X-射线致人淋巴细胞基因组差异表达及剂量—效应关系研究
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摘要
目的:在全基因组水平筛选辐射反应基因,验证X-射线照射诱导人外周血淋巴细胞基因表达谱的差异及剂量-效应关系,为阐明辐射生物学效应信号通路和利用基因表达改变作为辐射生物剂量计提供实验依据。
方法:以剂量为0.5、2.0、5.0Gy的X-射线照射正常人外周血淋巴细胞,用Agilent人基因组表达芯片检测照射后24小时全基因组mRNA表达改变,通过芯片扫描和软件分析,得到3个剂量组的差异表达谱,通过统计学检验分析、倍数变化和交集分析筛选出特异表达和共表达的差异基因。用SOM分析筛选辐射剂量依赖性表达上调基因,用统计分析软件检验和建立剂量-效应关系模型,并用实时荧光定量PCR进行验证。用层次聚类、Gene Ontology和GenMapp等数据库分析辐射相关信号通路,以流式细胞术和激光共聚焦显微技术检测细胞凋亡的变化。
结果:
1.基因表达谱芯片技术检测到各剂量组差异表达基因共计522个,在共表达的38个辐射反应基因中,18个已知基因的表达呈剂量依赖性上调,其中DDB2、BBC3、PCNA、IFNG、TNFSF4、TNFSF8、ZNF788、TMEM30A、DACH2、TNFRSF10B等基因表达的变化与辐照剂量呈直线回归关系,GADD45A、PHLDA3、ISG20L、APOBEC3H、E2F7、PPM1D等基因的表达与剂量间的变化关系符合指数模型。
2.实时荧光定量PCR验证2.0和5.0Gy照射组GADD45A、XPC、BAX、P21基因表达上调,Ku70、XRCC3基因表达下调,与基因芯片结果相同。GADD45基因表达在1.0～5.0Gy照射后存在剂量-效应关系,曲线拟合符合指数模型,与基因芯片结果一致。在2.0GyX射线照射后该基因表达的时间变化规律为1h开始上升,4h达到高峰,并持续到照射后72h,照后96 h基因表达水平恢复正常。p21基因表达在1.0～3.0Gy照射后与照射剂量存在线性剂量-效应关系。
3.芯片的数据分析表明,参与辐射反应的基因涉及DNA应激、DNA损伤修复、细胞周期控制、细胞凋亡、免疫反应等生物学过程。其中11个基因为P53直接调控的靶基因。信号通路分析揭示辐射主要诱导p53信号通路、Fas和热休克蛋白介导的凋亡通路、细胞周期调控和炎症反应通路。
结论
1.基因芯片技术可作为筛选辐射反应基因的有力工具。
2.不同剂量X-射线照射淋巴细胞后可致基因表达谱的特征性改变,部分基因的差异表达具有剂量-效应关系和时间-效应关系。
3. X-射线的辐射损伤效应,涉及一些特定的辐射反应基因和细胞信号通路。特征性的辐射反应基因有可能成为潜在的辐射生物剂量计。
Objective To screen radiation responsive genes at the level of global genome to search for dose-response relationship induced by X-ray irradiation to explore the possibility of specific gene expression patterns and signal pathways as radiation biodosimetry.
Methods Agilent human oligo microarray was used to screen differential expressive genes in homo sapien PBLs exposed to 0.5, 2.0, 5.0Gy X-ray irradiation. Radiation responsive genes were selected by statistics significance, fold change and intersection analysis. Dose-dependent up-regulated genes were mapped out with SOM method and confirmed by Quantitative RT-PCR. Radiation responsive signal pathways were analyzed through hierarchical clustering, Gene Ontology and GenMAPP databases. Cell apoptosis was detected with flow cytometry and laser confocal microscope.
Results
1. Total of 522 radiation responsive genes were screened out with oligo microarray. Among 38 coexpression radiation responsive genes, 18 were identified as dose-dependent responsive genes. A linear dose response relationship was established between IR and gene expression changes for DDB2, BBC3, PCNA, IFNG, TNFSF4, TNFSF8, ZNF788, TMEM30A, DACH2, TNFRSF10B and an exponential model for GADD45A, PHLDA3, ISG20L, APOBEC3H, E2F7, PPM1D over the dose range from 0.5 to 5.0Gy .
