Radiation makes cells select the form of death dependent on external or internal exposure: apoptosis or pyroptosis.

Internal radiation exposure from neutron-induced radioisotopes environmentally activated following atomic bombing or nuclear accidents should be considered for a complete picture of pathologic effects on survivors. Acute and localized high dose radiation exposure from hot particles taken into the body must induce cell death and severe damage to tissues, whether they are proliferating or not. However, very little the cellular and molecular mechanisms underlying this internal radiation pathology has been investigated. Male Wistar rats were internally exposed to 56MnO2 powder by inhalation. Small intestine samples were investigated by histological staining at acute phase (6 h, 3 days and 14 days) and late phase (2, 6 and 8 months) after the exposure. Histological location and chemical properties of the hot particles embedded in small intestinal tissues were analyzed by synchrotron radiation-X-ray fluorescence-X-ray absorption near-edge structure (SR-XRF-XANES). Hot particles located in the intestinal cavity were identified as accumulations of Mn and iron. Pathological changes showed evidence of crypt shortening, massive cell death at the position of stem cell zone, including apoptosis and pyroptosis from 6 h through 8 months in the internal exposed rats.

The effects of cystine and theanine mixture on the chronic survival rate and tumor incidence of rats after total body X-ray irradiation†.

Cystine and theanine (CT), an amino acid mixture, provides the substrates cysteine and glutamic acid that promote glutathione synthesis. We previously reported that CT pre-treatment significantly improved the acute survival rate and reduced acute radiation injury of the small intestine and bone marrow of rats after 5 Gy of total body X-ray irradiation. To examine the long-term effects of CT administration after irradiation, we investigated the effects of CT pre-treatment and pre- and post-treatment on the chronic survival rate and solid tumor (spleen, skin and subcutis, and thyroid) incidence after irradiation using 7-week-old male Wistar rats. CT pre-treatment of 280 mg/kg was administered orally for 5 days before 5 Gy irradiation, and CT pre- and post-treatment was administered 5 days before and 5 days after irradiation. A 0.5% carboxymethyl cellulose solution was administered as a control. The chronic survival rate of the pre-treated rats was higher than that of the control rats at 441 days after irradiation (40 vs 8.1%, P = 0.011). However, the survival rate did not significantly differ between the pre- and post-treatment and control rats at 467 days after irradiation (33.8 vs 30.2%, P = 0.792). In addition, more solid tumors, especially subcutis sarcomas, were observed in the pre-treatment rats (26.1%, 6/23) than in the control rats (4.5%, 1/22) after irradiation. Therefore, pre-administration of CT improves the chronic survival rate after irradiation; however, the occurrence of solid tumors was not suppressed.

Microdistribution of internal radiation dose in biological tissues exposed to 56Mn dioxide microparticles.

Manganese-56 (56Mn) was one of the dominant neutron-activated radionuclides during the first hours following the atomic-bombing of Hiroshima and Nagasaki. The radiation spectrum of 56Mn and the radiation emission from excited levels of 56Fe following 56Mn beta-decay include gamma-quanta, beta-particles, Auger electrons and X-rays. The dispersion of neutron activated 56Mn in the air can lead to entering of radioactive microparticles into the lungs. The investigation of spatial microdistribution of an internal dose in biological tissue exposed to 56Mn is an important matter with regards to the possible elevated irradiation of the lung alveoli and alveolar ducts. The Monte Carlo code (MCNP-4C) was used for the calculation of absorbed doses in biological tissue around 56Mn dioxide microparticles. The estimated absorbed dose has a very essential gradient in the epithelium cells of lung alveoli and alveolar duct: from 61 mGy/decay on the surface of simple squamous cells of epithelium to 0.15 mGy/decay at distance of 0.3 µm, which is maximal cell thickness. It has been concluded that epithelial cells of these pulmonary microstructures are selectively irradiated by low-energy electrons: short-range component of beta-particles spectrum and Auger electrons. The data obtained are important for the interpretation of biological experiments implementing dispersed neutron-activated 56Mn dioxide powder.

External dose estimates of laboratory rats and mice during exposure to dispersed neutron-activated 56Mn powder.

