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Correlating Gene-specific DNA Methylation Changes with Expression and Transcriptional Activity of Astrocytic KCNJ10 (Kir4.1)
Published on: September 26, 2015
Axin gene methylation status correlates with radiosensitivity of lung cancer cells
Lian-He Yang1, Yang Han, Guang Li
1Department of Pathology, First Affiliated Hospital and College of Basic Medical Sciences, China Medical University, Shenyang, Liaoning, China.
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.
Insights
X-ray irradiation enhances Axin expression in lung cancer cells by demethylating the gene, inhibiting tumor growth. Axin gene methylation status predicts tumor radiosensitivity.
Area of Science:
- Oncology
- Molecular Biology
- Radiotherapy
Background:
- Axin1 (Axin) is frequently down-regulated in lung cancer.
- X-ray irradiation was previously shown to increase Axin expression and inhibit lung cancer cells, but the underlying mechanisms were unclear.
Purpose of the Study:
- To investigate the role of Axin gene methylation in lung cancer response to X-ray irradiation.
- To elucidate the molecular mechanisms by which X-ray irradiation affects Axin expression and lung cancer cell behavior.
Main Methods:
- Assessed Axin gene methylation status in four lung cancer cell lines with and without X-ray treatment.
- Quantified Axin expression using real-time PCR and protein levels via Western blot.
- Evaluated biological behaviors including cell proliferation, invasion, and apoptosis using various assays and xenograft models.
Main Results:
- Hypermethylation of the Axin gene was found in 2 of 4 cell lines, inversely correlating with Axin expression.
- X-ray treatment significantly upregulated Axin in hypermethylated cell lines (H446, H157) but not in unmethylated ones (LTE, H460).
- X-ray irradiation reduced colony formation, xenograft growth, and invasiveness, while increasing apoptosis in hypermethylated H157 cells, with greater effects compared to unmethylated LTE cells.
Conclusions:
- X-ray-induced lung cancer cell inhibition is likely mediated by increased Axin expression through DNA demethylation and histone acetylation.
- The methylation status of the Axin gene influences tumor radiosensitivity, potentially serving as a predictive biomarker.
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