P33 Effects of hydrogen sulfide on apoptosis protein and antioxidant enzymes in the lungs from allergic mice

Abstract

Oxidative stress plays an important role in the pathogenesis of airway allergic. In allergic asthma, activated eosinophils and macrophages produce large amounts of reactive oxygen species (ROS) that may contribute to tissue injury. ROS were also involved in apoptotic pathway which includes enhanced ROS production by eosinophils. On the other hand, antioxidants agents can prevent spontaneous eosinophil apoptosis involving a major role for oxidative stress in eosinophil apoptosis. Apoptosis pathways can be initiated by diverse stimuli but all of them converge into common mechanisms involving cleavage of specific aspartic acids (caspases), as caspases 3, 6, 7, 8 and 9. Activation of these caspases results in activation of executioner caspases, such as caspase-3, that go ahead into cell apoptosis. Caspase-3 has been shown to be an important effector caspase in eosinophils following mitochondrial activation involving the Bcl-2 family protein Bax [1] . Our previous study showed that the pre-treatment of OVA-sensitized mice with hydrogen sulfide (H2S)-donor, NaHS, resulted in significant reduction of eosinophil migration to the lungs and increase in superoxide dismutase (SOD), glutathione peroxidase (GPx) and glutathione reductase (GR) antioxidant enzyme activities [2] . We, herein, investigated the effect of H2S on the expression of apoptotic proteins caspase-3 and Bax and on the level of reduced glutathione (GSH) and oxidized glutathione (GSSG) in the lungs of allergic mice. BALB/c mice were subcutaneously sensitized with ovalbumin (OVA) and, 7 days later, were intranasally challenged with OVA twice-daily for 2 consecutive days. Half of the challenged mice were treated with NaHS or with the irreversible inhibitor of CSE enzyme, propargylglicine (PAG), 30 min before the OVA challenge. All the mice were killed 48 h after the first challenge; the lungs were removed and homogenized in buffer phosphate containing a protease inhibitor cocktail. Caspase-3 and Bax protein expression were assessed by Western blotting and GSH and GSSG levels by HPLC. Results demonstrated that the treatment of allergic mice with NaHS increased the GSH levels into lungs, but GSSG was not affected. NaHS-treatment also produced significant reduction of caspase-3 expression and this effect was reverted by treatment of allergic mice with PAG. The expression of protein Bax was modified neither by NaHS nor by PAG treatments. Our results suggest that the H2S could prevent allergic lungs damage through antioxidant and antiapoptosis effects. As the GSH is considered one of the most important antioxidant agents against free radicals, the decrease in caspase-3 expression can be a consequence of this enzyme activity in the lung of allergic mice.