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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.

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The comparison of xenograft tumor growth between the cell line with a hypermethylated Axin gene (H157) and the cell line with an unmethylated Axin gene (LTE) after X-ray treatment. Figure A shows asignificant decrease in tumor mass in the mice transplanted with X-ray treated cancer cells, particularly in the mice with treated H157 cells in comparison with each control. Figure B shows the curves of time dependent tumor growth. Note X-ray treatment significantly suppresses the growth of tumor xenografts, with a more significant effect noted in H157 cells than in LTE cells.
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Figure 5: The comparison of xenograft tumor growth between the cell line with a hypermethylated Axin gene (H157) and the cell line with an unmethylated Axin gene (LTE) after X-ray treatment. Figure A shows asignificant decrease in tumor mass in the mice transplanted with X-ray treated cancer cells, particularly in the mice with treated H157 cells in comparison with each control. Figure B shows the curves of time dependent tumor growth. Note X-ray treatment significantly suppresses the growth of tumor xenografts, with a more significant effect noted in H157 cells than in LTE cells.

Mentions: H157 and LTE cells with or without X-ray irradiation (2 Gy) were inoculated into nude mice, respectively, and the tumors were completely excised 4 weeks later (Figure 5A). The weight of tumor was markedly reduced in H157 cells receiving irradiation from 1.15 ± 0.37g to 0.28 ± 0.08 g (P<0.01), and the size of tumor was decreased from 1.77 ± 0.63 cm3 to 0.44 ± 0.12 cm3 (P<0.01). The rate of tumor inhibition in the H157 cell line (76.63%, weight) was significantly higher than in the LTE cell line (38.5%, from 1.06 ± 0.22 g to 0.65 ± 0.21 g, P<0.01). There is no statistically significant difference in the rates of xenograft growth between the 2 cell lines without irradiation in tumor size and growth, but the difference is statistically significant between H157 cells with irradiation and LTE cells with irradiation in tumor size and growth (P<0.01) (Figure 5B). While X-ray irradiation showed the suppression of tumor growth in both cell lines, the extent of suppression in H157 cells was much more prominent than in LTE cells.


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 comparison of xenograft tumor growth between the cell line with a hypermethylated Axin gene (H157) and the cell line with an unmethylated Axin gene (LTE) after X-ray treatment. Figure A shows asignificant decrease in tumor mass in the mice transplanted with X-ray treated cancer cells, particularly in the mice with treated H157 cells in comparison with each control. Figure B shows the curves of time dependent tumor growth. Note X-ray treatment significantly suppresses the growth of tumor xenografts, with a more significant effect noted in H157 cells than in LTE cells.
© Copyright Policy - open-access
Related In: Results  -  Collection

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

Figure 5: The comparison of xenograft tumor growth between the cell line with a hypermethylated Axin gene (H157) and the cell line with an unmethylated Axin gene (LTE) after X-ray treatment. Figure A shows asignificant decrease in tumor mass in the mice transplanted with X-ray treated cancer cells, particularly in the mice with treated H157 cells in comparison with each control. Figure B shows the curves of time dependent tumor growth. Note X-ray treatment significantly suppresses the growth of tumor xenografts, with a more significant effect noted in H157 cells than in LTE cells.
Mentions: H157 and LTE cells with or without X-ray irradiation (2 Gy) were inoculated into nude mice, respectively, and the tumors were completely excised 4 weeks later (Figure 5A). The weight of tumor was markedly reduced in H157 cells receiving irradiation from 1.15 ± 0.37g to 0.28 ± 0.08 g (P<0.01), and the size of tumor was decreased from 1.77 ± 0.63 cm3 to 0.44 ± 0.12 cm3 (P<0.01). The rate of tumor inhibition in the H157 cell line (76.63%, weight) was significantly higher than in the LTE cell line (38.5%, from 1.06 ± 0.22 g to 0.65 ± 0.21 g, P<0.01). There is no statistically significant difference in the rates of xenograft growth between the 2 cell lines without irradiation in tumor size and growth, but the difference is statistically significant between H157 cells with irradiation and LTE cells with irradiation in tumor size and growth (P<0.01) (Figure 5B). While X-ray irradiation showed the suppression of tumor growth in both cell lines, the extent of suppression in H157 cells was much more prominent than in LTE cells.

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