2. The up-regulated genes of XPC, BAX, P21 and down-regulated genes of Ku70, XRCC3 were verified with Quantitative RT-PCR corresponded to the microarray results. The expression of GADD45 gene was dose-dependent ranging from 1.0Gy to 5.0Gy, rapidly increased at 1h, peaked at 4h, remained high for 72h and retuned to basal level by 96h post irradiation. A linear dose response relationship of gene expression for P21 was also verified over the range from 1.0 to 3.0 Gy.
3. The radiation responsive genes were functional in DNA stress, DNA damage repair, cell cycle control, apoptosis and immune response. Among them 11 genes were downstream target genes of P53. Signal pathway analysis revealed that P53 signal pathway, Fas ligand, apoptosis modulation via HSP70 pathway, cell cycle control and inflammatory response pathways were involved in radiation response to X-ray exposure.
Conclusions
1. Microarray technique is a powerful tool in screening radiation responsive genes.
2. Characteristic of differential expression induced by X-ray is explored in lymphocytes with specific dose-response and time-response patterns.
3. X-ray induced genome changes include radiation responsive genes and signaling pathways which might be applied as potential radiation biodosimetry.
方法:以剂量为0.5、2.0、5.0Gy的X-射线照射正常人外周血淋巴细胞,用Agilent人基因组表达芯片检测照射后24小时全基因组mRNA表达改变,通过芯片扫描和软件分析,得到3个剂量组的差异表达谱,通过统计学检验分析、倍数变化和交集分析筛选出特异表达和共表达的差异基因。用SOM分析筛选辐射剂量依赖性表达上调基因,用统计分析软件检验和建立剂量-效应关系模型,并用实时荧光定量PCR进行验证。用层次聚类、Gene Ontology和GenMapp等数据库分析辐射相关信号通路,以流式细胞术和激光共聚焦显微技术检测细胞凋亡的变化。
结果:
1.基因表达谱芯片技术检测到各剂量组差异表达基因共计522个,在共表达的38个辐射反应基因中,18个已知基因的表达呈剂量依赖性上调,其中DDB2、BBC3、PCNA、IFNG、TNFSF4、TNFSF8、ZNF788、TMEM30A、DACH2、TNFRSF10B等基因表达的变化与辐照剂量呈直线回归关系,GADD45A、PHLDA3、ISG20L、APOBEC3H、E2F7、PPM1D等基因的表达与剂量间的变化关系符合指数模型。
2.实时荧光定量PCR验证2.0和5.0Gy照射组GADD45A、XPC、BAX、P21基因表达上调,Ku70、XRCC3基因表达下调,与基因芯片结果相同。GADD45基因表达在1.0～5.0Gy照射后存在剂量-效应关系,曲线拟合符合指数模型,与基因芯片结果一致。在2.0GyX射线照射后该基因表达的时间变化规律为1h开始上升,4h达到高峰,并持续到照射后72h,照后96 h基因表达水平恢复正常。p21基因表达在1.0～3.0Gy照射后与照射剂量存在线性剂量-效应关系。
3.芯片的数据分析表明,参与辐射反应的基因涉及DNA应激、DNA损伤修复、细胞周期控制、细胞凋亡、免疫反应等生物学过程。其中11个基因为P53直接调控的靶基因。信号通路分析揭示辐射主要诱导p53信号通路、Fas和热休克蛋白介导的凋亡通路、细胞周期调控和炎症反应通路。
结论
1.基因芯片技术可作为筛选辐射反应基因的有力工具。
2.不同剂量X-射线照射淋巴细胞后可致基因表达谱的特征性改变,部分基因的差异表达具有剂量-效应关系和时间-效应关系。
3. X-射线的辐射损伤效应,涉及一些特定的辐射反应基因和细胞信号通路。特征性的辐射反应基因有可能成为潜在的辐射生物剂量计。
Objective To screen radiation responsive genes at the level of global genome to search for dose-response relationship induced by X-ray irradiation to explore the possibility of specific gene expression patterns and signal pathways as radiation biodosimetry.
