced inhibition of lung cancer cells may be mediated by enhanced expression of Axin via genomic DNA demethylation and histone acetylation. Lung cancer cells with a different methylation status of the Axin gene showed different radiosensitivity, suggesting that the methylation status of the Axin gene may be one important factor to predict radiosensitivity of the tumor.

Show MeSH

Related in: MedlinePlus

The effect of X-ray irradiation on DNMTs, MeCP2, acetylated histones and factors of the Wnt signaling pathway. Western blot analysis shows the effects of X-ray irradiation on the expression of DNMTs, MeCP2, acetylated histones, Axin, β-catenin, Cyclin D1 and MMP-7 at 24h after X-ray treatment. DNMT1, DNMT3B and MeCP2 are significantly down-regulated in H157 cells, and acetylated histone H3 and H4 are up-regulated in H157 cells (A, B) is the histogram of A. DNMT 1 and 3B are also down-regulated significantly in LTE cells. Slightly decreased MeCP2 expression and up-regulation of acetylated H3, H4 are noted in LTE cells (C and D), with the degree of change less significant than in H157 cells. Increased Axin expression and decreased β-catenin, Cyclin D1 and MMP-7 expression are noted in the H157 cell line after X-ray irradiation (E and F), but no significant change in these factors are detected in LTE cells (G and H). Note a dose dependent pattern of the changes in both cell lines, with more prominent changes in the H157 than in the LTE cell line. * P<0.05, ** P<0.01.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC3750238&req=5

Figure 3: The effect of X-ray irradiation on DNMTs, MeCP2, acetylated histones and factors of the Wnt signaling pathway. Western blot analysis shows the effects of X-ray irradiation on the expression of DNMTs, MeCP2, acetylated histones, Axin, β-catenin, Cyclin D1 and MMP-7 at 24h after X-ray treatment. DNMT1, DNMT3B and MeCP2 are significantly down-regulated in H157 cells, and acetylated histone H3 and H4 are up-regulated in H157 cells (A, B) is the histogram of A. DNMT 1 and 3B are also down-regulated significantly in LTE cells. Slightly decreased MeCP2 expression and up-regulation of acetylated H3, H4 are noted in LTE cells (C and D), with the degree of change less significant than in H157 cells. Increased Axin expression and decreased β-catenin, Cyclin D1 and MMP-7 expression are noted in the H157 cell line after X-ray irradiation (E and F), but no significant change in these factors are detected in LTE cells (G and H). Note a dose dependent pattern of the changes in both cell lines, with more prominent changes in the H157 than in the LTE cell line. * P<0.05, ** P<0.01.

Mentions: It has been reported that X-ray irradiation could induce demethylation by inhibiting DNMTs and MeCP2 [14-21]. DNA methylation is regulated by DNMTs, a family of enzymes catalyzing transfer of methyl groups to genomic DNA [13]. We examined the protein levels of DNMT1 and 3B at 24 hours after 1 Gy and 2 Gy X-ray irradiation, respectively, in two NSCLC cell lines: H157 (hypermethylated Axin gene) and LTE (unmethylated Axin gene). Both DNMT1 and DNMT3B were significantly down-regulated in the two cell lines (Figure 3A-D) (P<0.01), with more significant effects seen in the H157 cell line than in the other.


Axin gene methylation status correlates with radiosensitivity of lung cancer cells.

Yang LH, Han Y, Li G, Xu HT, Jiang GY, Miao Y, Zhang XP, Zhao HY, Xu ZF, Stoecker M, Wang E, Xu K, Wang EH - BMC Cancer (2013)

The effect of X-ray irradiation on DNMTs, MeCP2, acetylated histones and factors of the Wnt signaling pathway. Western blot analysis shows the effects of X-ray irradiation on the expression of DNMTs, MeCP2, acetylated histones, Axin, β-catenin, Cyclin D1 and MMP-7 at 24h after X-ray treatment. DNMT1, DNMT3B and MeCP2 are significantly down-regulated in H157 cells, and acetylated histone H3 and H4 are up-regulated in H157 cells (A, B) is the histogram of A. DNMT 1 and 3B are also down-regulated significantly in LTE cells. Slightly decreased MeCP2 expression and up-regulation of acetylated H3, H4 are noted in LTE cells (C and D), with the degree of change less significant than in H157 cells. Increased Axin expression and decreased β-catenin, Cyclin D1 and MMP-7 expression are noted in the H157 cell line after X-ray irradiation (E and F), but no significant change in these factors are detected in LTE cells (G and H). Note a dose dependent pattern of the changes in both cell lines, with more prominent changes in the H157 than in the LTE cell line. * P<0.05, ** P<0.01.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3750238&req=5