Estimates of external absorbed dose in experimental animals exposed to sprayed neutron-activated 56Mn powder are necessary for comparison with internal absorbed doses estimated under the same exposure conditions, which is required for a correct interpretation of the observed biological effects. It has been established that the measured dose of external absorbed dose as a result of gamma irradiation range 1-15 mGy, which is order of magnitude less than the maximal dose of internal gamma and beta irradiation of the whole body of the same experimental animals irradiated under the same conditions: according to the available literature data, the maximal values ​​of absorbed dose of internal gamma-beta irradiation of the whole body are in the range of 330 mGy-1200 mGy for mice and 100 mGy-150 mGy for rats. It is concluded that under the conditions of experiments with dispersed neutron-activated powder 56MnO2, internal gamma-beta irradiation of experimental animals is the main factor of radiation exposure compared to external gamma irradiation.

Pathological observation of the effects of exposure to radioactive microparticles on experimental animals.

Internal radiation exposure from neutron-induced radioisotopes that were environmentally activated following an atomic bombing or nuclear accident should be considered for a complete picture of the pathologic effects on survivors. Inhaled hot particles expose neighboring tissues to very high doses of particle beams, which can cause local tissue damage. Experimentally, a few µm of 55MnO2 powder was irradiated with neutrons at a nuclear reactor in order to generate 56MnO2 that emits ß-rays. Rats were irradiated via inhalation. Pathological changes in various rat tissues were examined. In addition, the 56Mn ß energy spectrum around the particles was calculated to determine the local dose rate and the cumulative dose. This review focuses on our latest pathological findings in lungs with internal radiation injury and discusses the pathological changes of early event damage caused by localized, very high-dose internal radiation exposure, including apoptosis, elastin stigma, emphysema, hemorrhage and severe inflammation. The pathological findings of lung tissue due to internal radiation exposure of 0.1 Gy were severe, with no pathological changes observed due to external exposure to γ radiation at a dose of 2.0 Gy. Therefore, it is suggested that new pathological analysis methods for internal exposure due to radioactive microparticles are required.

Age-dependent effects on radiation-induced carcinogenesis in the rat thyroid.

Childhood radiation exposure is a known thyroid cancer risk factor. This study evaluated the effects of age on radiation-induced thyroid carcinogenesis in rats irradiated with 8 Gy X-rays. We analyzed cell proliferation, cell death, DNA damage response, and autophagy-related markers in 4-week-old (4W) and 7-month-old (7M) rats and the incidence of thyroid tumors in 4W, 4-month-old (4M), and 7M rats 18 months after irradiation. Cell death and DNA damage response were increased in 4W rats compared to those in controls at 1 month post-irradiation. More Ki-67-positive cells were observed in 4W rats at 12 months post-irradiation. Thyroid tumors were confirmed in 61.9% (13/21), 63.6% (7/11), and 33.3% (2/6) of irradiated 4W, 4M, and 7M rats, respectively, compared to 0%, 14.3% (1/7), and 16.7% (1/6) in the respective nonirradiated controls. There were 29, 9, and 2 tumors in irradiated 4W, 4M, and 7M rats, respectively. The expression of several autophagy components was downregulated in the area surrounding radiation-induced thyroid carcinomas in 4W and 7M rats. LC3 and p62 expression levels decreased in radiation-induced follicular carcinoma in 4W rats. Radiosensitive cells causing thyroid tumors may be more prevalent in young rats, and abrogation of autophagy may be associated with radiation-induced thyroid carcinogenesis.

Internal doses in experimental mice and rats following exposure to neutron-activated 56MnO2 powder: results of an international, multicenter study.