Methods Agilent human oligo microarray was used to screen differential expressive genes in homo sapien PBLs exposed to 0.5, 2.0, 5.0Gy X-ray irradiation. Radiation responsive genes were selected by statistics significance, fold change and intersection analysis. Dose-dependent up-regulated genes were mapped out with SOM method and confirmed by Quantitative RT-PCR. Radiation responsive signal pathways were analyzed through hierarchical clustering, Gene Ontology and GenMAPP databases. Cell apoptosis was detected with flow cytometry and laser confocal microscope.
Results
1. Total of 522 radiation responsive genes were screened out with oligo microarray. Among 38 coexpression radiation responsive genes, 18 were identified as dose-dependent responsive genes. A linear dose response relationship was established between IR and gene expression changes for DDB2, BBC3, PCNA, IFNG, TNFSF4, TNFSF8, ZNF788, TMEM30A, DACH2, TNFRSF10B and an exponential model for GADD45A, PHLDA3, ISG20L, APOBEC3H, E2F7, PPM1D over the dose range from 0.5 to 5.0Gy .
2. The up-regulated genes of XPC, BAX, P21 and down-regulated genes of Ku70, XRCC3 were verified with Quantitative RT-PCR corresponded to the microarray results. The expression of GADD45 gene was dose-dependent ranging from 1.0Gy to 5.0Gy, rapidly increased at 1h, peaked at 4h, remained high for 72h and retuned to basal level by 96h post irradiation. A linear dose response relationship of gene expression for P21 was also verified over the range from 1.0 to 3.0 Gy.
3. The radiation responsive genes were functional in DNA stress, DNA damage repair, cell cycle control, apoptosis and immune response. Among them 11 genes were downstream target genes of P53. Signal pathway analysis revealed that P53 signal pathway, Fas ligand, apoptosis modulation via HSP70 pathway, cell cycle control and inflammatory response pathways were involved in radiation response to X-ray exposure.
Conclusions
1. Microarray technique is a powerful tool in screening radiation responsive genes.
2. Characteristic of differential expression induced by X-ray is explored in lymphocytes with specific dose-response and time-response patterns.
3. X-ray induced genome changes include radiation responsive genes and signaling pathways which might be applied as potential radiation biodosimetry.
引文
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(1) George A. Alexandera et al.BiodosEPR-2006 Meeting: Acute dosimetry consensus committee recommendations on biodosimetry applications in events involving uses of radiation by terrorists and radiation accident .Radiation Measurements. 2007 42:972 - 996
(2) Amundson SA, Do Shahab S, et a1.Identification of potential mRNA biomarkers in per peripheral blood lymphocytes for human exposure to ionizing radiation J. Radiat Res. 2000,154(3):342-346
(3) Grace MB, McLeland CB, Blakely .Real-time quantitative RT-PCR assay of GADD45 gene expression changes as a biomarker for radiation biodesimetry. J Radiat Biol. 2002;78(11):1011-1021
(4) Grace Marcy B., Christopher B. McLeland, Steven J. Gagliardi, Jeffrey M. Smith, William E. Jackson, and William F. Blakely. Development and Assessment of a Quantitative Reverse Transcription-PCR Assay for Simultaneous Measurement of Four Amplicons. Clin. Chem. Sep 2003; 49: 1467– 1475
(5) Blakely W.F., Miller A.C., Grace M.B., McLeland C.B., Luo L., Muderhwa J.M., Miner V.L., Prasanna P.G... Radiation biodosimetry: applications for spaceflight. Adv. Space Res. 2003a;31 (6):1487–1493
(6) Amundson S.A., Grace M.B., McLeland C.B., Epperly M.W., Yeager A., Zhan Q., Greenberger J.S., Fornace Jr. A.J.,. Human in vivo radiation-induced biomarkers: gene expression changes in radiotherapy patients. Cancer Res. 2004;64 (18): 6368-6371.
(7) Blakely W.F., Prasanna P.G.S., Kolanko C.J., Pyle M.D., Mosbrook D.M., Loats A.S., Rippeon T.L., Loats H. Application of the premature chromosome condensation assay in simulated partial-body radiation exposures: evaluation of the use of an automated metaphasefinder.Stem Cells. 1995;13 (Suppl. 1):223–230.
