氧化苦参碱通过自噬途径改善糖尿病大鼠心肌缺血再灌注急性肺损伤的作用机制

曾晓; 谢志斌; 许阳

doi:10.3969/j.issn.2095-5227.2020.12.011

氧化苦参碱通过自噬途径改善糖尿病大鼠心肌缺血再灌注急性肺损伤的作用机制

Oxymatrine attenuates acute lung injury by myocardial ischemia reperfusion via autophagy in diabetic rats

摘要

摘要:
目的探讨氧化苦参碱(oxymatrine，OMT)通过自噬途径对糖尿病大鼠心肌缺血再灌注急性肺损伤的保护作用。
方法采用链脲佐菌素腹腔注射建立糖尿病大鼠模型。心脏行左前降支结扎，缺血30 min再灌注120 min诱导急性肺损伤。实验随机分为正常对照组(n=15)和糖尿病非手术组(n=15)、糖尿病假手术组(n=15)、模型组糖尿病+心肌缺血再灌注(ischemia/reperfusion，I/R)(n=15)、自噬抑制剂组(0.25 mg/kg，n=15)、自噬诱导剂组(0.25 mg/kg，n=15)、OMT低剂量组(12.5 mg/kg，n=15)、OMT中剂量组(25 mg/kg，n=15)、OMT高剂量组(50 mg/kg，n=15)。采用ELISA法检测大鼠心肌组织匀浆中肌酸激酶同工酶(creatine kinase isoenzyme，CK-MB)含量变化；采用Western-blotting检测给予自噬抑制剂3-甲基腺嘌呤、诱导剂雷帕霉素、不同浓度氧化苦参碱(12.5 mg/kg、25 mg/kg、50 mg/kg)后自噬相关蛋白LC-3、Beclin-1、Atg5和 P62的表达，以观察急性肺损伤状态自噬对糖尿病肺损伤的影响。
结果模型组(糖尿病 +I/R 组)大鼠心肌缺血再灌注引起CK-MB含量升高至(5 770.67±252.55) U/L，明显高于正常对照组(1 566.67±404.15) U/L 和糖尿病非手术组(3 566.67±1 401.19) U/L；模型组肺组织HE染色显示肺泡内和肺间质水肿、出血、炎症细胞浸润。肺组织损伤评分：正常对照组0.83±0.69，糖尿病假手术组1.00±0.58，糖尿病非手术组2.00±0.58，模型组(糖尿病+I/R 组)3.88±0.37，模型组显著高于前三组（P<0.05）。模型组的肺功能显著降低，肺组织损伤显著加重，肺组织湿/干比增高，自噬相关蛋白LC-3 Ⅱ/LC-3 Ⅰ、Beclin-1、Atg5表达显著上调，P62表达下调。OMT(12.5 mg/kg、25 mg/kg、50 mg/kg)呈浓度依赖地改善糖尿病大鼠心肌I/R肺损伤。
结论 OMT可减轻糖尿病心肌I/R大鼠肺损伤，其机制可能与抑制自噬有关。

Abstract:
Objective To investigate the protective effect of oxymatrine on acute lung injury induced by myocardial ischemia/reperfusion (I/R) in diabetic rats via autophagy pathway.
Methods Diabetic rat model was established by intraperitoneal injection of streptozotozin. Then, the left anterior descending artery was ligated. Acute lung injury was induced by ischemia for 30 min and reperfusion for 120 min. Rats were randomly divided into the following groups: control group, diabetes group, diabetes+sham group, diabetes+I/R group, diabetes+I/R+rapamycin group, diabetes+I/R+3-methyladenine group, diabetes+I/R+oxymatrine group (12.5 mg/kg, 25 mg/kg, 50 mg/kg). ELISA method was used to determine the level of CK-MB content in tissue homogenate. Autophagy-related proteins LC-3 Ⅱ/LC-3 Ⅰ, Beclin-1, Atg5 and P62 were detected by western blotting.
Results The results showed that myocardial I/R in diabetic rats caused a significant increase in CK-MB content, and it was 1 566.67±404.15 in control group, 3 566.67±1 401.19 in diabetic group, and 5 776.67±252.55 in diabetic+I/R group, respectively. HE staining showed alveolar and interstitial edema, hemorrhage and inflammatory cell infiltration. The lung tissue injury scores were 0.83±0.69 in control group, 1.00±0.58 in sham group, 2.00±0.58 in diabetic group and 3.88±0.37 in diabetic+I/R group, respectively. Lung function deteriorated significantly, the ratio of WET/DRY in lung tissue increased, the expressions of autophagy-related proteins LC-3 Ⅱ/LC-3 Ⅰ, Beclin-1, Atg5 were significantly up-regulated and P62 expression was down-regulated. OMT reversed myocardial I/R induced lung injury concentration dependently in diabetic rats.
Conclusion OMT can alleviate myocardial I/R induced lung injury in diabetic rats, which may be related to inhibition of autophagy.

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