The experiment was performed in support of a Japanese initiative to investigate the biological effects of irradiation from residual neutron-activated radioactivity that resulted from the A-bombing. Radionuclide 56Mn (T1/2 = 2.58 h) is one of the main neutron-activated emitters during the first hours after neutron activation of soil dust particles. In our previous studies (2016-2017) related to irradiation of male Wistar rats after dispersion of 56MnO2 powder, the internal doses in rats were found to be very inhomogeneous: distribution of doses among different organs ranged from 1.3 Gy in small intestine to less than 0.0015 Gy in some of the other organs. Internal doses in the lungs ranged from 0.03 to 0.1 Gy. The essential pathological changes were found in lung tissue of rats despite a low level of irradiation. In the present study, the dosimetry investigations were extended: internal doses in experimental mice and rats were estimated for various activity levels of dispersed neutron-activated 56MnO2 powder. The following findings were noted: (a) internal radiation doses in mice were several times higher in comparison with rats under similar conditions of exposure to 56MnO2 powder. (b) When 2.74 × 108 Bq of 56MnO2 powder was dispersed over mice, doses of internal irradiation ranged from 0.81 to 4.5 Gy in the gastrointestinal tract (small intestine, stomach, large intestine), from 0.096 to 0.14 Gy in lungs, and doses in skin and eyes ranged from 0.29 to 0.42 Gy and from 0.12 to 0.16 Gy, respectively. Internal radiation doses in other organs of mice were much lower. (c) Internal radiation doses were significantly lower in organs of rats with the same activity of exposure to 56MnO2 powder (2.74 × 108 Bq): 0.09, 0.17, 0.29, and 0.025 Gy in stomach, small intestine, large intestine, and lungs, respectively. (d) Doses of internal irradiation in organs of rats and mice were two to four times higher when they were exposed to 8.0 × 108 Bq of 56MnO2 (in comparison with exposure to 2.74 × 108 Bq of 56MnO2). (e) Internal radiation doses in organs of mice were 7-14 times lower with the lowest 56MnO2 amount (8.0 × 107 Bq) in comparison with the highest amount, 8.0 × 108 Bq, of dispersed 56MnO2 powder. The data obtained will be used for interpretation of biological effects in experimental mice and rats that result from dispersion of various levels of neutron-activated 56MnO2 powder, which is the subject of separate studies.

Impact of Local High Doses of Radiation by Neutron Activated Mn Dioxide Powder in Rat Lungs: Protracted Pathologic Damage Initiated by Internal Exposure.

Internal radiation exposure from neutron-induced radioisotopes environmentally activated following atomic bombing or nuclear accidents should be considered for a complete picture of pathologic effects on survivors. Inhaled hot particles expose neighboring tissues to locally ultra-high doses of ß-rays and can cause pathologic damage. 55MnO2 powder was activated by a nuclear reactor to make 56MnO2 which emits ß-rays. Internal exposures were compared with external γ-rays. Male Wistar rats were administered activated powder by inhalation. Lung samples were observed by histological staining at six hours, three days, 14 days, two months, six months and eight months after the exposure. Synchrotron radiation - X-ray fluorescence - X-ray absorption near-edge structure (SR-XRF-XANES) was utilized for the chemical analysis of the activated 56Mn embedded in lung tissues. 56Mn beta energy spectrum around the particles was calculated to assess the local dose rate and accumulated dose. Hot particles located in the bronchiole and in damaged alveolar tissue were identified as accumulations of Mn and iron. Histological changes showed evidence of emphysema, hemorrhage and severe inflammation from six hours through eight months. Apoptosis was observed in the bronchiole epithelium. Our study shows early event damage from the locally ultra-high internal dose leads to pathogenesis. The trigger of emphysema and hemorrhage was likely early event damage to blood vessels integral to alveolar walls.

Relationship between thyroid tumor radiosensitivity and nuclear localization of DNA-dependent protein kinase catalytic subunit.

Thyroid tumors are the most common types of endocrine malignancies and are commonly treated with radioactive iodine (RAI) to destroy remaining cancer cells following surgical intervention. We previously reported that the expression levels of double-stranded DNA-dependent protein kinase catalytic subunit (DNA-PKcs), which plays a key role in non-homologous end joining, are correlated with the radiosensitivity of cancer cells. Specifically, cells expressing high levels of DNA-PKcs exhibited radiation resistance, whereas cells expressing low levels were sensitive to radiation treatment. In this study, we observed full-length native DNA-PKcs (460 kDa) in radiation-resistant FRO and KTC-2 cells through western blot analysis using an antibody against the C-terminus of DNA-PKcs. In contrast, cleaved DNA-PKcs (175 kDa) were observed in radiation-sensitive TPC-1 and KTC-1 cells. Almost equal amounts of DNA-PKcs were observed in moderately radiation-sensitive WRO cells. We also describe a simple method for the prediction of radiation therapy efficacy in individual cases of thyroid cancers based on staining for DNA-PKcs in human cancer cell lines. Immunofluorescent staining showed that native DNA-PKcs was localized largely in the cytoplasm and only rarely localized in the nuclei of radiation-resistant thyroid cancer cells, whereas in radiation-sensitive cancer cells a 175-kDa cleaved C-terminal fragment of DNA-PKcs was localized mainly inside the nuclei. Therefore, DNA-PKcs moved to the nucleus after γ-ray irradiation. Our results suggest a new method for classifying human thyroid tumors based on their cellular distribution patterns of DNA-PKcs in combination with their radiosensitivity.