(8) Kerri E. Rieger and Gilbert Chu. Portrait of transcriptional responses to ultraviolet and ionizing radiation in human cells.Nucleic Acids Research. 2004;32(16): 4786–4803
(9) Kuang-Yu Jen and Vivian G. Cheung Transcriptional Response ofLymphoblastoid Cells to Ionizing Radiation Genome Res. 2003;13:2092-2100
(10) Amundson SA, Bittner M, Chen Y, Trent J, Meltzer P,Fornace AJ Jr. Fluorescent cDNA microarray hybridization reveals complexity and heterogeneity of cellular genotoxic stress responses. Oncogene 1999;18:3666–72
(11) Amundson SA, Bittner M, Meltzer P, Trent J, Fornace AJ Jr. Induction of gene expression as a monitor of exposure to ionizing radiation. Radiat Res. 2001;156:657–61
(12) Sally A. Amundson, R. Anthony Lee, Christine A. Koch-Paiz, Michael L. Bittner, Paul Meltzer, Jeffrey M. Trent and Albert J. Fornace, Jr Differential Responses of Stress Genes to Low Dose-Rate Irradiation. Mol. Cancer Res. 2003; 1: 445.
(13) Zhan Q , Bae I , Kastan MB et al . The p532dependent gamma-ray response of GADD45. Cancer Res 1994 ;54 :2755-2760.
(14) Kearsey JM, Coates PJ ,Prescott AR et al . GADD45 is a nuclear cell cycle regulated protein which interacts with p21 (CIP1) . Oncogene 1995;11:1675-1683.
(15) Kastan MB, Onyekwere O , Sidransky D..et al . Participation of p53 protein in the cellular response to DNA damage. Cancer Res .1991;51:6304-6311.
(16) Torres M, Al-Buhairi M and Alsbeih G.. Induction of p53 and p21 proteins by gamma radiation in skin fibroblasts derived from breast cancer patients. Int J Radiat Oncol Biol Phys. 2004; 58(2): 479-484.
(17) FU H Q, JU G Z, SU X et al. J Radiat Res Process. 2000;18(2):86-90
(18) Baranov AE, Guskova AK, Nadejina NM, et al. Chernobyl experience: Biological indicators of exposure to ionizing radiation. Stem Cells, 1995;13(Suppl l) :69-77.
(19) Hall D, Hall EJ. Radiobiology for the Radiologist . Philadelphia:Lippincott JB Co. 1993. 316-320
(20) Amundson S. A, Do K. T., and Fornace A. J. Induction of stress genes by low doses of gamma rays. Radiat. Res., 1999; 152: 225– 231.
(21) Han-Fei Ding et al. Essential Role for Caspase-8 in Transcription-independent Apoptosis Triggered by p53, the JOURNAL OFBIOLOGICAL CHEMISTRY 2000,276(49): 38905–38911
(22) Valerie K and L.f. Povirk. Regulation and mechanism of mammalian double strand break repair.Oncogene. 2003.;22(37):5792-812
(23) Atsushi Hirao, Young-Yun Kong, Shuhei Matsuoka, Andrew Wakeham, Jürgen Ruland, Hiroki Yoshida, Dou Liu, Stephen J. Elledge and Tak W. Ma DNA Damage-Induced Activation of p53 by the Checkpoint Kinase Chk2 Science. 2000; 287: 1824-34
(24) Zhou B.B.and S.J.Elledgethe DNA damage response : putting checkpoints in perpective , Nature.2000;408(6811):433-9
(25) Vogelstein B., D.Lane et al .Surfing the p53 network. Nature.200;408(6810):323-31
(26) Levine A.J. et al P53, the cellular gate keeper for growth and division. Cell. 1997;88(3):323-31
(27) Maya R.and M. Balass et al. ATM-dependent phosphorylation of Mdm2 on serine role in p53 activation by DNA damage . Genes Dev. 2001;15(9):1067-77
(28) Takimoto R. and W.s El Deiry Wild–type P53 transactivates the KILLER/DR5 gene through on intronic sequence specific DNA-binding site . Oncogene. 2000,19(14):1735-43
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(5) Kuang Yu Jen and Vivian G. Cheung. Identification of Novel p53 Target Genes in Ionizing Radiation Response. Cancer Res. 2005; 65: (17):7666-7673
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(7) Amundson SA, Shahab S, Bittner M, Meltzer P, Trent J, Fornace AJ Jr. Identification of potential mRNA markers in peripheral blood lymphocytes for human exposure to ionizing radiation. Radiation Res. 2000;154:342-346.
