Project description:Transcriptomic profiling of normal mouse thyroid tissue following 211At irradiation Introduction The cellular response to stimuli can be studied on different molecular levels, including gene expression regulation. RNA microarray is a high-throughput technique that enables comparison of genome-wide transcriptional status between samples. 211At-labeled radiopharmaceuticals are potentially useful for tumor therapy. Metabolically released 211At accumulates in the thyroid gland, which is one of the main critical organs for such therapy. The aim of this study was to determine the effect of absorbed dose, dose-rate, and time after 211At exposure on transcriptional expression in mouse thyroid gland. Method BALB/c mice were i.v. injected with 1.7, 7.5 or 100 kBq 211At. Animals injected with 1.7 kBq were killed after 1, 6, or 168 h with mean absorbed doses to thyroid of 0.023, 0.32, and 1.4 Gy, respectively. Animals injected with 7.5 and 100 kBq were killed after 6 and 1 h, respectively, resulting in mean absorbed doses to thyroid of 1.4 Gy. Animals were killed through cardiac puncture under anesthesia, and thyroids were removed and snap-frozen. Total RNA was extracted from pooled thyroid tissue samples and mRNA levels were determined with the Illumina RNA microarray platform. Nexus Expression 3.0 was used to identify differentially expressed transcripts (adjusted p-value <0.01, fold change >1.5) and enriched GO terms (p-value <0.05) between irradiated groups and controls. Results In total, 1232 differentially expressed transcripts were detected after 211At administration, demonstrating a profound effect on gene regulation. The number of regulated transcripts increased with higher initial dose-rate/absorbed dose when the exposure time was 1 or 6 h. However, the number of regulated transcripts decreased with mean absorbed dose/time after 1.7 kBq 211At administration. This suggests that initial dose-rate (thus injected activity) is an important parameter in radiation-induced responses. Furthermore, the regulation profiles of groups administered 1.7 kBq were similar. Few previously proposed radiation responsive genes were identified in the present study, but, e.g. Gadd45g and Cdkn1a were among the regulated genes identified in the present study. A response involving immunological processes was identified especially at 1, 6, and 168 h after 1.7 kBq administration (0.023, 0.32, 1.8 Gy). Conclusion The present study allowed a comprehensive assessment of how variations in exposure parameters affect transcriptional regulation. The regulation profiles were dependent on both absorbed dose and time after exposure, but, initial dose-rate was the most influential parameter. Functional annotation of regulated genes revealed exposure-specific effects on biological processes. Additionally, we have identified several recurrently regulated genes after 211At exposure that may play a role in how mouse thyroid tissue responds to 211At exposure. Introduction The cellular response to stimuli can be studied on different molecular levels, including gene expression regulation. RNA microarray is a high-throughput technique that enables comparison of genome-wide transcriptional status between samples. 211At-labeled radiopharmaceuticals are potentially useful for tumor therapy. Metabolically released 211At accumulates in the thyroid gland, which is one of the main critical organs for such therapy. The aim of this study was to determine the effect of absorbed dose, dose-rate, and time after 211At exposure on transcriptional expression in mouse thyroid gland. Method BALB/c mice were i.v. injected with 1.7, 7.5 or 100 kBq 211At. Animals injected with 1.7 kBq were killed after 1, 6, or 168 h with mean absorbed doses to thyroid of 0.023, 0.32, and 1.4 Gy, respectively. Animals injected with 7.5 and 100 kBq were killed after 6 and 1 h, respectively, resulting in mean absorbed doses to thyroid of 1.4 Gy. Animals were killed through cardiac puncture under anesthesia, and thyroids were removed and snap-frozen. Total RNA was extracted from pooled thyroid tissue samples and mRNA levels were determined with the Illumina RNA microarray platform. Nexus Expression 3.0 was used to identify differentially expressed transcripts (adjusted p-value <0.01, fold change >1.5) and enriched GO terms (p-value <0.05) between irradiated groups and controls. Results In total, 1232 differentially expressed transcripts were detected after 211At administration, demonstrating a profound effect on gene regulation. The number of regulated transcripts increased with higher initial dose-rate/absorbed dose when the exposure time was 1 or 6 h. However, the number of regulated transcripts decreased with mean