Protective effects of a cystine and theanine mixture against acute radiation injury in rats.

This study aims to examine the effects of cystine and theanine (CT), which increases glutathione biosynthesis, on the survival rate and acute radiation injury of the small intestine and bone marrow using a rat model. CT pre-treatment (280 mg/kg for 5 days) significantly improved weight loss and survival rate of rats as compared with the control group after 5 Gy. CT pre-treatment significantly increased the rate of mucosa and crypt length, and decreased the number of apoptotic cells, TUNEL and cleaved caspase-3 positive cells, while increasing the number of mitotic cells and Ki-67 positive cells in jejunal crypts and villi compared to control rats post-irradiation. CT also suppressed bone marrow cell loss and reduced the number of apoptotic cells in bone marrow. These results suggest a protective effect of CT pre-treatment for acute injury after irradiation through apoptosis inhibition and increased proliferative activity in jejunal crypt cells and bone marrow cells.

Wortmannin, a specific inhibitor of phosphatidylinositol-3-kinase, induces accumulation of DNA double-strand breaks.

Wortmannin, a fungal metabolite, is a specific inhibitor of the phosphatidylinositol 3-kinase (PI3K) family, which includes double-stranded DNA dependent protein kinase (DNA-PK) and ataxia telangiectasia mutated kinase (ATM). We investigated the effects of wortmannin on DNA damage in DNA-PK-deficient cells obtained from severe combined immunodeficient mice (SCID cells). Survival of wortmannin-treated cells decreased in a concentration-dependent manner. After treatment with 50 µM wortmannin, survival decreased to 60% of that of untreated cells. We observed that treatment with 20 and 50 µM wortmannin induced DNA damage equivalent to that by 0.37 and 0.69 Gy, respectively, of γ-ray radiation. The accumulation of DNA double-strand breaks (DSBs) in wortmannin-treated SCID cells was assessed using pulsed-field gel electrophoresis. The maximal accumulation was observed 4 h after treatment. Moreover, the presence of DSBs was confirmed by the ability of nuclear extracts from γ-ray-irradiated SCID cells to produce in vitro phosphorylation of histone H2AX. These results suggest that wortmannin induces cellular toxicity by accumulation of spontaneous DSBs through inhibition of ATM.

Reactivation of heat-inactivated Ku proteins by heat shock cognate protein HSC73.

PURPOSE: Mouse double-stranded DNA-dependent protein kinase (DNA-PK) activity is heat sensitive. Recovery of heat-inactivated DNA repair activity is a problem after combination therapy with radiation and heat. We investigated the mechanism of recovery of heat-inactivated DNA-PK activity. METHODS: Hybrid cells containing a fragment of human chromosome 8 in scid cells (RD13B2) were used. DNA-PK activity was measured by an in vitro assay. Immunoprecipitation of the nuclear extract was performed with an anti-Ku80 antibody. Proteins co-precipitated with Ku80 were separated by sodium dodecyl sulfate (SDS) polyacrylamide gel electrophoresis and detected by Western blotting using anti-heat shock protein (HSP)72 and anti-heat shock cognate protein (HSC)73 antibodies. HSC73 was overexpressed with the pcDNA3.1 vector. Short hairpin (sh)RNA was used to downregulate HSC73 and HSP72. RESULTS: The activity of heat-inactivated DNA-PK recovered to about 50% of control during an additional incubation at 37 °C after heat treatment at 44 °C for 15 min in the presence of cycloheximide (which inhibits de novo protein synthesis). Maximal recovery was observed within 3 h of incubation at 37 °C after heat treatment. Constitutively expressed HSC73, which folds newly synthesized proteins, reached maximal levels 3 h after heat treatment using a co-immunoprecipitation assay with the Ku80 protein. Inhibiting HSC73, but not HSP72, expression with shRNA decreased the recovery of DNA-PK activity after heat treatment. CONCLUSIONS: These results suggest that de novo protein synthesis is unnecessary for recovery of some heat-inactivated DNA-PK. Rather, it might be reactivated by the molecular chaperone activity of HSC73, but not HSP72.

