Biomarkers of Atherosclerotic Vascular Disease in Workers Chronically Exposed to Ionizing Radiation.

ABSTRACT: It is well established that cohorts of individuals exposed to ionizing radiation demonstrate increased risks of cardio- and cerebrovascular diseases. However, mechanisms of these radiation-induced diseases developing in individuals exposed to ionizing radiation remain unclear. To identify biomarkers of the atherosclerotic vessel damage in workers chronically exposed to ionizing radiation, this study considered 49 workers of the Russian nuclear production facility-the Mayak Production Association (mean age of 68.73 ± 6.92 years)-and 38 unexposed individuals (mean age of 68.84 ± 6.20 y) who had never been exposed to ionizing radiation (control). All workers were chronically exposed to combined radiation (external gamma rays and internal alpha particles). The mean cumulative liver absorbed dose from external gamma-ray exposure was 0.18 ± 0.12 Gy; the mean cumulative liver absorbed dose from internal alpha-particles was 0.14 ± 0.21 Gy. Levels of biomarkers in blood serum of the study participants were measured using the ELISA method. Elevated levels of apolipoprotein B, superoxide dismutase, monocyte chemoattractant protein 1, vascular cell adhesion protein 1, and a decreased level of endothelin-1 were observed in blood serum of Mayak PA workers chronically exposed to combined radiation compared to control individuals. A significant positive correlation was demonstrated between the vascular cell adhesion protein 1 level and cumulative liver absorbed doses from external gamma radiation and internal alpha radiation. Findings of the study suggest that molecular changes in blood of individuals occupationally exposed to ionizing radiation (combined internal exposure to alpha particles and external exposure to gamma rays) may indicate dyslipidemia, oxidative stress, inflammation, and endothelial dysfunction involved in atherosclerosis development.

Immunological Markers of Chronic Occupational Radiation Exposure.

This study aimed to identify immunological biomarkers for prolonged occupational radiation exposure and thus studied a random sample of the Mayak Production Association worker cohort (91 individuals). The control group included 43 local individuals never employed at the Mayak Production Association. To identify biomarkers, two groups of workers were formed: the first one included workers chronically exposed to external gamma rays at cumulative doses of 0.5-3.0 Gy (14 individuals); the second one included workers exposed to combined radiation-external gamma rays at doses ranging from 0.7 to 5.1 Gy and internal alpha radiation from incorporated plutonium with 0.3-16.4 kBq body burden (77 individuals). The age range of the study individuals was 66-91 y. Peripheral blood serum protein concentrations of cytokines, immunoglobulins, and matrix metalloproteinase-9 were analyzed using enzyme-linked immunoassay following the manufacturer's protocol. Flow cytometry was used to analyze levels of various lymphocyte subpopulations. The findings of the current study demonstrate that some immunological characteristics may be considered as biomarkers of prolonged chronic radiation exposure for any radiation type (in the delayed period after the exposure) based on fold differences from controls: M immunoglobulin fold differences were 1.75 ± 0.27 (p = 0.0001) for external gamma-ray exposure and 1.50 ± 0.27 (p = 0.0003) for combined radiation exposure; matrix metalloproteinase-9 fold differences were 1.5 ± 0.22 (p = 0.008) for external gamma-ray exposure and 1.69 ± 0.24 (p = 0.00007) for combined radiation exposure; A immunoglobulin fold differences were 1.61 ± 0.27 (p = 0.002) for external gamma-ray exposure and 1.56 ± 0.27 (p = 0.00002) for combined radiation exposure; relative concentration of natural killer cell fold differences were 1.53 ± 0.23 (p = 0.01) for external gamma-ray exposure and 1.35 ± 0.22 (p = 0.001) for combined radiation exposure; and relative concentration of T-lymphocytes fold differences were 0.89 ± 0.04 (p = 0.01) for external gamma-ray exposure and 0.95 ± 0.05 (p = 0.003) for combined radiation exposure. Based on fold differences from controls, interferon-gamma (3.50 ± 0.65, p = 0.031), transforming growth factor-beta (2.91 ± 0.389, p = 0.026), and relative blood serum levels of T-helper cells (0.90 ± 0.065, p = 0.02) may be used as immunological markers of chronic external gamma-ray exposure. Moreover, there was a significant inverse linear association of relative concentration of T-helper cells with dose from external gamma rays accumulated over an extended period.

Expression of blood serum proteins and lymphocyte differentiation clusters after chronic occupational exposure to ionizing radiation.

