MRNA Expression data from AG 1522 human fibroblasts post X-ray radiation
ABSTRACT: Radioadaptive response (RAR) in mammalian cells refers to the phenomenon where a low-dose ionizing irradiation alters the gene expression profiles, and protects the cells from the detrimental effects of a subsequent high dose exposure. We focused on mRNA microarray studies, and aimed to characterize the transcriptome for RAR in AG 1522 human skin fibroblasts and to examine the functional regulatory networks at the genetic level. AG 1522 cells were exposed at a specific time point to a challenging dose of 2 Gy in the RAR group, or a priming dose of 5 cGy in the low-dose group. We aimed to have a comprehensive investigation on the RAR induced in the AG 1522 human fibroblasts first exposed to 5 cGy (priming dose) and then followed by 2 Gy (challenge dose) of X-ray through comparisons to those cells which had only received a single 2 Gy dose.
Project description:Radioadaptive response (RAR) in mammalian cells refers to the phenomenon where a low-dose ionizing irradiation alters the gene expression profiles, and protects the cells from the detrimental effects of a subsequent high dose exposure. We focused on microRNA microarray studies, and aimed to characterize the transcriptome for RAR in AG 1522 human skin fibroblasts and to examine the functional regulatory networks at the genetic level. AG 1522 cells were exposed at a specific time point to a challenging dose of 2 Gy in the RAR group, or a priming dose of 5 cGy in the low-dose group. We aimed to have a comprehensive investigation on the RAR induced in the AG 1522 human fibroblasts first exposed to 5 cGy (priming dose) and then followed by 2 Gy (challenge dose) of X-ray through comparisons to those cells which had only received a single 2 Gy dose.
Project description:Primary term human trophoblasts were derived from placentas after a healthy pregnancy, and exposed to ionizing irradiation (vs sham) in vitro Primary human trophoblasts were irradiated 24 h after initial plating, defined as time zero. Cells were irradiated at 10 Gy using a Clinac 600C (Varian Medical Systems, Palo Alto, CA) with a 6 MV photon beam and a dose rate of 250 cGy/min. The flasks containing the cells were placed on 1.5 cm of bolus (a tissue equivalent material) since the maximum irradiation depth was 1.5 cm, which corresponded to the plated cell layer. Cells were analyzed 4, 8, and 24 h after irradiation or sham.
Project description:The development of a biodosimetric assay based on the combined gene expression analysis of ex vivo-irradiated human peripheral blood (PB) and humans treated with total body irradiation has been developed. Gene expression profile signature of human radiation injury into a rapid and high-throughput chemical ligation-dependent probe amplification assay (CLPA) has been shown to discriminate 6 radiation dose levels in human PB. Candidate gene lists were first developed assaying human ex vivo and human TBI samples on the Affymetrix platform. For the Ex Vivo samples, healthy consented adult donor was randomly assigned to a radiation group and labeled with the target dose of 0 cGy, 150 cGy, 300 cGy, or 600 cGy. In addition to the radiation exposures, the samples were also treated with either no treatment, GCSF, or LPS at time points 6 hours and 24 hours. Radiation expire times from the Cs137 irradiator were calculated to achieve target doses specific for tubes filled with blood and inserted in the roaring test tube holder. The dose rate was 480 cGy/min for the duration of the study. The healthy donors were enrolled to participate in this study following a protocol to collect PB samples that was previously approved by the Duke University Institutional Review Board. For the TBI samples, adult patients, ages 21 to 66, were evaluated at the Duke University Adult Bone Marrow Transplantation Program enrolled in a Duke IRB-approved protocol to collect PB prior to and post TBI conditioning. With approval from the Duke University Institutional Review Board (IRB), between 5-12 mL of peripheral blood was collected from consented patients prior to and 6 hours following TBI with 150-200 cGy as part of their pre-transplantation conditioning. All patients receiving non-myeloablative conditioning were treated with 200 cGy of TBI from a linear accelerator at a dose rate of 20 cGy/min. All patients who underwent TBI-based myeloablative allogeneic or autologous stem-cell transplantation received radiation fractionated at 150 cGy per fraction at 20 cGy/min. All patients had PB collected (50 ml) prior to and 6 hours following exposure to either 200 cGy or 150 cGy radiation treatment. The dose exposure for TBI patients used in this study were 0 cGy, 150 cGy, 450 cGy and 1050 cGy at 0, 6, 30 and 78 hours post irradiation. Total RNA was extracted and assayed on the Affymetrix platform.
