Defining the pathway of radiation induced lung toxicity in mice overexpressing extracellular superoxide dismutase

Abstract

Radiation induced lung injury is mediated by reactive oxygen species. Superoxide dismutase is involved in the processing of these free radicals. Overexpression of extracellular superoxide dismutase (EC-SOD) has been shown to protect against radiation induced lung injury. The purpose of this study is to evaluate if the transforming growth factor beta (TGFβ)/smad3 signal transduction pathway is involved in the pathogenesis of radiation induced lung toxicity. B6C3 wildtype and transgenic mice were randomized to wildtype /no radiation (n = 3), wildtype/radiation (n = 5), transgenic/no radiation (n = 4), and transgenic/radiation (n = 6) groups. Irradiated animals received 15 Gy to the whole lung. Upon sacrifice (at 6 months), lung tissue was embedded in paraffin blocks. Immunohistochemistry(IHC) was performed to evaluate for active TGFβ1, Integrin-β6, smad-3 and phosphorylated smad3 (p-smad3). After scanning the whole lung section and calculating the extent of damaged lung parenchyma, three representative fields (20x objective) were selected and analyzed. The relative expression of Integrin-β6, smad-3 and psmad-3 were measured (positively stained cells/total cell count x 100). Active TGFβ1 was calculated as the positively stained area/total parenchymal area x 100. TGFbβ1 is elevated in WT mice undergoing whole lung RT. This expression is significantly decreased in the transgenic animals. No difference was noted in the expression of the Integrin-β6 expression in the RT groups, although, both are higher than the groups not receiving RT. Smad-3 and psmad-3 are both elevated in the wildtype RT group. Again, in the transgenic mice undergoing RT, the staining was decreased. * p =< 0.05; ∧p = 0.06 This study demonstrates that there is an alteration in the TGFβ/smad3 signal transduction pathway after whole lung irradiation. Wildtype mice express elevated active TGFβ1 after RT. This increase is ameliorated by the presence of extracellular SOD. The study shows a significant increase in smad-3 and psmad-3 staining after whole lung RT in wildtype mice. There is a significant decrease in staining in animals overexpressing extracellular SOD. This indicates that protection against oxidative stress by SOD ameliorates radiation-induced lung damage through down regulation of TGFβ/smad3 signal transduction pathway.