Transcriptional response of BALB/c mouse thyroids following in vivo astatine-211 exposure reveals distinct gene expression profiles

Abstract

Background: Astatine-211 (211At) is an alpha particle emitting halogen with almost optimal linear energy transfer for creating DNA double-strand breaks and is thus proposed for radionuclide therapy when bound to tumor-seeking agents. Unbound 211At accumulates in the thyroid gland, and the concept of basal radiation-induced biological effects in the thyroid tissue is, to a high degree, unknown and is most valuable.

Methods: Female BALB/c nude mice were intravenously injected with 0.064 to 42 kBq of 211At, resulting in absorbed doses of 0.05 to 32 Gy in the thyroid gland. Thyroids were removed 24 h after injection; total RNA was extracted from pooled thyroids and processed in triplicate using Illumina MouseRef-8 Whole-Genome Expression Beadchips.

Results: Thyroids exposed to 211At revealed distinctive gene expression profiles compared to non-irradiated controls. A larger number of genes were affected at low absorbed doses (0.05 and 0.5 Gy) compared to intermediate (1.4 Gy) and higher absorbed doses (11 and 32 Gy). The proportion of dose-specific genes increased with decreased absorbed dose. Additionally, 1.4 Gy often exerted opposite regulation on gene expression compared to the other absorbed doses. Using Gene Ontology data, an immunological effect was detected at 0.05 and 11 Gy. Effects on cellular response to external stress and cell cycle regulation and proliferation were detected at 1.4 and 11 Gy.

Conclusions: Conclusively, the cellular response to ionizing radiation is complex and differs with absorbed dose. The response acquired at high absorbed doses cannot be extrapolated down to low absorbed doses or vice versa. We also demonstrated that the thyroid - already at absorbed doses similar to those obtained in radionuclide therapy - responds with expression of a high number of genes. Due to the increased heterogeneous irradiation at low absorbed doses, we suggest that this response partly originates from non-irradiated cells in the tissue, i.e., bystander cells.

Figure 1

Proportion of shared genes at the different absorbed dose levels. The distribution of regulated transcripts is shown in a radar plot for the different absorbed dose levels: (A) 0.05 Gy, (B) 0.5 Gy, (C) 1.4 Gy, (D) 11 Gy, and (E) 32 Gy. For example, in (A), 91%, 18%, 36%, and 29% of the transcripts regulated at 0.05 Gy were also regulated at 0.5, 1.4, 11, and 32 Gy. For the exact distribution of regulated transcripts, see Additional file 2.

Figure 2

Differentially expressed genes present at all absorbed doses. Genes categorized by known gene expression in mouse thyroid according to UniGene and The Human Protein Atlas. Second column: genes are listed together with their log₂ ratio value where the red color scale and the green color scale represent the magnitude of gene down-regulation and up-regulation, respectively. Genes annotated with ‘Unknown’ are genes with unknown gene and/or protein expression in normal thyroid tissue, and genes annotated with ‘o’ indicate that the gene is not expressed in normal thyroid tissue. For gene expression in mouse thyroid tissue, ‘+’ indicates gene expression, whereas for protein expression, ‘+’, ‘++’, or ‘+++’ indicates weak, moderate, or strong protein expression levels, respectively. The fifth column shows the log₂ values of gene expression in non-irradiated mouse thyroids (the present study). Genes with a value equal to or greater than log₂ 6.8 are considered as expressed in normal mouse thyroid.

Figure 3

Gene expression patterns for a selection of genes vs. absorbed dose. A selection of genes chosen in order to illustrate the different gene expression patterns among the 130 differentially expressed genes at all absorbed doses studied. Values for gene expression are given as comparison log₂ ratio values related to non-irradiated tissue.