Expression of the DNA-dependent protein kinase catalytic subunit is associated with the radiosensitivity of human thyroid cancer cell lines.

The prognosis and treatment of thyroid cancer depends on the type and stage of the disease. Radiosensitivity differs among cancer cells owing to their varying capacity for repair after irradiation. Radioactive iodine can be used to destroy thyroid cancer cells. However, patient prognosis and improvement after irradiation varies. Therefore, predictive measures are important for avoiding unnecessary exposure to radiation. We describe a new method for predicting the effects of radiation in individual cases of thyroid cancer based on the DNA-dependent protein kinase (DNA-PK) activity level in cancer cells. The radiation sensitivity, DNA-PK activity, and cellular levels of DNA-PK complex subunits in five human thyroid cancer cell lines were analyzed in vitro. A positive correlation was observed between the D10 value (radiation dose that led to 10% survival) of cells and DNA-PK activity. This correlation was not observed after treatment with NU7441, a DNA-PK-specific inhibitor. A significant correlation was also observed between DNA-PK activity and expression levels of the DNA-PK catalytic subunit (DNA-PKcs). Cells expressing low DNA-PKcs levels were radiation-sensitive, and cells expressing high DNA-PKcs levels were radiation-resistant. Our results indicate that radiosensitivity depends on the expression level of DNA-PKcs in thyroid cancer cell lines. Thus, the DNA-PKcs expression level is a potential predictive marker of the success of radiation therapy for thyroid tumors.

Autoradiographic analysis of internal plutonium radiation exposure in Nagasaki atomic bomb victims.

BACKGROUND: Radiation doses received by Hiroshima and Nagasaki atomic bomb survivors has been evaluated from data related only to external exposure because there was no reliable evidence for internal exposure in atomic bomb victims. However, we assumed that the contribution of internal exposure cannot be ruled out. METHODS: Autoradiography was carried out with the 70-year old paraffin-embedded specimens taken from Nagasaki atomic bomb victims who died within 5 months after the bombing. After exposure to photo emulsion for 6 months alpha-tracks were revealed in the specimens. We confirmed the alpha-tracks were emitted from deposited plutonium (Pu) in reference to the track length of the 8.787 MeV alpha-particle of thorium series from Polonium-212. Radioactivity concentration of Pu was obtained by counting alpha-tracks. The absorbed dose of each cell nucleus penetrated by an alpha-particle was estimated by calculating the absorbed energy from the particle. RESULTS: Using old paraffin embedded sections processed about 70 years ago, we demonstrated for the first time that conditions in the aftermath of the bombing led to internal exposure to alpha-particles emitted from Pu, the fissile material of the Nagasaki atomic bomb. Dose rate of internal exposure was higher in the victims exposed outdoors than those indoors. Radioactivity concentration was relatively uniform among organs examined in a victim. CONCLUSION: Pu was deposited in the bodies of the Nagasaki A-bomb victims presumably via various routes. Organ dose from Pu of the Nagasaki A-bomb victims studied was during their surviving period, which is lower compared with external exposure. However, the impact to the individual cell nucleus by a single alpha-particle might not be negligible, It would be meaningful; to analyze the relationship of the impact of internal exposure at the cellular level and organ dose. The 70-year old pathological specimens utilized in our study are an invaluable source for understanding internal radiation exposure and are crucial in elucidating experimentally unreproducible phenomena.

Internal exposure to neutron-activated 56Mn dioxide powder in Wistar rats: part 1: dosimetry.

