1. Volume 26, Issue 10, Pages 2456-2465 (October 2018)
Disruption of Chromosomal Architecture of cox2 Locus Sensitizes Lung Cancer Cells to Radiotherapy 
Yuxiang Sun, Hui Dai, Shaopeng Chen, Yajun Zhang, Tao Wu, Xianbin Cao, Guoping Zhao, An Xu, Jun Wang, Lijun Wu 
Molecular Therapy 
Volume 26, Issue 10, Pages (October 2018)
DOI: /j.ymthe
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2. Figure 1
Cytotoxicity of Combination Treatment of Aspirin and Radiation
Cells were treated with radiation (5 Gy) with or without pre-treatment of 1 mM aspirin. (A) Cell death was measured by colony formation assay (*p < 0.05). (B) FACS analysis of A549 and NHLF cells stained with Annexin V-FITC and PI. The lower right and upper right quadrants represent apoptotic populations. These results were obtained from three independent experiments. (C) Quantification of FACS analysis (*p < 0.05). (D) Cell viability of A549 and NHLF cells was measured by MTT assay with 24 hr post-irradiation (***p < 0.001).
Molecular Therapy  , DOI: ( /j.ymthe )
Copyright © 2018 The Author(s) Terms and Conditions

3. Figure 2
The Crucial Role of cox2 in Aspirin-Mediated Radiosensitization of A549 Cells
(A) Western blot analysis of cox2 in A549 cells after radiation with or without pre-treatment of 1 mM aspirin (***p < 0.001). (B) MTT (*p < 0.05) and (C) colony formation assays were performed on A549 cells transfected with cox2 siRNA for 48 hr to determine the cell death and viability after radiation (***p < 0.001).
Molecular Therapy  , DOI: ( /j.ymthe )
Copyright © 2018 The Author(s) Terms and Conditions

4. Figure 3
Disruption of Chromatin Architecture Suppressed cox2 Expression in Response to Aspirin Treatment
(A) Schematic diagram of 3C analysis of the cox2 locus. (B) After aspirin treatment for 24 hr, the intra-chromosomal interactions between A1 and A2 sites were measured by nested PCR. Interaction frequency was determined by normalizing the 3C PCR signal relative to that of the positive control (***p < 0.001). (C) The specificity of PCR products was validated by DNA sequencing.
Molecular Therapy  , DOI: ( /j.ymthe )
Copyright © 2018 The Author(s) Terms and Conditions

5. Figure 4
The Role of p65 in Modulating Aspirin-Mediated Disruption of Chromatin Architecture
(A) Aspirin treatment inhibited nuclear translocation of p65. A549 cells were incubated with 1 mM aspirin for 24 hr. Levels of nuclear and cytosolic p65 were determined by western blot analysis. Values represent mean ± SD (***p < 0.001). (B) Left: aspirin treatment reduced binding of p65 on cox2 promoter and enhancer regions. After aspirin treatment aspirin for 24 hr, the cox2 promoter region (A1) and enhancer regions (A2) in the chromatin precipitate were amplified by PCR. Rabbit IgG was included as a negative control. Right: error bars represent SEM of three independent ChIP assays (*p < 0.05). (C) A549 cells were transfected with control siRNA or p65 siRNA prior to aspirin treatment, and then 3C assay was used to measure the interaction between A1 and A2 (*p < 0.05).
Molecular Therapy  , DOI: ( /j.ymthe )
Copyright © 2018 The Author(s) Terms and Conditions

6. Figure 5
The Role of Chromosome Conformation in Radiosensitivity by Regulating Apoptosis
(A and B) Expression levels of apoptotic proteins were examined by western blot analysis in A549 (A) and NHLF (B) cells (*p < 0.05). (C) Expression levels of apoptotic proteins in cells treated with or without cox2 siRNA were examined by western blot analysis (*p < 0.05). (D) FACS analysis of cells transfected with or without p65 siRNA prior to radiation. Cells were stained with PI.
Molecular Therapy  , DOI: ( /j.ymthe )
Copyright © 2018 The Author(s) Terms and Conditions

7. Figure 6
Schematic Diagram of the Molecular Mechanism of Aspirin-Mediated Radiosensitization of A549 Cells
Aspirin suppresses cox2 expression by disrupting chromatin architecture at the cox2 locus, sensitizing cells to radiation-induced apoptosis.
Molecular Therapy  , DOI: ( /j.ymthe )
Copyright © 2018 The Author(s) Terms and Conditions