absorbed dose/time after 1.7 kBq 211At administration. This suggests that initial dose-rate (thus injected activity) is an important parameter in radiation-induced responses. Furthermore, the regulation profiles of groups administered 1.7 kBq were similar. Few previously proposed radiation responsive genes were identified in the present study, but, e.g. Gadd45g and Cdkn1a were among the regulated genes identified in the present study. A response involving immunological processes was identified especially at 1, 6, and 168 h after 1.7 kBq administration (0.023, 0.32, 1.8 Gy). Conclusion The present study allowed a comprehensive assessment of how variations in exposure parameters affect transcriptional regulation. The regulation profiles were dependent on both absorbed dose and time after exposure, but, initial dose-rate was the most influential parameter. Functional annotation of regulated genes revealed exposure-specific effects on biological processes. Additionally, we have identified several recurrently regulated genes after 211At exposure that may play a role in how mouse thyroid tissue responds to 211At exposure. Total RNA was isolated from fresh-frozen tissue samples (Normal balb/c mouse thyroids)

Project description:Transcriptomic profiling of normal mouse thyroid tissue following 211At irradiation Introduction The cellular response to stimuli can be studied on different molecular levels, including gene expression regulation. RNA microarray is a high-throughput technique that enables comparison of genome-wide transcriptional status between samples. 211At-labeled radiopharmaceuticals are potentially useful for tumor therapy. Metabolically released 211At accumulates in the thyroid gland, which is one of the main critical organs for such therapy. The aim of this study was to determine the effect of absorbed dose, dose-rate, and time after 211At exposure on transcriptional expression in mouse thyroid gland. Method BALB/c mice were i.v. injected with 1.7, 7.5 or 100 kBq 211At. Animals injected with 1.7 kBq were killed after 1, 6, or 168 h with mean absorbed doses to thyroid of 0.023, 0.32, and 1.4 Gy, respectively. Animals injected with 7.5 and 100 kBq were killed after 6 and 1 h, respectively, resulting in mean absorbed doses to thyroid of 1.4 Gy. Animals were killed through cardiac puncture under anesthesia, and thyroids were removed and snap-frozen. Total RNA was extracted from pooled thyroid tissue samples and mRNA levels were determined with the Illumina RNA microarray platform. Nexus Expression 3.0 was used to identify differentially expressed transcripts (adjusted p-value <0.01, fold change >1.5) and enriched GO terms (p-value <0.05) between irradiated groups and controls. Results In total, 1232 differentially expressed transcripts were detected after 211At administration, demonstrating a profound effect on gene regulation. The number of regulated transcripts increased with higher initial dose-rate/absorbed dose when the exposure time was 1 or 6 h. However, the number of regulated transcripts decreased with mean absorbed dose/time after 1.7 kBq 211At administration. This suggests that initial dose-rate (thus injected activity) is an important parameter in radiation-induced responses. Furthermore, the regulation profiles of groups administered 1.7 kBq were similar. Few previously proposed radiation responsive genes were identified in the present study, but, e.g. Gadd45g and Cdkn1a were among the regulated genes identified in the present study. A response involving immunological processes was identified especially at 1, 6, and 168 h after 1.7 kBq administration (0.023, 0.32, 1.8 Gy). Conclusion The present study allowed a comprehensive assessment of how variations in exposure parameters affect transcriptional regulation. The regulation profiles were dependent on both absorbed dose and time after exposure, but, initial dose-rate was the most influential parameter. Functional annotation of regulated genes revealed exposure-specific effects on biological processes. Additionally, we have identified several recurrently regulated genes after 211At exposure that may play a role in how mouse thyroid tissue responds to 211At exposure. Introduction The cellular response to stimuli can be studied on different molecular levels, including gene expression regulation. RNA microarray is a high-throughput technique that enables comparison of genome-wide transcriptional status between samples. 