This study aimed to assess effects of chronic occupational exposure on immune status in Mayak workers chronically exposed to ionizing radiation (IR). The study cohort consists of 77 workers occupationally exposed to external gamma-rays at total dose from 0.5 to 3.0 Gy (14 individuals) and workers with combined exposure (external gamma-rays at total dose range 0.7-5.1 Gy and internal alpha-radiation from incorporated plutonium with a body burden of 0.3-16.4 kBq). The control group consists of 43 age- and sex-matched individuals who never were exposed to IR, never involved in any cleanup operations following radiation accidents and never resided at contaminated areas. Enzyme-linked immunoassay and flow cytometry were used to determine the relative concentration of lymphocytes and proteins. The concentrations of T-lymphocytes, interleukin-8 and immunoglobulins G were decreased in external gamma-exposed workers relative to control. Relative concentrations of NKT-lymphocytes, concentrations of transforming growth factor-ß, interferon gamma, immunoglobulins A, immunoglobulins M and matrix proteinase-9 were higher in this group as compared with control. Relative concentrations of T-lymphocytes and concentration of interleukin-8 were decreased, while both the relative and absolute concentration of natural killers, concentration of immunoglobulins A and M and matrix proteinase-9 were increased in workers with combined exposure as compared to control. An inverse linear relation was revealed between absolute concentration of T-lymphocytes, relative and absolute concentration of T-helpers cells, concentration of interferon gamma and total absorbed dose from external gamma-rays in exposed workers. For workers with incorporated plutonium, there was an inverse linear relation of absolute concentration of T-helpers as well as direct linear relation of relative concentration of NKT-lymphocytes to total absorbed red bone marrow dose from internal alpha-radiation. In all, chronic occupational IR exposure of workers induced a depletion of immune cells in peripheral blood of the individuals involved.

Radiation-induced lung adenocarcinoma is associated with increased frequency of genes inactivated by promoter hypermethylation.

Epigenetic inactivation of genes by promoter hypermethylation, a major mechanism in the initiation and progression of tobacco-induced cancer, has also been associated with lung cancer induced through environmental and occupational exposures. Our previous study of gene methylation in workers from the MAYAK nuclear enterprise identified a significantly higher prevalence for methylation of the p16 gene (CDKN2A) in adenocarcinomas from workers compared to tumors from non-worker controls. The purpose of this investigation was to determine whether genes in addition to p16 are "targeted" for silencing and whether overall gene methylation was more common in radiation-induced adenocarcinoma. A significant increase in the prevalence of methylation of GATA5 was seen in tumors from workers compared to tumors from controls. The prevalence for methylation of PAX5 beta and H-cadherin did not differ in tumors from workers and controls. Evaluating the frequency for methylation of a five-gene panel revealed that 93% of adenocarcinomas from workers compared to 66% of tumors from controls were methylated for at least one gene. Moreover, a twofold increase was seen in the number of tumors methylated for three or more genes for tumors from workers compared to controls. Increased frequency for inactivation of genes by promoter hypermethylation and targeting of tumor suppressor genes such as GATA5 may be factors that contribute to the increased risk for lung cancer associated with radiation exposure.

Plutonium targets the p16 gene for inactivation by promoter hypermethylation in human lung adenocarcinoma.

Lung cancer from radon or (239)plutonium exposure has been linked to alpha-particles that damage DNA through large deletions and point mutations. We investigated the involvement of an epigenetic mechanism, gene inactivation by promoter hypermethylation in adenocarcinomas from plutonium-exposed workers at MAYAK, the first Russian nuclear enterprise established to manufacture weapons plutonium. Adenocarcinomas were collected retrospectively from 71 workers and 69 non-worker controls. Lung adenocarcinomas were examined from workers and non-worker controls for methylation of the CDKN2A (p16), O(6)-methylguanine-DNA methyltransferase (MGMT), death associated protein kinase (DAP-K), and Ras effector homolog 1 genes (RASSF1A). The prevalence for methylation of the MGMT or DAP-K genes did not differ between workers and controls, while a higher prevalence for methylation of the RASSF1A gene was seen in tumors from controls. In marked contrast, the prevalence for methylation of p16, a key regulator of the cell cycle, was increased significantly (P = 0.03) in tumors from workers compared with non-worker controls. Stratification of plutonium exposure into tertiles also revealed a striking dose response for methylation of the p16 gene (P = 0.008). Workers in the plutonium plant where exposure to internal radiation was highest had a 3.5 times (C.I. 1.5, 8.5; P = 0.001) greater risk for p16 methylation in their tumors than controls. This increased probability for methylation approximated the 4-fold increase in relative risk for adenocarcinoma in this group of workers exposed to plutonium. In addition, a trend (P = 0.08) was seen for an increase in the number of genes methylated (> or =2 genes) with plutonium dose. Here we demonstrate that exposure to plutonium may elevate the risk for adenocarcinoma through specifically targeting the p16 gene for inactivation by promoter methylation.