Skip to main content

Quantification of DNA double-strand break induced by radiation in cervix cancer cells: in vitro study

  • Sothing Vashum (a1), Rabi Raja Singh I (a1), Saikat Das (a1), Mohammed Azharuddin KO (a2) and Prabhakaran Vasudevan (a2)...

DNA double-strand break (DSB) results in the phosphorylation of the protein, H.2AX histone. In this study, the effect of radiotherapy and chemotherapy on DNA DSB in cervical cancer cells is analysed by the phosphorylation of the protein.


The cervical cancer cells (HeLa cells) were cultured and exposed to ionising radiation. Radiation sensitivity was measured by clonogenic survival fraction after exposing to ionising radiation. Since the phosphorylation of H.2AX declines with time, the DNA damage was quantified at different time points: 1 hour, 3 hours and 1 week after exposed to the radiation. The analysis of γ-H.2AX was done by Western-blot technique. The protein expression was observed at different dose of radiation and combination of both radiation and paclitaxel.


Low-dose hypersensitivity was observed. By 1 week after radiation at 0·5, 0·8 and 2 Gy, there was no expression of phosphorylated H.2AX. Previous experiments on the expression of phosphorylated H.2AX (γ-H.2AX) in terms of foci analysis was found to peak at 1 hour and subsequently decline with time. In cells treated with the DNA damaging agents, the expression of phosphorylated H.2AX decreases in a dose-dependent manner when treated with radiation alone. However, when combined with paclitaxel, at 0·5 Gy, the expression peaked and reduces at 0·8 Gy and slightly elevated at 2 Gy.


In this study, the peak phosphorylation was observed at 3 hour post irradiation indicating that DSBs are still left unrepaired.

Corresponding author
Authors for correspondence: Sothing Vashum, Department of Radiation Therapy, Christian Medical College and Hospital, Ida Scudder Road, Vellore – 632004, Tamil Nadu, India. Tel: +91-8754867454. E-mail: Prabhakaran Vasudevan, Department of Neurosciences, Christian Medical College and Hospital, Ida Scudder Road, Vellore – 632004, Tamil Nadu, India. Tel: +91-4162282701. E-mail:
Hide All
1. Kuo, LJ, Yang, L. “Gamma-H2AX - a novel biomarker for DNA double-strand breaks.” In vivo 22 3 (2008): 305-9. Radiobiology Laboratory, California Pacific Medical Center Research Institute, San Francisco, CA 94118; 2St. Mary’s Medical Center, San Francisco, CA 94117, U.S.A.
2. Rogakuo, EP, Pilch, DR, Orr, AH, Ivanova, VS, Boriner, WM. DNA double strand breaks induce histone H2AX phosphorylation on serine 139. J Biol Chem 1998; 273 (10): 58585868.
3. Paull, TT, Rogakou, EP, Yamazaki, V, Kirchgessner, CU, Gellert, M, Bonner, WM. A critical role for histone H2AX in recruitment of repair factors to nuclear foci after DNA damage. Curr Biol 2000; 10 (15): 886895.
4. Fuhrman, CB, Kilgore, J, LaCoursiere, YD et al. Radiosensitization of cervical cancer cells via double-strand DNA break repair inhibition. Gynecol Oncol 2008; 110 (1): 9398.
5. MacPhail, SH, Banáth, JP, Yu, TY, Chu, EH, Lambur, H, Olive, PL. Expression of phosphorylated histone H2AX in cultured cell lines following exposure to X-rays. Int J Radiat Biol 2003; 79 (5): 351358.
6. Goutham, HV, Mumbrekar, KD, Vadhiraja, BM et al. DNA double-strand break analysis by γ-H2AX foci: a useful method for determining the overreactors to radiation-induced acute reactions among head-and-neck cancer patients. Int J Radiat Oncol Biol Phys 2012; 84 (5): e607e612. Epub Jul 24, 2012 .
7. Lowry, OH, Rosebrough, NJ, Farr, A, Randall, RJ. Protein measurement with the Folin phenol reagent. J Biol Chem 1951; 193 (1): 265275.
8. ImageJ Software National Institutes of Health, Bethesda, Maryland, USA.
9. Das, S, Singh, R, George, D, Vijaykumar, TS, John, S. Radiobiological response of cervical cancer cell line in low dose region: evidence of low dose hypersensitivity (HRS) and induced radioresistance (IRR).
Recommend this journal

Email your librarian or administrator to recommend adding this journal to your organisation's collection.

Journal of Radiotherapy in Practice
  • ISSN: 1460-3969
  • EISSN: 1467-1131
  • URL: /core/journals/journal-of-radiotherapy-in-practice
Please enter your name
Please enter a valid email address
Who would you like to send this to? *



Full text views

Total number of HTML views: 0
Total number of PDF views: 0 *
Loading metrics...

Abstract views

Total abstract views: 0 *
Loading metrics...

* Views captured on Cambridge Core between <date>. This data will be updated every 24 hours.

Usage data cannot currently be displayed