Elsevier

Radiotherapy and Oncology

Volume 81, Issue 2, November 2006, Pages 196-205
Radiotherapy and Oncology

Experimental radiotherapy
Systemic polyethylene glycol-modified (PEGylated) superoxide dismutase and catalase mixture attenuates radiation pulmonary fibrosis in the C57/bl6 mouse

https://doi.org/10.1016/j.radonc.2006.09.013Get rights and content

Abstract

Purpose

Since oxidative injury is implicated in radiation-induced tissue damage to the lung, we studied systemically administered polyethylene glycol (PEGylated) antioxidant enzymes (AOEs) as pulmonary radioprotectors in mice.

Methods and materials

C57/bl6 Mice received 13.5 Gy single-dose irradiation to the thorax. One cohort also received 100 μg of a 1:1 mixture of PEG-AOEs {PEG-catalase and PEG-superoxide dismutase (SOD)} intravenously, pre-irradiation and subgroups were evaluated at variable time-points for inflammation and fibrosis. Potential for AOE tumor protection was studied by thoracic irradiation of mice with Lewis lung carcinoma.

Results

At 48 h post-irradiation, control irradiated mice had marked elevations of tissue p21, Bax and TGF-β1 in lungs, not seen in irradiated, PEG-AOE-treated mice. TUNEL staining of lung sections was performed at just one time-point (24 h post-irradiation) and revealed a decrease in apoptotic cells with AOE treatment. At four months post-irradiation, these mice had significantly increased pulmonary fibrosis as measured by hydroxyproline content. Mice treated with PEG-AOE prior to irradiation had 4-month hydroxyproline levels that were similar to that of unirradiated controls (p = 0.28). This corresponded to less pulmonary fibrosis as visualized histologically when compared with mice irradiated without AOEs. PEG-AOEs did not prevent post-irradiation pulmonary inflammation or lung cancer response to irradiation.

Conclusions

A mixture of PEG-SOD and PEG-CAT successfully diminished radiation pulmonary fibrosis in mice. There was also a corresponding effect on several early biomarkers of lung injury and decreased apoptosis. There were no significant effects on acute pneumonitis or tumor protection.

Section snippets

Reagents

The following reagents were used in the study: PEG-catalase and PEG-SOD (Cu/Zn-SOD from bovine erythrocytes) from Sigma (St. Louis, MO), Bradford Bio-Rad protein microassay kit from (Bio-Rad Laboratories, Hercules, CA).

Animals

Our studies used female C57/bl6 mice, a strain well characterized in the field of pulmonary radioprotection [18]. Mice were obtained from Charles River (Wilmington, MA). Mice were irradiated at the age of approximately 6–8 weeks in groups of 8 mice per irradiation round. Ten mice

Effect of PEG-SOD and PEG-catalase mixture on acute radiation pneumonitis

C57/bl6 mice develop radiation pneumonitis as early as 3 weeks post single fraction XRT (13.5 GY) (Fig. 1). Systemic PEG-AOE (100 μg/mouse for each PEGylated AOE) given i.v. at the time of radiation did not alleviate lung injury or inflammation as judged by BAL protein levels or BAL neutrophils, respectively, at any time-point investigated (1, 2, 3, 7, 14, 21 and 120 days post XRT).

Effect of AOEs on oxidative modification and apoptosis in irradiated lung tissues

Malondialdehyde (MDA) is a product of lipid peroxidation, which in turn is a result of free radical interactions

Survival

Overall, no statistically significant survival differences were observed between irradiated, PEG-AOE-treated and non-drug treated controls (p = 0.8569). Nevertheless, the overall survival rate at 4 months yielded a sufficient number of animals to allow the semiquantitative and quantitative analysis of the fibrotic status of the lungs in each experiment.

Discussion

These studies provide strong evidence that systemically delivered, non-targeted antioxidant drug therapy (PEG-SOD and PEG-CAT enzymes) can decrease some of the manifestations of radiation pneumonopathy. Specifically, several early molecular events were diminished and the important “late” effect of fibrosis was also reduced. Interestingly, however, the intermediate phase of radiation lung injury – inflammation or pneumonitis – was not affected at all. This supports the hypothesis that

Conflict of interest

The co-authors of this manuscript state that there is no conflict of interest.

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    Funded in part by American Cancer Society IRG #78-002-23 (Machtay); National American Lung Association #RG-087-N, National Institute of Health NIH-1R21CA-(118111-01) and University of Pennsylvania Research Foundation (Christofidou-Solomidou).

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