(8) Amundson SA, Bittner M, Chen Y, Trent J, Meltzer P, Fornace AJ Jr. Fluorescent cDNA microarray hybridization reveals complexity and heterogeneity of cellular genotoxic stress responses. Oncogene. 1999;18:3666-3672.
(9) Amundson SA, Bittner M, Meltzer P, Trent J, Fornace AJ Jr. Induction of gene expression as a monitor of exposure to ionizing radiation. Radiat Res. 2001;156:657–61.
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(11) Amundson. Sally A, Marcy B. Grace, Christopher B. McLeland, Michael W. Epperly, Andrew Yeager, Qimin Zhan, Joel S. Greenberger and Albert J. Fornace. Human In vivo Radiation-Induced Biomarkers: Gene Expression Changes in Radiotherapy Patients Cancer Res. 2004; 64: 6368- 6371.
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(1) George A. Alexander et al.BiodosEPR-2006 Meeting: Acute dosimetry consensus committee recommendations on biodosimetry applications in events involving uses of radiation by terrorists and radiation accident. Radiation Measurements. 2007; 42:972 - 996
(2) Amundson SA, Do Shahab S. et a1.Identification of potential mRNA biomarkers in peripheral blood lymphocytes for human exposure to ionizing radiation J.Radiat. Res. 2000;154(3):342-346
(3) Grace MB, McLeland CB, Blakely. Real-time quantitative RT-PCR assay of GADD45 gene expression changes as a biomarker for radiation biodesimetry. J Radiat. Biol. 2002;78(11):1011-1021
(4) Grace Marcy B., Christopher B. McLeland, Steven J. Gagliardi, Jeffrey M. Smith, William E. Jackson and William F. Blakely. Development and Assessment of a Quantitative Reverse Transcription-PCR Assay for Simultaneous Measurement of Four Amplicons. Clin. Chem. 2003; 49: 1467-1475
(5) Blakely W.F., Miller A.C., Grace M.B., McLeland C.B., Luo L.,Muderhwa J.M., Miner V.L.,Prasanna P.G. Radiation biodosimetry: applications for spaceflight. Space Res. 2003;31 (6):1487-1493
(6) Amundson S.A., Grace M.B., McLeland C.B., Epperly M.W., Yeager A., Zhan Q., Greenberger J.S., Fornace Jr. A.J. Human in vivo radiation-induced biomarkers: gene expression changes in radiotherapy patients. Cancer Res. 2004;64 (18): 6368-6371.
(7) Blakely W.F., Prasanna P.G.S., Kolanko C.J., Pyle M.D., Mosbrook D.M., Loats A.S., Rippeon T.L., Loats H.. Application of the premature chromosomecondensation assay in simulated partial-body radiation exposures: evaluation of the use of an automated metaphase finder. Stem Cells. 1995;13 (Suppl.1): 223–230.
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(9) Kerri E. Rieger and Gilbert Chu Portrait of transcriptional responses to ultraviolet and ionizing radiation in human cells. Nucleic Acids Research. 2004;32(16): 4786-4803
(10) Kuang-Yu Jen and Vivian G. Cheung Transcriptional Response of Lymphoblastoid Cells to Ionizing Radiation. Genome Res. 2003;13:2092-2100
(11) Amundson Sally A., Marcy B. Grace, Christopher B. McLeland, Michael W. Epperly, Andrew Yeager, Qimin Zhan, Joel S. Greenberger and Albert J. Fornace Human In vivo Radiation-Induced Biomarkers: Gene Expression Changes in Radiotherapy Patients Cancer Res. 2004; 64: 6368 - 6371.
