目的:探究白藜芦醇(RSV)通过减弱中性粒细胞迁移和浸润改善脂多糖(LPS)诱导的急性肺损伤(ALI).方法:将BALB/c小鼠随机分为对照组、RSV组、模型组及模型+RSV组(LPS+RSV),模型组和LPS+RSV组采用LPS诱导小鼠ALI模型,RSV组和LPS+RSV组以40 mg/kg RSV灌胃,连续灌胃7 d.末次给药后,收集小鼠支气管肺泡灌洗液(BALF),检测BALF中白细胞、中性粒细胞和蛋白含量,评估肺组织髓过氧化物(MPO)活性,H-E染色观察肺组织病理损伤,检测肺组织和BALF中炎症细胞因子、趋化因子的表达.体外培养中性粒细胞,分别以1 μg/mL LPS、1 μg/mL RSV处理细胞,流式细胞术检测中性粒细胞凋亡、趋化因子受体(CXCR2、Mac-1)表达,活性氧荧光探针(DCFH-DA)检测细胞内活性氧(ROS)水平,评估中性粒细胞趋化性和迁移能力,免疫印迹检测核因子NF-κB p65磷酸化水平.结果:与模型组比较,LPS+RSV组BALF中细胞总数、中性粒细胞数、蛋白量、TNF-α、IL-6及CXCL2表达降低,肺组织MPO活性、TNF-α、IL-1β、IL-6及CXCL2 mRNA和蛋白表达降低.与对照组比较,模型组和LPS+RSV组中性粒细胞凋亡均降低,但模型组和LPS+RSV组中性粒细胞凋亡的比较,差异无统计学意义.与模型组比较,LPS+RSV组处理10、30 min时中性粒细胞内ROS活性降低,但其他时间点ROS活性比较,差异无统计学意义.与模型组比较,LPS+RSV组中性粒细胞趋化性、迁移能力、表面表达CXCR2和Mac-1的中性粒细胞数、CXCL2 mRNA及蛋白表达、p-NF-κB p65蛋白表达量降低.结论:白藜芦醇可改善LPS诱导的急性肺损伤,可能与白藜芦醇抑制中性粒细胞迁移和浸润有关.
Objective:To explore the mechanism of Resveratrol(RSV)improving lipopolysaccharide(LPS)-induced acute lung injury(ALI)by weakened migration and infiltration of neutrophils.Methods:BALB/c mice were randomly divided into control group,RSV group,model group,and model+RSV group(LPS+RSV).In the model group and LPS+RSV group,ALI mice were induced by LPS,while mice in the RSV group and LPS+RSV group were intragastrically administered with 40 mg/kg of RSV for 7 d.After the last administration,bronchoalveolar lavage fluid(BALF)was collected to detect levels of white blood cells,neutrophils and proteins.The activity of myeloperoxidase(MPO)in lung tissues was evaluated.The pathological damage of lung tissues was observed by H-E staining.The expressions of inflammatory cytokines and chemokines in lung tissues and BALF were detected.Neutrophils were cultured in vitro and treated with 1 μg/mL of LPS and RSV,respectively.Neutrophils apoptosis and expression of chemokines receptors(CXCR2,Mac-1)were detected by flow cytometry.The level of reactive oxygen species(ROS)in cells was detected by dichlorodihydrofluorescein diacetate(DCFH-DA)probe to evaluate chemotaxis and migration ability of neutrophils.The phosphorylation level of nuclear factor(NF-κB p65)was detected by Western blotting.Results:Compared with the model group,the total count of cells,the count of neutrophils,expression level of proteins,expression levels of TNF-α,IL-6 and CXCL2 in BALF were decreased in the LPS+RSV group,and MPO activity in lung tissues,expression levels of TNF-α,IL-1β,IL-6,CXCL2 mRNA and protein were decreased.Compared with the control group,the apoptosis rate of neutrophils was decreased in the model group and the LPS+RSV group,but there was no significant difference in neutrophils apoptosis between the model group and the LPS+RSV group.Compared with the mode group,ROS activity in neutrophils was declined after 10 and 30 minutes of treatment in the LPS+RSV group,but there was no significant difference in ROS activity between the two groups at the other time points.Compared with the mode group,chemotaxis and migration ability of neutrophils,the number of neutrophils expressing CXCL2 and Mac-1 on the surface,expression of CXCL2 mRNA and protein,and p-NF-κB p65 were decreased in the LPS+RSV group.Conclusion:RSV can improve LPS-induced ALI,which may be related to the migration and infiltration of neutrophils inhibited by RSV.
刘新伟;李颖;李中霞;陈嘉宇;杨洋;林雪容
河北北方学院附属第一医院急诊科,张家口 075000
急性肺损伤;白藜芦醇;脂多糖;中性粒细胞;迁移;浸润
acute lung injury;Resveratrol;lipopolysaccharide;neutrophil;migration;infiltration
《解剖学杂志》 2023 (006)
502-507,513 / 7
2021年度河北省医学科学研究(20211689)
10.3969/j.issn.1001-1633.2023.06.007
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