Molecules and Cells

Korean Society for Molecular and Cellular Biology (한국분자세포생물학회)
권호 표지

Low-Dose Radiation Stimulates the Proliferation of Normal Human Lung Fibroblasts Via a Transient Activation of Raf and Akt

  • Kim, Cha Soon (Radiation Health Research Institute, Korea Hydro and Nuclear Power Co., Ltd.) ;
  • Kim, Jin Kyoung (Radiation Health Research Institute, Korea Hydro and Nuclear Power Co., Ltd.) ;
  • Nam, Seon Young (Radiation Health Research Institute, Korea Hydro and Nuclear Power Co., Ltd.) ;
  • Yang, Kwang Hee (Radiation Health Research Institute, Korea Hydro and Nuclear Power Co., Ltd.) ;
  • Jeong, Meeseon (Radiation Health Research Institute, Korea Hydro and Nuclear Power Co., Ltd.) ;
  • Kim, Hee Sun (Radiation Health Research Institute, Korea Hydro and Nuclear Power Co., Ltd.) ;
  • Kim, Chong Soon (Korea Institute of Radiological and Medical Sciences) ;
  • Jin, Young-Woo (Radiation Health Research Institute, Korea Hydro and Nuclear Power Co., Ltd.) ;
  • Kim, Joon (School of Life Sciences and Biotechnology, Korea University)
  • Received : 2007.10.11
  • Accepted : 2007.11.01
  • Published : 2007.12.31
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Abstract

The biological effects of low-dose radiation have been investigated and debated for more than a century, but its cellular effects and regulatory mechanisms remain poorly understood. This study shows the human cellular responses to low-dose radiation in CCD-18 Lu cells, which are derived from normal human lung fibroblasts. We examined a colony-forming assay for cell survival by ionizing radiation. Live cell counting and cell cycle analysis were measured for cell proliferation and cell cycle progression following low-dose irradiation. We examined Raf and Akt phosphorylation to determine the proliferation mechanism resulting from low-dose radiation. We also observed that p53 and p21 were related to cell cycle response. We found that 0.05 Gy of ionizing radiation enhanced cell proliferation and did not change the progression of the cell cycle. In addition, 0.05 Gy of ionizing radiation transiently activated Raf and Akt, but did not change phospho-p53, p53 and p21 in CCD-18 Lu cells. However, 2 Gy of ionizing radiation induced cell cycle arrest, phosphorylation of p53, and expression of p53 and p21. The phosphorylation of Raf and Akt proteins induced by 0.05 Gy of ionizing radiation was abolished by pre-treatment with an EGFR inhibitor, AG1478, or a PI3k inhibitor, LY294002. Cell proliferation stimulated by 0.05 Gy of ionizing radiation was blocked by the suppression of Raf and Akt phosphorylation with these inhibitors. These results suggest that 0.05 Gy of ionizing radiation stimulates cell proliferation through the transient activation of Raf and Akt in CCD-18 Lu cells.