211At-labeled radiopharmaceuticals are potentially useful for tumor therapy. Metabolically released 211At accumulates in the thyroid gland, which is one of the main critical organs for such therapy. The aim of this study was to determine the effect of absorbed dose, dose-rate, and time after 211At exposure on transcriptional expression in mouse thyroid gland. Method BALB/c mice were i.v. injected with 1.7, 7.5 or 100 kBq 211At. Animals injected with 1.7 kBq were killed after 1, 6, or 168 h with mean absorbed doses to thyroid of 0.023, 0.32, and 1.4 Gy, respectively. Animals injected with 7.5 and 100 kBq were killed after 6 and 1 h, respectively, resulting in mean absorbed doses to thyroid of 1.4 Gy. Animals were killed through cardiac puncture under anesthesia, and thyroids were removed and snap-frozen. Total RNA was extracted from pooled thyroid tissue samples and mRNA levels were determined with the Illumina RNA microarray platform. Nexus Expression 3.0 was used to identify differentially expressed transcripts (adjusted p-value <0.01, fold change >1.5) and enriched GO terms (p-value <0.05) between irradiated groups and controls. Results In total, 1232 differentially expressed transcripts were detected after 211At administration, demonstrating a profound effect on gene regulation. The number of regulated transcripts increased with higher initial dose-rate/absorbed dose when the exposure time was 1 or 6 h. However, the number of regulated transcripts decreased with mean absorbed dose/time after 1.7 kBq 211At administration. This suggests that initial dose-rate (thus injected activity) is an important parameter in radiation-induced responses. Furthermore, the regulation profiles of groups administered 1.7 kBq were similar. Few previously proposed radiation responsive genes were identified in the present study, but, e.g. Gadd45g and Cdkn1a were among the regulated genes identified in the present study. A response involving immunological processes was identified especially at 1, 6, and 168 h after 1.7 kBq administration (0.023, 0.32, 1.8 Gy). Conclusion The present study allowed a comprehensive assessment of how variations in exposure parameters affect transcriptional regulation. The regulation profiles were dependent on both absorbed dose and time after exposure, but, initial dose-rate was the most influential parameter. Functional annotation of regulated genes revealed exposure-specific effects on biological processes. Additionally, we have identified several recurrently regulated genes after 211At exposure that may play a role in how mouse thyroid tissue responds to 211At exposure.

Project description:Introduction Iodine-131 (131I) is frequently used in nuclear medicine. Unbound or released 131I accumulate in the thyroid gland and may be detrimental to normal thyroid function. The aim of the present study was to identify biomarkers for 131I exposure in rat thyroid tissue and to assess the effect on thyroid function. Methods Thirty six male Sprague Dawley rats were i.v. injected with 150 Âµl saline solution containing 9.0, 88, 170, 260, 340, 760, 1300, or 4700 kBq (group A-H) 131I, or mock-treated with 150 Âµl saline solution only, and killed at 24 h after injection. Total RNA was extracted from individual thyroid tissue samples thyroids and mRNA levels were determined with the Agilent microarray platform. Results Estimated absorbed doses in treatment groups A-H was 0.0058, 0.057, 0.11, 0.17, 0.22, 0.5 Gy, 0.8 Gy, and 3 Gy. Totally, 429 transcripts were identified with a fold change fold change â¥ 1.5 and adjusted p-value â¤ 0.01. A trend with downregulation of thyroid hormone biosynthesis associated genes (e.g. thyroglobulin, thyroid peroxidase, the sodium-iodine symporter) was identified, but only statistically significant after 0.0058 and 0.22 Gy. Three transcripts coding for isoform 1 of the DBP protein showed a pattern with monotonous decrease in downregulation with absorbed dose between 0.0058-0.22 Gy. Changes in Dbp expression were not statistically significant between 0.5-3 Gy. However, a trend with downregulation at 0.5 and 0.8 Gy and upregulation and 3 Gy was identified. Previously, 131I (0.85-17 Gy) and 211At (0.023-32 Gy) exposure resulted in upregulation of Dbp in mice thyroid tissue 24 h after administrations. Additionally, a monotonous decrease in Dbp downregulation has been identified of in mouse kidney tissue at 8 and 12 months after 177Lu-octreotate administrations. Conclusion Conclusively, the Dbp gene is a promising candidate biomarker gene for exposure to 131I and possibly other internal radiation emitters. Further studies should be performed to establish how Dbp expression vary with dose-rate, absorbed dose, time after administration, different radiation qualities, and the function of Dbp. Total RNA was isolated from fresh-frozen individual thyroid tissue samples (Sprague Dawley rats). Each sample was run once. Four rats received the same treatment. Control samples (from non-irradiated rats) are included.