(12) Amundson SA, Bittner M, Chen Y, Trent J, Meltzer P, Fornace AJ Jr. Fluorescent cDNA microarray hybridization reveals complexity and heterogeneity of cellular genotoxic stress responses. Oncogene. 1999;18:3666–72
(13) Amundson SA, Bittner M, Meltzer P, Trent J, Fornace AJ Jr. Induction of gene expression as a monitor of exposure to ionizing radiation. Radiat Res. 2001;156:657–61
(14) Sally A. Amundson, R. Anthony Lee, Christine A. Koch-Paiz, Michael L. Bittner, Paul Meltzer, Jeffrey M. Trent, and Albert J. Fornace, Jr. Differential Responses of Stress Genes to Low Dose-Rate Irradiation. Mol. Cancer Res. 2003; 1: 445-452.
(15) Zhan Q, Bae I, Kastan MB. et al.. The p532 dependent gamma-ray response of GADD45. Cancer Res. 1994;54 :2755-2760.
(16) Kearsey JM, Coates PJ., Prescott AR, et al. GADD45 is a nuclear cell cycle regulated protein which interacts with p21 (CIP1). Oncogene 1995;11 :1675-1683.
(17) Kastan MB, Onyekwere O, Sidransky D,et al . Participation of p53 protein in the cellular response to DNA damage. Cancer Res. 1991;51 : 6304-6311.
(18) Torres M, Al-Buhairi M and Alsbeih G.. Induction of p53 and p21 proteinsby gamma radiation in skin fibroblasts derived from breast cancer patients. Int. J Radiat. Oncol Biol Phys. 2004; 58(2): 479-484.
(19) Fu H Q, Ju G Z, Su X et al. J Radiat Res Process 2000;18(2)86-90
(20) Baranov AE, Guskova AK, Nadejina NM, et al. Chernobyl experience: Biological indicators of exposure to ionizing radiation. Stem Cells, 1995;13(Suppl l) :69-77.
(21) Hall D, Hall EJ. Radiobiology for the Radiologist . Philadelphia:Lippincott JB Co. 1993. 316-320.
(22) Amundson S. A, Do K. T., and Fornace A. J. Induction of stress genes by low doses of gamma rays. Radiat. Res., 1999; 152: 225– 231.
(1) Akira Fujimori, Ryuichi Okayasu, Hiroshi Ishihara, Satoshi Yoshida, Kiyomi Eguchi-Kasai, Kumie Nojima, Satoru Ebisawa and Sentaro Takahashi. Extremely Low Dose Ionizing Radiation Up-regulates CXC Chemokines in Normal Human Fibroblasts. Cancer Res. Nov 2005; 65: 10159 - 10163.
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(3) Adam J. Krieg,Ester M. Hammond,and Amato J. Giaccia. Functional Analysis of p53 Binding under Differential Stresses Mol. Cell. Biol. 2006; 26: 7030 - 7045.
