Abstract
Damage to normal lung tissue is a limiting factor when ionizing radiation is used in clinical applications. In addition, radiation pneumonitis and fibrosis are a major cause of mortality following accidental radiation exposure in humans. Although clinical symptoms may not develop for months after radiation exposure, immediate events induced by radiation are believed to generate molecular and cellular cascades that proceed during a clinical latent period. Oxidative damage to DNA is considered a primary cause of radiation injury to cells. DNA can be repaired by highly efficient mechanisms while repair of oxidized proteins is limited. Oxidized proteins are often destined for degradation. We examined protein oxidation following 17 Gy (0.6 Gy/min) thoracic X-irradiation in C57BL/6J mice. Seventeen Gy thoracic irradiation resulted in 100% mortality of mice within 127–189 days postirradiation. Necropsy findings indicated that pneumonitis and pulmonary fibrosis were the leading cause of mortality. We investigated the oxidation of lung proteins at 24 h postirradiation following 17 Gy thoracic irradiation using 2-D gel electrophoresis and OxyBlot for the detection of protein carbonylation. Seven carbonylated proteins were identified using mass spectrometry: serum albumin, selenium binding protein-1, alpha antitrypsin, cytoplasmic actin-1, carbonic anhydrase-2, peroxiredoxin-6, and apolipoprotein A1. The carbonylation status of carbonic anhydrase-2, selenium binding protein, and peroxiredoxin-6 was higher in control lung tissue. Apolipoprotein A1 and serum albumin carbonylation were increased following X-irradiation, as confirmed by OxyBlot immunoprecipitation and Western blotting. Our findings indicate that the profile of specific protein oxidation in the lung is altered following radiation exposure.
Highlights
Pulmonary injuries are limiting factors for the use of radiotherapy for the treatment of thoracic cancers [1,2,3,4,5,6]
We previously provided evidence that proteins in the liver and bone marrow of mice are oxidized in normal tissue and that the profile of protein oxidation is altered in response to ionizing radiation exposure, with some proteins displaying enhanced carbonylation and other proteins displaying reduced carbonylation at 24 h postirradiation [25,26]
We wished to identify proteins oxidized in the lung at 24 h following a level of radiation exposure that would be sufficient to induce delayed pulmonary fibrosis, as these oxidized proteins could contribute to early alterations in cell function and survival
Summary
Pulmonary injuries are limiting factors for the use of radiotherapy for the treatment of thoracic cancers [1,2,3,4,5,6]. The delayed onset of radiation-induced lung injury is believed to be the result of a multi-step process involving repeated episodes of inflammation followed by failed repair due to loss of regenerative properties of the normal tissue and their replacement with activated fibroblasts [17,19]. Using primary lung endothelial cell cultures, we previously demonstrated that protein oxidation by radiation can lead to endoplasmic reticulum (ER) stress, that can lead to programmed cell death and accelerated senescence within 24 h postirradiation [23]. We previously provided evidence that proteins in the liver and bone marrow of mice are oxidized in normal tissue and that the profile of protein oxidation is altered in response to ionizing radiation exposure, with some proteins displaying enhanced carbonylation and other proteins displaying reduced carbonylation at 24 h postirradiation [25,26]. The present findings show that radiation exposure alters the profile of protein carbonylation in the lung, and that there are both similarities and differences in the identified proteins that are carbonylated in the lung compared with other tissues
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