Project description:Introduction Iodine-131 (131I) is frequently used in nuclear medicine. Unbound or released 131I accumulate in the thyroid gland and may be detrimental to normal thyroid function. The aim of the present study was to identify biomarkers for 131I exposure in rat thyroid tissue and to assess the effect on thyroid function. Methods Thirty six male Sprague Dawley rats were i.v. injected with 150 µl saline solution containing 9.0, 88, 170, 260, 340, 760, 1300, or 4700 kBq (group A-H) 131I, or mock-treated with 150 µl saline solution only, and killed at 24 h after injection. Total RNA was extracted from individual thyroid tissue samples thyroids and mRNA levels were determined with the Agilent microarray platform. Results Estimated absorbed doses in treatment groups A-H was 0.0058, 0.057, 0.11, 0.17, 0.22, 0.5 Gy, 0.8 Gy, and 3 Gy. Totally, 429 transcripts were identified with a fold change fold change ≥ 1.5 and adjusted p-value ≤ 0.01. A trend with downregulation of thyroid hormone biosynthesis associated genes (e.g. thyroglobulin, thyroid peroxidase, the sodium-iodine symporter) was identified, but only statistically significant after 0.0058 and 0.22 Gy. Three transcripts coding for isoform 1 of the DBP protein showed a pattern with monotonous decrease in downregulation with absorbed dose between 0.0058-0.22 Gy. Changes in Dbp expression were not statistically significant between 0.5-3 Gy. However, a trend with downregulation at 0.5 and 0.8 Gy and upregulation and 3 Gy was identified. Previously, 131I (0.85-17 Gy) and 211At (0.023-32 Gy) exposure resulted in upregulation of Dbp in mice thyroid tissue 24 h after administrations. Additionally, a monotonous decrease in Dbp downregulation has been identified of in mouse kidney tissue at 8 and 12 months after 177Lu-octreotate administrations. Conclusion Conclusively, the Dbp gene is a promising candidate biomarker gene for exposure to 131I and possibly other internal radiation emitters. Further studies should be performed to establish how Dbp expression vary with dose-rate, absorbed dose, time after administration, different radiation qualities, and the function of Dbp.

