[抗炎混合物通过调节 Beclin-1 介导的自噬作用减轻败血症诱发的大鼠急性肺损伤】。]

Q3 Medicine
Weilan Lu, Guoliang Yan, Yifan Shen, Haitao Li, Sai Wu, Tongrui Weng, Rui Zhang, Yanwen Huo
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引用次数: 0

摘要

目的研究抗炎混合物对败血症诱发的大鼠急性肺损伤(ALI)的保护作用及其可能的机制:将 40 只 Sprague-Dawley (SD) 大鼠随机分为假组、脓毒症 ALI 模型组(模型组)、3-甲基腺嘌呤(3-MA)对照组和抗炎混合物预处理组,每组 10 只。进行盲肠结扎和穿孔(CLP)以再现脓毒性 ALI 模型。假大鼠组只进行开腹和闭腹,不进行穿孔和结扎。两组大鼠均在手术前连续 3 天灌胃和腹腔注射生理盐水。3-MA 对照组在造模前连续 3 天腹腔注射生理盐水和自噬抑制剂 3-MA 15 mg/kg。抗炎混合物预处理组在造模前连续 3 天给大鼠灌胃 8.8 mL/kg 抗炎混合物[抗炎混合物的成分:大黄 15 g(后下)、黄连 15 g、黄芩 12 g、厚朴 12 g、白芷 30 g]和生理盐水。各组大鼠术后 24 小时麻醉,因腹主动脉采血而死亡。采用酶联免疫吸附试验(ELISA)检测血清炎性细胞因子白细胞介素(IL-1β和IL-6)的水平。取肺组织,然后收集支气管肺泡灌洗液(BALF),用酶联免疫吸附试验(ELISA)检测 IL-1β 和 IL-6 的水平。测量肺干湿重量比(W/D)。苏木精-伊红(HE)染色后,在光镜下观察肺组织病理学变化。用 Western 印迹法检测肺组织中自噬标记物微管相关蛋白 1 轻链 3- II/I (LC3- II/I)和 Beclin-1 蛋白的表达。透射电子显微镜观察肺组织中的自噬体:结果:与假组相比,模型组大鼠肺组织结构破坏严重,炎性细胞浸润明显,肺W/D比值、血清和BALF中IL-1β和IL-6水平明显升高,LC3- II/I和Beclin-1蛋白表达下调,自噬体增多。与模型组相比,3-甲基丙烯酸对照组大鼠的肺组织损伤更为严重,肺W/D比值以及血清和BALF中的炎性细胞因子水平进一步升高,LC3- II/I和Beclin-1蛋白的表达与假组相比仍呈下降趋势,自噬体数量少于模型组。与模型组相比,抗炎混合物预处理组的肺组织损伤较轻,炎性细胞浸润量极少,肺W/D比值明显降低(7.07±1.02 vs. 11.33±1.85,P<0.05),血清和BALF中IL-1β和IL-6水平均明显下降[血清中IL-1β(ng/L):26.04±3.86 vs. 40.83±5.46,P<0.05]。83±5.46,17.75±2.02 vs. 26.86±4.32;IL-6(ng/L):91.28±10.15 vs. 129.44±13.05,76.06±7.51 vs. 120.91±7.47,均P<0.05],LC3- II/I 比值和 Beclin-1 蛋白表达均显著增加[LC3- II/I 比值:1.23±0.02 vs. 0.60±0.02,Beclin-1 蛋白(Beclin-1/GAPDH):2.37±0.33 vs. 0.62±0.05,均 P <0.05]。此外,还观察到自噬体数量的增加:结论:抗炎混合物可改善 CLP 诱导的败血症大鼠的肺损伤并降低炎症水平,这可能是通过上调 Beclin-1 介导的自噬作用实现的。
本文章由计算机程序翻译,如有差异,请以英文原文为准。
[Anti-inflammatory mixture alleviates acute lung injury induced by sepsis in rats by modulating Beclin-1-mediated autophagy].