(4) Kuang-Yu Jen and Vivian G. Cheung. Identification of Novel p53 Target Genes in Ionizing Radiation Response. Cancer Res. 2005; 65: (17).7666-7673
(5) Amundson SA, Bittner M, Chen Y, Trent J, Meltzer P, Fornace AJ Jr. Fluorescent cDNA microarray hybridization reveals complexity and heterogeneity of cellular genotoxic stress responses. Oncogene 1999;18:3666–72
(6) Tusher,V.G..et al. Significance analysis of microarrays applied to the ionizing radiation response. Pro. Natl. Acad. Aci. 2001;98(9):5116-21
(7) Jen, K. Y. and V. G. Cheung . Transcriptional response of lymphoblastiod cells to ionizing radiation. The FASEB Journal. 2003; 13(9):2092-100
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(11) http://www.genmapp.org/
(12)李瑶等基因芯片数据分析与处理.化学工业出版社, 2006第一版157-177
(13) Han-Fei Ding et al. Essential Role for Caspase-8 in Transcription-independent Apoptosis Triggered by p53, the Journal of biological Chemistry. 2000;276(49):38905-38911
(14) Valerie K. and L.f. Povirk. Regulation and mechanism of mammalian double strand break repair.Oncogene. 2003.;22(37):5792-812
(15) Atsushi Hirao, Young-Yun Kong, Shuhei Matsuoka, Andrew Wakeham, Jürgen Ruland, Hiroki Yoshida, Dou Liu, Stephen J. Elledge, and Tak W. Ma. DNA Damage-Induced Activation of p53 by the Checkpoint Kinase Chk2. Science. 2000; 287: 1824-9
(16) Zhou B.B. and S.J.Elledgethe DNA damage response : putting checkpoints in perpective, Nature. 2000;408(6811):433-9
(17) Vogelstein B., D.Lane et al .Surfing the p53 network. Nature. 200;408(6810):323-31
(18) Levine A.J. et al P53, the cellular gatekeeper for growth and division. Cell. 1997;88(3):323-31
(19) Maya R., M. Balass, et al. ATM-dependent phosphorylation of Mdm2 on serine role in p53 activation by DNA damage . Genes Dev. 2001;15(9):1067-77
(20) Takimoto R. and W.s El Deiry Wild–type P53 transactivates the KILLER/DR5 gene through on intronic sequence specific DNA-binding site . Oncogene. 2000; 19(14):1735-43
(21) Amundson SA, Do Shahab S,eta1.Identification of potential mRNA biomarkers in peripheral blood lymphocytes for human exposure to ionizing radiation J Radiat Res. 2000;154(3):342-346
(22) Amundson SA, .Marcy B. Grace,Christopher B. McLeland,Michael W. Epperly,Andrew Yeager,Qimin Zhan,Joel S. Greenberger, and Albert J. Fornace, Jr. Human In vivo Radiation-Induced Biomarkers: Gene Expression Changes in Radiotherapy Patients Cancer Res.2004; 64: 6368 - 6371.
(23) Sally A. Amundson, R. Anthony Lee, Christine A. Koch-Paiz, Michael L. Bittner, Paul Meltzer, Jeffrey M. Trent and Albert J. Fornace, Jr Differential Responses of Stress Genes to Low Dose-Rate Irradiation. Mol. Cancer Res. 2003; 1: 445.
(1) George A. Alexandera et al.BiodosEPR-2006 Meeting: Acute dosimetry consensus committee recommendations on biodosimetry applications in events involving uses of radiation by terrorists and radiation accident .Radiation Measurements. 2007 42:972 - 996
(2) Amundson SA, Do Shahab S, et a1.Identification of potential mRNA biomarkers in per peripheral blood lymphocytes for human exposure to ionizing radiation J. Radiat Res. 2000,154(3):342-346
(3) Grace MB, McLeland CB, Blakely .Real-time quantitative RT-PCR assay of GADD45 gene expression changes as a biomarker for radiation biodesimetry. J Radiat Biol. 2002;78(11):1011-1021
(4) Grace Marcy B., Christopher B. McLeland, Steven J. Gagliardi, Jeffrey M. Smith, William E. Jackson, and William F. Blakely. Development and Assessment of a Quantitative Reverse Transcription-PCR Assay for Simultaneous Measurement of Four Amplicons. Clin. Chem. Sep 2003; 49: 1467– 1475
(5) Blakely W.F., Miller A.C., Grace M.B., McLeland C.B., Luo L., Muderhwa J.M., Miner V.L., Prasanna P.G... Radiation biodosimetry: applications for spaceflight. Adv. Space Res. 2003a;31 (6):1487–1493
(6) Amundson S.A., Grace M.B., McLeland C.B., Epperly M.W., Yeager A., Zhan Q., Greenberger J.S., Fornace Jr. A.J.,. Human in vivo radiation-induced biomarkers: gene expression changes in radiotherapy patients. Cancer Res. 2004;64 (18): 6368-6371.
(7) Blakely W.F., Prasanna P.G.S., Kolanko C.J., Pyle M.D., Mosbrook D.M., Loats A.S., Rippeon T.L., Loats H. Application of the premature chromosome condensation assay in simulated partial-body radiation exposures: evaluation of the use of an automated metaphasefinder.Stem Cells. 1995;13 (Suppl. 1):223–230.