Project description:Rationale: Auger electrons bear a low energy (0-10 keV) which is deposited in a very small volume (a few nm3) around the site of emission. From a radio-toxicological point of view, the effects of Auger electrons on normal tissues are largely unknown, understudied and generally assumed negligible. In this context, the discovery that the Auger electron emitter 99mTc can induce stunning on primary thyrocytes in vitro, at low doses, is intriguing. Extrapolated in vivo, this observation suggests that, a radioisotope as commonly used in nuclear medicine as 99mTc may significantly influence thyroid biology. The aim of the present study was to evaluate the absorbed dose of 99mTc pertechnetate (99mTcO4−) required to induce thyroid stunning in vivo and to analyse the biological events associated/concomitant with this effect. Methods: SPECT/CT imaging of thyroid uptake of 99mTcO4− was used to demonstrate stunning effect in mice and quantitative analysis of the images was used for dosimetric calculations. The biological effects associated with this phenomenon were evaluated by Western blotting, quantitative RT-PCR, histology, immuno-histochmistry and global gene expression pattern analysis. Results: Auger electrons-mediated thyroid stunning can be observed in vivo, in mouse thyroid. The threshold of the absorbed dose to the thyroid required to obtain a significant stunning effect is in the range of 20 Gy. This effect is associated with a reduction in the levels of functional Na/I symporter (NIS) protein without any significant cell death. It is reversible within days. At the cellular level, a decrease in NIS mRNA, the activation of the p53 pathway and the generation of double-strand DNA breaks can be observed. Conclusion: Auger electrons emitted by 99mTc can induce thyroid stunning in vivo, in mice but the thyroid needs to be exposed to a dose of at least 20 Gy, a dose unlikely to be encountered in the clinical practice. Nevertheless, this report represents, to our knowledge, the first example of a significant action of Auger electrons on a normal tissue, in vivo and provides a unique experimental set-up to understand the fine molecular mechanisms involved in the biological effects of Auger electrons. One color design, corresponding to 2 conditions performed on 4 mice: TcO4- (Tc)-treated versus Control (Ct) mice for a total of 8 samples.

Project description:Purpose: The International Commission on Radiological Protection (ICRP) recently recommended reducing the occupational equivalent dose limit for the lens of the eye. Based primarily on a review of epidemiological data, the absorbed dose threshold is now considered to be 0.5 Gy for induction of acute opacities, reduced from the previous threshold of 2 Gy. However, direct mechanistic evidence to support an understanding of the underlying molecular mechanisms of damage is still lacking. To this end, we explored the effects of a broad dose-range of ionizing radiation exposure on gene expression changes in a human lens epithelial (HLE) cell-line in order to better understand the shape of the dose-response relationship and identify transcriptional thresholds of effects. Methods: HLE cells were exposed to doses of 0, 0.01, 0.05, 0.25, 0.5, 2, and 5 Gy of X-ray radiation at two dose rates (1.62 cGy/min and 38.2 cGy/min). Cell culture lysates were collected 20 h post-exposure and analysed using whole-genome RNA-sequencing. Pathways and dose-thresholds of biological effects were identified using benchmark dose (BMD) modeling. Results: Transcriptional responses were minimal at doses less than 2 Gy. At higher doses there were a significant number of differentially expressed genes (DEGs) (p < 0.05, fold change > |1.5|) at both dose rates, with 1308 DEGs for the low dose rate (LDR) and 840 DEGs for the high dose rate (HDR) exposure. Dose-response modeling showed that a number of genes exhibited non-linear bi-phasic responses, which was verified by digital droplet PCR. BMD analysis showed the majority of the pathways responded at BMD median values in the dose range of 1.5-2.5 Gy, with the lowest BMD median value being 0.6 Gy for the HDR exposure. The minimum pathway BMD median value for LDR exposure, however, was 2.5 Gy. Although the LDR and HDR exposures shared pathways involved in extracellular matrix reorganization and collagen production with BMD median value of 2.9 Gy, HDR exposures were more effective in activating pathways associated with DNA damage response, apoptosis, and cell cycling relative to LDR exposure. Conclusion: Overall, the results suggest that radiation induces complex non-linear transcriptional dose-response relationships that are dose-rate dependent. Pathways shared between the two dose rates may be important contributors to radiation-induced cataractogenesis. BMD analysis suggests that the majority of pathways are activated above 0.6 Gy, which supports current ICRP identified dose thresholds for deterministic effects to the lens of the eye of 0.5 Gy.

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Transcriptional response of mouse thyroids following in vivo 211At exposure reveals distinct gene expression profiles

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