Objective: To investigate the protective effects of an anti-inflammatory mixture on acute lung injury (ALI) induced by sepsis in rats, as well as its possible mechanisms.

Methods: A total of 40 Sprague-Dawley (SD) rats were randomly divided into the sham group, septic ALI model group (model group), 3-methyladenine (3-MA) control group, and anti-inflammatory mixture pretreatment group, with 10 rats in each group. Cecal ligation and perforation (CLP) was performed to reproduce a septic ALI model. The rats in the sham group only underwent opening and closing the abdomen without perforation and ligation. Both groups were given saline gavage and intraperitoneal injection for 3 consecutive days before surgery. The 3-MA control group was given intraperitoneal injection of saline and autophagy inhibitor 3-MA 15 mg/kg for 3 consecutive days before modeling. The anti-inflammatory mixture pretreatment group was given 8.8 mL/kg of anti-inflammatory mixture by gavage [the composition of anti-inflammatory mixture: rhubarb 15 g (after the next), coptis chinensis 15 g, baical skullcap root 12 g, magnoliae cortex 12 g, dahurian patrinia herb 30 g] and saline intraperitoneal injection for 3 consecutive days before modeling. The rats in each group were anesthetized 24 hours after surgery and died due to abdominal aortic blood collection. Enzyme-linked immunosorbent assay (ELISA) was used to detect the levels of serum inflammatory cytokines interleukins (IL-1β and IL-6). Lung tissue was taken and then the bronchoalveolar lavage fluid (BALF) was collected, and the levels of IL-1β and IL-6 were detected by ELISA. Lung wet/dry weight (W/D) ratio was measured. After hematoxylin-eosin (HE) staining, the histopathological changes of the lungs were observed under light microscopy. Western blotting was used to detect the expression of autophagy markers microtubule-associated protein 1 light chain 3- II/I (LC3- II/I) and Beclin-1 protein in lung tissue. Autophagosomes in lung tissue were observed with transmission electron microscopy.

Results: Compared with the sham group, the rats in the model group exhibited severe destruction of lung tissue structure, with significant infiltration of inflammatory cells, the lung W/D ratio and the levels of IL-1β and IL-6 in serum and BALF were significantly increased, the expressions of LC3- II/I and Beclin-1 protein were down-regulated, the autophagosomes were more. The rats in the 3-MA control group exhibited more severe lung tissue injury as compared with the model group, the lung W/D ratio and the levels of inflammatory cytokines in serum and BALF were further increased, the expressions of LC3- II/I and Beclin-1 protein still showed a decrease tendency as compared with the sham group, and the autophagosomes were less than that in the model group. Compared with the model group, the anti-inflammatory mixture pretreatment group showed milder lung tissue injury with a minimal amount of inflammatory cell infiltration, the lung W/D ratio was significantly reduced (7.07±1.02 vs. 11.33±1.85, P < 0.05), the levels of IL-1β and IL-6 in both serum and BALF were significantly decreased [IL-1β (ng/L): 26.04±3.86 vs. 40.83±5.46 in serum, 17.75±2.02 vs. 26.86±4.32 in BALF; IL-6 (ng/L): 91.28±10.15 vs. 129.44±13.05 in serum, 76.06±7.51 vs. 120.91±7.47 in BALF, all P < 0.05], and the ratio of LC3- II/I and Beclin-1 protein expression were significantly increased [LC3- II/I ratio: 1.23±0.02 vs. 0.60±0.02, Beclin-1 protein (Beclin-1/GAPDH): 2.37±0.33 vs. 0.62±0.05, both P < 0.05]. Furthermore, an increase in the number of autophagosomes was observed.

Conclusions: The anti-inflammatory mixture improves lung injury in rats with sepsis induced by CLP and reduce inflammation levels, potentially through upregulation of Beclin-1-mediated autophagy.

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来源期刊
Zhonghua wei zhong bing ji jiu yi xue
Zhonghua wei zhong bing ji jiu yi xue Medicine-Critical Care and Intensive Care Medicine
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