Keywords

Acknowledgement

Supported by : KHNP, MOST

References

  1. Andjekovic, M., Jakubowicz, T., Cron, P., Ming, X. F., Han, J. W., et al. (1996) Activation and phosphorylation of a pleckstrin homology domain containing protein kinase (RACPK/ PKB) promoted by serum and protein phosphatase inhibitors. Proc. Natl. Acad. Sci. USA 93, 5699-5704
  2. Bernhard, E. J., Maity, A., Muschel, R. J., and Mckenna, W. G. (1995) Effects of ionizing radiation on cell cycle progression. A review. Radiat. Environ. Biophys. 34, 79-83 https://doi.org/10.1007/BF01275210
  3. Blalock, W. L., Weinstein-Oppenheimer, C., Chang, F., Hoyle, P. E., Wang, X. Y., et al. (1999) Signal transduction, cell cycle regulatory, and anti-apoptotic pathways regulated by IL-3 in hematopoietic cells: possible sites for intervention with antineoplastic drugs. Leukemia 13, 1109-1166 https://doi.org/10.1038/sj/leu/2401493
  4. Brooks, A. L. (2003) Developing a scientific basis for radiation risk estimates: goal of the DOE low dose research program. Health Phys. 85, 85-93 https://doi.org/10.1097/00004032-200307000-00016
  5. Cai, L. (1999) Research of the adaptive response induced by low-dose radiation: where have we been and where should we go? Hum. Exp. Toxicol. 18, 419-425 https://doi.org/10.1191/096032799678840291
  6. Cantley, L. C. (2002) The phosphoinositide 3-kinase pathway. Science 296, 1655
  7. Contessa, J. N., Hampton, J., Lammering, G., Mikkelsen, R. B., Dent, P., et al. (2002) Ionizing radiation activates Erb-B receptorr dependent Akt and p70 S6 kinase signaling in carcinoma cells. Oncogene 21, 4032-4041 https://doi.org/10.1038/sj.onc.1205500
  8. Datta, S. R., Brunet, A., and Greenberg, M. E. (1999) Cellular survival: a play in three Akts, Genes Dev. 13, 2905-2927 https://doi.org/10.1101/gad.13.22.2905
  9. Dent, P., Reardon, D. B., Park, J. S., Bowers, G., Logsdon, C., et al. (1999) Radiation-induced release of transforming growth factor alpha activates the epidermal growth factor receptor and mitogen-activated protein kinase pathway in carcinoma cells, leading to increased proliferation and protection from radiation-induced cell death. Mol. Biol. Cell 10, 2493-2506 https://doi.org/10.1091/mbc.10.8.2493
  10. Dougherty, M. K., Muller, J., Ritt, D. A., Zhou, M., Zhou, X. Z., et al. (2005) Regulation of Raf-1 by direct feedback phosphorylation. Mol. Cell 17, 215-224 https://doi.org/10.1016/j.molcel.2004.11.055
  11. Edwards, E., Geng, L., Tan, J., Onishko, H., Donnelly, E., et al. (2002) Phosphatidylinositol 3-kinase/Akt signaling in the response of vascular endothelium to ionizing radiation. Cancer Res. 62, 4671-4677
  12. El-Deiry, W. S., Tokino, T., Velculescu, V. E., Lwvy, D. B., Parsons, R., et al. (1993) WAF1, a potential mediator of p53 tumor suppression. Cell 75, 817-825 https://doi.org/10.1016/0092-8674(93)90500-P
  13. Fornace, A. J. Jr., Amundson, S. A., Do, K. T., Meltzer, P., Trent, J., et al. (2002) Stress-gene induction by low-dose gamma irradiation. Mil. Med. 167, 13-15 https://doi.org/10.1093/milmed/167.suppl_1.13
  14. Franke, T. F., Kaplan, D. R., Cantley, L. C., and Toker, A. (1997) Direct regulation of the Akt proto-oncogene product by phosphatidylinositol- 3,4-bisphosphate. Science 275, 665
  15. Goldkorn, T., Balaban, N., Shannon, M., and Matsukuma, K. (1997) EGF receptor phosphorylation is affected by ionizing radiation. Biochim. Biophys. Acta 1358, 289-299 https://doi.org/10.1016/S0167-4889(97)00063-3
  16. Grana, T. M., Rusyn, E. V., Zhou, H., Sartor, C. I., and Cox, A. D. (2002) Ras mediates radioresistance through both phosphatidylinositol 3-kinase-dependent and Raf-dependent but mitogenactivated protein kinase/extracellular signal-regulated kinase kinase independent signaling pathways. Cancer Res. 62, 4142-4150
  17. Grant, S., Fisher, P. B., and Dent, P. (2002a) The role of signal transduction pathways in drug and radiation resistance. Cancer Treat. Res. 112, 89-108
