Biological role and related mechanism of autophagy in acute lung injury of hemorrhagic shock mice
Objective To study the biological role and related mechanism of autophagy in acute lung injury(ALI)of hemorrhagic shock mice.Methods According to random number table method,wild-type male C57BL/6 mice were divided into control group,ALI group,rapamycin group and 3-methyladenine(3-MA)group,with 8 mice in each group.Light chain 3(LC3)gene knockout mice with C57BL/6 background were divided into LC3 knockout group and LC3 knockout+ALI group,with 8 mice in each group.Control group,ALI group,LC3 knockout group,LC3 knockout+ALI group were intraperitoneally injected with 2 mL/kg normal saline,rapamycin group was intraperitoneally injected with 3 mg/kg autophagy activator rapamycin,3-MA group was intraperitoneally injected with 15 mg/kg autophagy inhibitor 3-MA,all of which were given for 3 consecutive days.2 hours after the last administration,the hemorrhagic shock induced ALI model was established.24 hours after modeling,the lung index was calculated.Hematoxylin-eosin(HE)staining was used to observe the pathological changes of lung tissue and lung injury score was performed.The expressions of autophagy genes LC3-Ⅱ/LC3-Ⅰ and Beclin-1 in lung tissue were detected by Western blotting.The contents of tumor necrosis factor-α(TNF-α),interleukin-6(IL-6)and malondialdehyde(MDA)in lung tissue were detected according to the steps of the kit.Results Compared with the control group,the lung tissue structure was destroyed and exudation increased,lung index,lung injury score,the expressions of LC3-Ⅱ/LC3-Ⅰ,Beclin-1,and the contents of TNF-α,IL-6 and MDA in lung tissue significantly increased in the ALI group.Compared with the ALI group,the structural damage and exudation of lung tissue were reduced in the rapamycin group,lung index,lung injury score and the contents of TNF-α,IL-6 and MDA in lung tissue decreased,while the expressions of LC3-Ⅱ/LC3-Ⅰ and Beclin-1 in lung tissue increased[lung index:(7.56±0.39)％vs.(9.12±0.59)％,lung injury score:3.04±0.58 vs.9.32±2.14,TNF-α(ng/mg):1.85±0.32 vs.3.51±0.62,IL-6(ng/mg):1.61±0.32 vs.2.52±0.44,MDA(nmol/mg):1.03±0.16 vs.1.88±0.24,LC3-Ⅱ/LC3-Ⅰ:1.21±0.12 vs.0.39±0.05,Beclin-1/β-actin:1.10±0.12 vs.0.58±0.06,all P＜0.05],while lung tissue structure damage was aggravated and exudation was further increased in the 3-MA group,lung index,lung injury score and the contents of TNF-α,IL-6 and MDA in lung tissue increased,the expressions of LC3-Ⅱ/LC3-Ⅰ and Beclin-1 in lung tissue decreased[lung index:(10.44±0.62)％vs.(9.12±0.59)％,lung injury score:11.59±2.28 vs.9.32±2.14,TNF-α(ng/mg):4.77±0.71 vs.3.51±0.62,IL-6(ng/mg):3.44±0.52 vs.2.52±0.44,MDA(nmol/mg):2.71±0.42 vs.1.88±0.24,LC3-Ⅱ/LC3-Ⅰ:0.25±0.04 vs.0.39±0.05,Beclin-1/β-actin:0.21±0.03 vs.0.58±0.06,all P＜0.05].Lung index,lung injury score and the contents of TNF-α,IL-6 and MDA in lung tissue of LC3 knockout ALI mice were higher than those of wild-type ALI mice[lung index:(10.44±0.75)％vs.(9.12±0.59)％,lung injury score:12.41±2.86 vs.9.32±2.14,TNF-α(ng/mg):4.85±0.72 vs.3.51±0.62,IL-6(ng/mg):3.28±0.51 vs.2.52±0.44,MDA(nmol/mg):2.75±0.41 vs.1.88±0.24,all P＜0.05].Conclusion Autophagy plays a protective role in ALI of hemorrhagic shock mice,and the related molecular mechanism is the inhibition of inflammatory response and oxidative stress response.
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