Effect of methylene blue preconditioning on ischemia-reperfusion injury in isolated rat lungs

Abstract

Objective To evaluate the effect of methylene blue(MB)preconditioning on ischemia-reperfusion(I/R)injury in isolated rat lungs. Methods Eighteen pathogen-free healthy male Sprague-Dawley rats, aged 3 months, weighing 240-320 g, were divided into 3 groups(n=6 each)using a random number table: sham operation group(group Sham), lung I/R group(group I/R)and methylene blue preconditioning group(group MB). A model of isolated lung I/R injury was established in pentobarbital sodium-anesthetized rats.MB 2 mg/kg was intraperitoneally injected at 2 h before stopping perfusion in group MB.Isolated lungs were perfused for 20 min, followed by 45-min ischemia, and then reperfused for 60 min in I/R and MB groups.At 60 min of reperfusion, the activity of lactic dehydrogenase(LDH)in the perfusate was detected, wet weight(W)and dry weight(D)was determined, W/D ratio was calculated, and the levels of malondialdehyde(MDA), ATP, reactive oxygen species(ROS)and superoxide dismutase(SOD)were measured in lung tissues.Mitochondria and cytoplasm were isolated from lung tissues for determination of mitochondrial membrane potential(MMP), degree of mitochondrial swelling and content of cytochrome C(Cyt c)in cytoplasm.Apoptotic cells in lung tissues were detected using TUNEL, and apoptotic index was calculated. Results Compared with group Sham, the activity of LDH in perfusate, W/D ratio, levels of ROS, MDA and Cyt c in cytoplasm and apoptosis index were significantly increased, the degree of mitochondrial swelling was aggravated, and the content of ATP and MMP were decreased in I/R and MB groups, and the SOD activity was significantly decreased in group I/R(P<0.05). Compared with group I/R, the activity of LDH in perfusate, W/D ratio, levels of ROS, MDA and Cyt c in cytoplasm and apoptosis index were significantly decreased, the degree of mitochondrial swelling was attenuated, and the activity of SOD, content of ATP and MMP were increased in group MB(P<0.05). Conclusion Methylene blue preconditioning can reduce I/R injury in isolated rat lungs, and the mechanism may be related to improving mitochondrial function and inhibiting cell apoptosis. Key words: Methylene blue; Ischemic preconditioning; Reperfusion injury; Lung