(8) Kerri E. Rieger and Gilbert Chu. Portrait of transcriptional responses to ultraviolet and ionizing radiation in human cells.Nucleic Acids Research. 2004;32(16): 4786–4803
(9) Kuang-Yu Jen and Vivian G. Cheung Transcriptional Response ofLymphoblastoid Cells to Ionizing Radiation Genome Res. 2003;13:2092-2100
(10) Amundson SA, Bittner M, Chen Y, Trent J, Meltzer P,Fornace AJ Jr. Fluorescent cDNA microarray hybridization reveals complexity and heterogeneity of cellular genotoxic stress responses. Oncogene 1999;18:3666–72
(11) Amundson SA, Bittner M, Meltzer P, Trent J, Fornace AJ Jr. Induction of gene expression as a monitor of exposure to ionizing radiation. Radiat Res. 2001;156:657–61
(12) Sally A. Amundson, R. Anthony Lee, Christine A. Koch-Paiz, Michael L. Bittner, Paul Meltzer, Jeffrey M. Trent and Albert J. Fornace, Jr Differential Responses of Stress Genes to Low Dose-Rate Irradiation. Mol. Cancer Res. 2003; 1: 445.
(13) Zhan Q , Bae I , Kastan MB et al . The p532dependent gamma-ray response of GADD45. Cancer Res 1994 ;54 :2755-2760.
(14) Kearsey JM, Coates PJ ,Prescott AR et al . GADD45 is a nuclear cell cycle regulated protein which interacts with p21 (CIP1) . Oncogene 1995;11:1675-1683.
(15) Kastan MB, Onyekwere O , Sidransky D..et al . Participation of p53 protein in the cellular response to DNA damage. Cancer Res .1991;51:6304-6311.
(16) Torres M, Al-Buhairi M and Alsbeih G.. Induction of p53 and p21 proteins by gamma radiation in skin fibroblasts derived from breast cancer patients. Int J Radiat Oncol Biol Phys. 2004; 58(2): 479-484.
(17) FU H Q, JU G Z, SU X et al. J Radiat Res Process. 2000;18(2):86-90
(18) Baranov AE, Guskova AK, Nadejina NM, et al. Chernobyl experience: Biological indicators of exposure to ionizing radiation. Stem Cells, 1995;13(Suppl l) :69-77.
(19) Hall D, Hall EJ. Radiobiology for the Radiologist . Philadelphia:Lippincott JB Co. 1993. 316-320
(20) Amundson S. A, Do K. T., and Fornace A. J. Induction of stress genes by low doses of gamma rays. Radiat. Res., 1999; 152: 225– 231.
(21) Han-Fei Ding et al. Essential Role for Caspase-8 in Transcription-independent Apoptosis Triggered by p53, the JOURNAL OFBIOLOGICAL CHEMISTRY 2000,276(49): 38905–38911
(22) Valerie K and L.f. Povirk. Regulation and mechanism of mammalian double strand break repair.Oncogene. 2003.;22(37):5792-812
(23) Atsushi Hirao, Young-Yun Kong, Shuhei Matsuoka, Andrew Wakeham, Jürgen Ruland, Hiroki Yoshida, Dou Liu, Stephen J. Elledge and Tak W. Ma DNA Damage-Induced Activation of p53 by the Checkpoint Kinase Chk2 Science. 2000; 287: 1824-34
(24) Zhou B.B.and S.J.Elledgethe DNA damage response : putting checkpoints in perpective , Nature.2000;408(6811):433-9
(25) Vogelstein B., D.Lane et al .Surfing the p53 network. Nature.200;408(6810):323-31
(26) Levine A.J. et al P53, the cellular gate keeper for growth and division. Cell. 1997;88(3):323-31
(27) Maya R.and M. Balass et al. ATM-dependent phosphorylation of Mdm2 on serine role in p53 activation by DNA damage . Genes Dev. 2001;15(9):1067-77
(28) Takimoto R. and W.s El Deiry Wild–type P53 transactivates the KILLER/DR5 gene through on intronic sequence specific DNA-binding site . Oncogene. 2000,19(14):1735-43