Figure 3: The effect of X-ray irradiation on DNMTs, MeCP2, acetylated histones and factors of the Wnt signaling pathway. Western blot analysis shows the effects of X-ray irradiation on the expression of DNMTs, MeCP2, acetylated histones, Axin, β-catenin, Cyclin D1 and MMP-7 at 24h after X-ray treatment. DNMT1, DNMT3B and MeCP2 are significantly down-regulated in H157 cells, and acetylated histone H3 and H4 are up-regulated in H157 cells (A, B) is the histogram of A. DNMT 1 and 3B are also down-regulated significantly in LTE cells. Slightly decreased MeCP2 expression and up-regulation of acetylated H3, H4 are noted in LTE cells (C and D), with the degree of change less significant than in H157 cells. Increased Axin expression and decreased β-catenin, Cyclin D1 and MMP-7 expression are noted in the H157 cell line after X-ray irradiation (E and F), but no significant change in these factors are detected in LTE cells (G and H). Note a dose dependent pattern of the changes in both cell lines, with more prominent changes in the H157 than in the LTE cell line. * P<0.05, ** P<0.01.
Mentions: It has been reported that X-ray irradiation could induce demethylation by inhibiting DNMTs and MeCP2 [14-21]. DNA methylation is regulated by DNMTs, a family of enzymes catalyzing transfer of methyl groups to genomic DNA [13]. We examined the protein levels of DNMT1 and 3B at 24 hours after 1 Gy and 2 Gy X-ray irradiation, respectively, in two NSCLC cell lines: H157 (hypermethylated Axin gene) and LTE (unmethylated Axin gene). Both DNMT1 and DNMT3B were significantly down-regulated in the two cell lines (Figure 3A-D) (P<0.01), with more significant effects seen in the H157 cell line than in the other.

Bottom Line: The mechanisms, however, were not clear.Real-time PCR was used to quantify the expression of Axin, and western blot analysis was applied to measure protein levels of Axin, β-catenin, Cyclin D1, MMP-7, DNMTS, MeCP2 and acetylated histones.Hypermethylated Axin gene was detected in 2 of 4 cell lines, and it correlated inversely with Axin expression.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Pathology, First Affiliated Hospital and College of Basic Medical Sciences, China Medical University, Shenyang, Liaoning, China.

ABSTRACT

Background: We previously reported that Axin1 (Axin) is down-regulated in many cases of lung cancer, and X-ray irradiation increased Axin expression and inhibited lung cancer cells. The mechanisms, however, were not clear.

Methods: Four lung cancer cell lines were used to detect the methylation status of Axin with or without X-ray treatment. Real-time PCR was used to quantify the expression of Axin, and western blot analysis was applied to measure protein levels of Axin, β-catenin, Cyclin D1, MMP-7, DNMTS, MeCP2 and acetylated histones. Flow cytometric analysis, colony formation assay, transwell assay and xenograft growth experiment were used to study the biological behavior of the cells with hypermethylated or unmethylated Axin gene after X-ray treatment.

Results: Hypermethylated Axin gene was detected in 2 of 4 cell lines, and it correlated inversely with Axin expression. X-ray treatment significantly up-regulated Axin expression in H446 and H157 cells, which possess intrinsic hypermethylation of the Axin gene (P<0.01), but did not show up-regulation in LTE and H460 cells, which have unmethylated Axin gene. 2Gy X-ray significantly reduced colony formation (from 71% to 10.5%) in H157 cells, while the reduction was lower in LTE cells (from 71% to 20%). After X-ray irradiation, xenograft growth was significantly decreased in H157 cells (from 1.15 g to 0.28 g) in comparison with LTE cells (from 1.06 g to 0.65 g). Significantly decreased cell invasiveness and increased apoptosis were also observed in H157 cells treated with X-ray irradiation (P<0.01). Down-regulation of DNMTs and MeCP2 and up-regulation of acetylated histones could be detected in lung cancer cells.

Conclusions: X-ray-induced inhibition of lung cancer cells may be mediated by enhanced expression of Axin via genomic DNA demethylation and histone acetylation. Lung cancer cells with a different methylation status of the Axin gene showed different radiosensitivity, suggesting that the methylation status of the Axin gene may be one important factor to predict radiosensitivity of the tumor.

Show MeSH
Related in: MedlinePlus