There were two sources of ionizing irradiation after the atomic bombings of Hiroshima and Nagasaki: (1) initial gamma-neutron irradiation at the moment of detonation and (2) residual radioactivity. Residual radioactivity consisted of two components: radioactive fallout containing fission products, including radioactive fissile materials from nuclear device, and neutron-activated radioisotopes from materials on the ground. The dosimetry systems DS86 and DS02 were mainly devoted to the assessment of initial radiation exposure to neutrons and gamma rays, while only brief considerations were given for the estimation of doses caused by residual radiation exposure. Currently, estimation of internal exposure of atomic bomb survivors due to dispersed radioactivity and neutron-activated radioisotopes from materials on the ground is a matter of some interest, in Japan. The main neutron-activated radionuclides in soil dust were 24Na, 28Al, 31Si, 32P, 38Cl, 42K, 45Ca, 46Sc, 56Mn, 59Fe, 60Co, and 134Cs. The radionuclide 56Mn (T 1/2 = 2.58 h) is known as one of the dominant beta- and gamma emitters during the first few hours after neutron irradiation of soil and other materials on ground, dispersed in the form of dust after a nuclear explosion in the atmosphere. To investigate the peculiarities of biological effects of internal exposure to 56Mn in comparison with external gamma irradiation, a dedicated experiment with Wistar rats exposed to neutron-activated 56Mn dioxide powder was performed recently by Shichijo and coworkers. The dosimetry required for this experiment is described here. Assessment of internal radiation doses was performed on the basis of measured 56Mn activity in the organs and tissues of the rats and of absorbed fractions of internal exposure to photons and electrons calculated with the MCNP-4C Monte Carlo using a mathematical rat phantom. The first results of this international multicenter study show that the internal irradiation due to incorporated 56Mn powder is highly inhomogeneous, and that the most irradiated organs of the experimental animals are: large intestine, small intestine, stomach, and lungs. Accumulated absorbed organ doses were 1.65, 1.33, 0.24, 0.10 Gy for large intestine, small intestine, stomach, and lungs, respectively. Other organs were irradiated at lower dose levels. These results will be useful for interpretation of the biological effects of internal exposure of experimental rats to powdered 56Mn as observed by Shichijo and coworkers.

Internal exposure to neutron-activated 56Mn dioxide powder in Wistar rats-Part 2: pathological effects.

To fully understand the radiation effects of the atomic bombing of Hiroshima and Nagasaki among the survivors, radiation from neutron-induced radioisotopes in soil and other materials should be considered in addition to the initial radiation directly received from the bombs. This might be important for evaluating the radiation risks to the people who moved to these cities soon after the detonations and probably inhaled activated radioactive "dust." Manganese-56 is known to be one of the dominant radioisotopes produced in soil by neutrons. Due to its short physical half-life, 56Mn emits residual radiation during the first hours after explosion. Hence, the biological effects of internal exposure of Wistar rats to 56Mn were investigated in the present study. MnO2 powder was activated by a neutron beam to produce radioactive 56Mn. Rats were divided into four groups: those exposed to 56Mn, to non-radioactive Mn, to 60Co γ rays (2 Gy, whole body), and those not exposed to any additional radiation (control). On days 3, 14, and 60 after exposure, the animals were killed and major organs were dissected and subjected to histopathological analysis. As described in more detail by an accompanying publication, the highest internal radiation dose was observed in the digestive system of the rats, followed by the lungs. It was found that the number of mitotic cells increased in the small intestine on day 3 after 56Mn and 60Co exposure, and this change persisted only in 56Mn-exposed animals. Lung tissue was severely damaged only by exposure to 56Mn, despite a rather low radiation dose (less than 0.1 Gy). These data suggest that internal exposure to 56Mn has a significant biological impact on the lungs and small intestine.

Association between p53-binding protein 1 expression and genomic instability in oncocytic follicular adenoma of the thyroid.