Project description:This study investigates three radiation exposure scenarios in BALB/c and C57BL/6 mice: (1) low dose (LD) group -- four weekly doses of 7.5 cGy, (2) high dose (HD) group -- four weekly doses of 1.8 Gy, (3) unexposed group -- four weekly sham exposures. We then used comparative expression profiles of the mouse mammary gland and cardiac blood to build a model of candidate tissue functions associated with LD cancer susceptibility in these strains and murine and human knowledgebases to characterize these tissue functions and their relevance to breast cancer. All samples were assessed on Affymetrix MOE 430 A arrays (HT_MG-430A; GPL8759). Mammary gland tissues: 43 samples (2 strains, 3 treatments, 2 timepoints, 2-4 replicates). For Cardiac blood tissue: 8 samples (2 strains, 4 replicates)
Project description:Skin is usually exposed during human exposures to ionizing radiation, however there are few experiments that evaluate the radiation responsiveness of the cells of the epidermis (keratinocytes) and those of the dermis (fibroblasts) in the same studies. We evaluated the transcriptional responses of quiesent primary keratinocytes and fibroblasts from the same individual and compared them with quiescent keratinocytes and fibroblasts that were immortalized by human telomerase (hTert). The primary transcriptional responses to 10-500 cGy ionizing radiation were p53-mediated responses; however, we did identify distinct responses between the keratinocytes and the fibroblasts. Experiment Overall Design: Four cell types (primary keratinocytes, hTert immortalized keratinocytes, primary fibroblasts, hTert immortalized fibroblasts) grown to quiescence, treated with 0, 10, 100 or 500 cGy gamma irradiation, RNA collected at 4 hrs.
Project description:Most studies have analysed the effects of high dose radiation such as atomic bomb survivors in Japan, people exposed during the Chernobyl nuclear accident, patients undergoing radiation therapy, uranium miners, etc. However, it has been difficult to measure and assess the risk of cancer in people exposed to lower doses of ionising radiation, such as the people living at high altitudes, who are exposed to more natural background radiation from cosmic rays than people at sea level. We measured the genomic response to X-ray ionising radiation (10 cGy and 100 cGy) in a skin tissue model to compare the effects of low and high dose ionising radiation at different time points. The microarray data was then analysed using state-of-the art “upside-down pyramid” computational systems biology methods to identify genes contributing to the difference in the response to the different radiation doses. The model is reconstructed skin tissue, which is composed of keratinocytes that make up the epidermal layer, and fibroblasts that make up the dermal layer of the skin. Tissues were irradiated with 0, 10, and 100 cGy X-ray radiation. Skin plugs were harvested at 0, 3, 8, and 24 hours post irradiation.
Project description:Accumulating data suggest that the biological responses to high and low doses of radiation are qualitatively different, necessitating the direct study of low dose responses. Most such studies have utilized 2-dimensional culture systems, which may not fully represent responses in 3-dimensional tissues. To gain insight into low dose responses in tissue, we have profiled global gene expression in EPI-200, a 3-dimensional tissue model from MatTek that imitates the structure and function of human epidermis, at 4, 16 and 24 hours after exposure to high (2.5 Gy) and low (0.1 Gy) doses of low LET protons. Untreated controls and samples exposed to 10 cGy or to 2.5 Gy were analyzed at three different times (4, 16 or 24 hours after exposure). Three biological repeats were performed for each condition
Project description:While modern radiotherapy technologies can precisely deliver higher doses of radiation to tumors; thus, reducing overall radiation exposure to normal tissues, moderate dose and normal tissue toxicity still remains a significant limitation. The present study profiled the global effects on transcript and miR expression in Human Coronary Artery Endothelial Cells (HCAECs) using single-dose irradiation (SD, 10Gy) or multi-fractionated irradiation (MF, 2Gy x 5) regimens. Longitudinal timepoints were collected after a SD or final dose of MF irradiation for analysis using Agilent Human Gene Expression and miRNA microarray platforms. Compared to SD, the exposure to MF resulted in robust transcript and miR expression changes in terms of the number and magnitude. For data analysis, statistically significant mRNAs (2-fold) and miRs (1.5-fold) were processed by Ingenuity Pathway Analysis (IPA) to uncover miRs associated with target transcripts from several cellular pathways post-irradiation. Interestingly, MF radiation induced a cohort of mRNAs and miRs that coordinate the induction of immune response pathway under tight regulation. Additionally, mRNAs and miRs associated with DNA replication, recombination and repair, apoptosis, cardiovascular events and angiogenesis were revealed. Human Coronary Artery Endothelial Cells (HCAECs) were irradiated in a PANTAK high frequency X-ray generator (Precision X-ray Inc., N. Bedford, CT), operated at 300kV and 10MA. The dose rate was 1.6 Gy per min. Cells were plated into T75cm2 flasks (1-1.5 x 10^6 for single dose radiation and 0.6-0.8 x 10^6 for fractionated radiation). After 24h, cells were exposed to a total of 10 Gy radiation administered either as a single-dose radiation (SD), or as multi-fractionated radiation of 2 Gy x 5 (MF). These non-isoeffective doses were selected to simulate clinical hypofractionated and conventionally fractionated radiotherapy regimens. For the MF protocol, cells were exposed to 2 Gy radiation twice a day, at 6h interval. The cells were approximately 90% confluent at the time of harvesting. For both protocols, radiation-induced changes were analyzed at 6h and 24h after a SD and 6h and 24h after the final dose of fractionated irradiation. Separate controls were maintained for SD and MF radiation protocols.
Project description:The role of RB1 in response to radiation was examined in human osteoblasts. We demonstrate that RB1 induced SASP genes, a response which was attenuated in RB1 knockdown osteoblasts. Subconfluent hOB shEV and shRB1 cells were irradiated at 4 Gy using a 137Cesium source at a dose rate of 1.7 min/Gy and cell pelllets collected 0, 2, 4, 8, 16 and 24 hrs after irradiation.