  18. Kasid, U., Suy, S., Dent, P., Ray, S., Whiteside, T. L., et al. (1996) Activation of Raf by ionizing radiation. Nature (Lond.) 382, 813-816 https://doi.org/10.1038/382813a0
  19. Kavanagh, B. D., Dent, P., Schmidt-Ullrich, R. K., Chen, P., and Mikkelsen, R. B. (1998) Calcium-dependent stimulation of mitogen-activated protein kinase activity in A431 cells by low doses of ionizing radiation. Radiat. Res. 149, 579-587 https://doi.org/10.2307/3579904
  20. Korystov, Y. N., Eliseeva, N. A., Kublik, L. N., and Narimanov, A. A. (1996) The effect of low-dose irradiation on proliferation of mammalian cells in vitro. Radiat. Res. 146, 329-332 https://doi.org/10.2307/3579464
  21. Lee, S. W., Fang, L., Igarashi, M., Ouchi, T., Lu, K. P., et al. (2000) Sustained activation of Ras/Raf/mitogen-activated protein kinase cascade by the tumor suppressor p53. Proc. Natl. Acad. Sci. USA 97, 8302-8305
  22. Macklis, R. M., Beresford, B. A., Palayoor, S., Sweeney, S., and Humm, J. L. (1993) Cell cycle alterations, apoptosis, and response to low-dose rate radioimmunotherapy in lymphoma cells. Int. J. Radiat. Oncol. Biol. Phys. 27, 643-650 https://doi.org/10.1016/0360-3016(93)90391-8
  23. Maity, A., McKenna, W. G., and Muschel, R. J. (1994) The molecular basis for cell cycle delays following ionizing radiation: a review. Radiother. Oncol. 31, 1-13 https://doi.org/10.1016/0167-8140(94)90408-1
  24. Marshall, M. S. (1995) Ras target proteins in eukaryotic cells. FASEB J. 9, 1311-1318 https://doi.org/10.1096/fasebj.9.13.7557021
  25. Mercer, K., Giblett, S., Oakden, A., Brown, J., Marais R., et al. (2005) A-Raf and Raf-1 work together to influence transient ERK phosphorylation and Gl/S cell cycle progression. Oncogene 24, 5207-5217 https://doi.org/10.1038/sj.onc.1208707
  26. Mizumatsu, S., Monje, M., Morhardt, D., Rola, R., Palmer, T., et al. (2003) Extreme sensitivity of adult neurogenesis to low doses of X-irradiation. Cancer Res. 63, 4021-4027
  27. Morrison, D. K. and Cutler, R. E. (1997) The complexity of Raf- 1 regulation. Curr. Opin. Cell Biol. 9, 174-179 https://doi.org/10.1016/S0955-0674(97)80060-9
  28. Park, W. Y., Hwang, C. I., Im, C. N., Kang, M. J., Woo, J. H., et al. (2002) Identification of radiation-specific responses from gene expression profile. Oncogene 21, 8521-8525 https://doi.org/10.1038/sj.onc.1205977
  29. Rupnow, B. A., Murtha, A. D., Alarcon, R. M., Giaccia, A. J., and Knox, S. J. (1998) Direct evidence that apoptosis enhances tumor responses to fractionated radiotherapy. Cancer Res. 58,1779-1784
  30. Rushworth, L. K., Hindley, A. D., O'Neill, E., and Kolch, W. (2006) Regulation and role of Raf-1/B-Raf heterodimerization. Mol. Cell. Biol. 26, 2262−2272
  31. Schmidt-Ullrich, R. K., Dent, P., Grant, S., Mikkelsen, R. B., and Valerie, K. (2000) Signal transduction and cellular radiation responses. Radiat. Res. 153, 245−257
  32. Sizemore, N., Cox, A. D., Barnard, J. A., Oldham, S. M., Reynolds, E. R., et al. (1999) Pharmacological inhibition of Rastransformed epithelial cell growth is linked to down-regulation of epidermal growth factor-related peptides. Gastroenterology 117, 567−576
  33. Spadinger, I., Marples, B., Matthews, J., and Skov, K. (1994) Can colony size be used to detect low-dose effects? Radiat. Res. 138, S21−S24
  34. Suzuki, K., Kodama, S., and Watanabe, M. (2001) Extremely low-dose ionizing radiation causes activation of mitogenactivated protein kinase pathway and enhances proliferation of normal human diploid cells. Cancer Res. 61, 5396−5401
  35. Vivanco, I. and Sawyers, C. L. (2002) The phosphatidylinositol 3-Kinase AKT pathway in human cancer. Nat. Rev. Cancer 2, 489
  36. Yin, E., Nelson, D. O., Coleman, M. A., Peterson, L. E., and Wyrobek, A. J. (2003) Gene expression changes in mouse brain after exposure to low-dose ionizing radiation. Int. J. Radiat. Biol. 79, 759−775
  37. Kandel, E. S., Skeen, J., Majewski, N., Di Cristofano, A., Pandolfi, P. P., et al. (2002) Activation of Akt/protein kinase B overcomes a G(2)/m cell cycle checkpoint induced by DNA damage. Mol. Cell. Biol. 22, 7831−7841