Oncocytic follicular adenomas (FAs) of the thyroid are neoplasms of follicular cell origin that are predominantly composed of large polygonal cells with eosinophilic and granular cytoplasm. However, the pathological characteristics of these tumors are largely unexplored. Both the initiation and progression of cancer can be caused by an accumulation of genetic mutations that can induce genomic instability. Thus, the aim of this study was to evaluate the extent of genomic instability in oncocytic FA. As the presence of p53-binding protein 1 (53BP1) in nuclear foci has been found to reflect DNA double-strand breaks that are triggered by various stresses, the immunofluorescence expression pattern of 53BP-1 was assessed in oncocytic and conventional FA. The association with the degree of DNA copy number aberration (CNA) was also evaluated using array-based comparative genomic hybridization. Data from this study demonstrated increased 53BP1 expression (i.e., "unstable" expression) in nuclear foci of oncocytic FA and a higher incidence of CNAs compared with conventional FA. There was also a particular focus on the amplification of chromosome 1p36 in oncocytic FA, which includes the locus for Tumor protein 73, a member of the p53 family implicated as a factor in the development of malignancies. Further evaluations revealed that unstable 53BP1 expression had a significant positive correlation with the levels of expression of Tumor protein 73. These data suggest a higher level of genomic instability in oncocytic FA compared with conventional FA, and a possible relationship between oncocytic FA and abnormal amplification of Tumor protein 73.

Effect of age on the sensitivity of the rat thyroid gland to ionizing radiation.

Exposure to ionizing radiation during childhood is a well-known risk factor for thyroid cancer. Our study evaluated the effect of age on the radiosensitivity of rat thyroid glands. Four-week-old (4W), 7 -week-old (7W), and 8-month-old (8M) male Wistar rats were exposed to 8 Gy of whole-body X-ray irradiation. Thyroids were removed 3-72 h after irradiation, and non-irradiated thyroids served as controls. Ki67-positivity and p53 binding protein 1 (53BP1) focus formation (a DNA damage response) were evaluated via immunohistochemistry. Amounts of proteins involved in DNA damage response (p53, p53 phosphorylated at serine 15, p21), apoptosis (cleaved caspase-3), and autophagy (LC3, p62) were determined via western blotting. mRNA levels of 84 key autophagy-related genes were quantified using polymerase chain reaction arrays. Ki67-positive cells in 4W (with high proliferative activity) and 7W thyroids significantly decreased in number post-irradiation. The number of 53BP1 foci and amount of p53 phosphorylated at serine 15 increased 3 h after irradiation, regardless of age. No increase in apoptosis or in the levels of p53, p21 or cleaved caspase-3 was detected for any ages. Levels of LC3-II and p62 increased in irradiated 4W but not 8M thyroids, whereas expression of several autophagy-related genes was higher in 4W than 8M irradiated thyroids. Irradiation increased the expression of genes encoding pro-apoptotic proteins in both 4W and 8M thyroids. In summary, no apoptosis or p53 accumulation was noted, despite the expression of some pro-apoptotic genes in immature and adult thyroids. Irradiation induced autophagy in immature, but not in adult, rat thyroids.

Radiation-associated small cell neuroendocrine carcinoma of the thyroid: a case report with molecular analyses.

BACKGROUND: Neuroendocrine tumor (NET) of the thyroid other than medullary carcinoma is extremely rare. We describe here a case of calcitonin-negative small cell neuroendocrine carcinoma (SCNEC), which occurred in a thyroid gland that had previously been irradiated at high dose (60 Gy) for pharyngeal cancer, with molecular analyses for follicular cell origin. PATIENT FINDINGS: The tumor cells were small with fine chromatin, inconspicuous nucleoli, and inapparent cytoplasm, and showed neuroendocrine architectures such as palisading, rosettes, and trabeculae. Mitotic figures were numerous exceeding 10 mitoses per 10 high-power fields. The tumor cells invaded into several vessels and metastasized to regional lymph nodes. Immunohistochemically, the tumor cells were strongly positive for neuroendocrine markers and thyroglobulin (Tg), a marker of thyroid follicular cells but negative for calcitonin and carcinoembryonic antigen (CEA). Expression of Tg and thyrotropin receptor (TSHR) were confirmed by quantitative real-time polymerase chain reaction (RT-PCR). Ki-67 labeling index was more than 70% in the tumor cells. Taken together, the tumor was diagnosed as SCNEC of the thyroid. Genetic analyses also revealed microsatellite abnormalities of the phosphatase and tensin homolog (PTEN) gene, suggesting that functional loss of PTEN contributes to carcinogenesis. CONCLUSIONS: This is the first report describing a SCNEC of the thyroid with molecular analyses that provide evidence for a follicular epithelial origin.