利拉鲁肽对脓毒症小鼠急性肺损伤的保护作用及其机制

Protective effect of liraglutide on acute lung injury in septic mice and its mechanism

目的探讨利拉鲁肽对脓毒症小鼠急性肺损伤的保护作用及其可能机制。方法选取8-12周龄、健康清洁级FVB/NJ雄性小鼠36只,随机(随机数字法)分为正常对照组、急性肺损伤组(Acute lung injury group,ALI组)和利拉鲁肽干预组(ALI+LIRA组)三组,每组12只。采用气管内注射铜绿假单胞菌混悬液制备脓毒症急性肺损伤小鼠模型,正常对照组给予气管内注射等体积生理盐水;利拉鲁肽干预组在造模0.5 h后给予利拉鲁肽(2 mg/kg)皮下注射干预,正常对照组和急性肺损伤组给予等体积生理盐水皮下注射。造模24 h后处死小鼠,收集小鼠肺组织,采用苏木精-伊红染色评估肺脏病理损伤,免疫荧光法检测肺组织表面活性蛋白D(surfactant associated protein D,SP-D)表达;收集支气管肺泡灌洗液(broncho-alveolar lavage fl uid,BALF),BCA法检测总蛋白浓度,酶联免疫吸附试验检测白细胞介素6(interleukin-6,IL-6)和肿瘤坏死因子α(tumor necrosis factorα,TNF-α)水平,Western blot法检测BALF和肺组织中SP-D蛋白表达水平。使用SPSS软件进行统计学分析,组间比较采用单因素方差分析。结果与对照组相比,ALI组小鼠的肺组织病理学损伤评分、BALF中总蛋白浓度、IL-6和TNF-α水平均升高,肺组织中SP-D阳性细胞数减少,分泌到BALF中SP-D和肺组织中的SP-D的蛋白表达下调(均P<0.05)。与ALI组相比,ALI+LIRA组小鼠的肺组织病理学损伤评分、BALF中总蛋白浓度、IL-6和TNF-α水平均下降,肺组织中SP-D阳性细胞数增加,分泌到BALF中SP-D和肺组织中的SP-D的蛋白表达上调(均P<0.05)。结论利拉鲁肽可减轻脓毒症小鼠急性肺损伤程度,其保护作用可能通过与促进SP-D分泌相关。

Objective To explore the protective effects of liraglutide on acute lung injury in septic mice and its mechanisms.Methods Thirty-six male FVB/NJ mice were randomly(random number)divided into three groups:control group(Control,n=12),acute lung injury group(ALI,n=12)and liraglutide intervention group(ALI+LIRA,n=12).Mice model of acute lung injury were prepared by intratracheal instillation of Pseudomonas aeruginosa suspension,while the control group were given intratracheal instillation of equal volume of physiological saline;the mice in ALI+LIRA group were received subcutaneous injection of liraglutide(2 mg/kg)30 minutes post-induction,while both the mice in control group and ALI group were received subcutaneous injection of equal volume physiological saline.After 24 hours,the mice were euthanized,the lung tissues and bronchoalveolar lavage fluid(BALF)were collected,the lung pathological damage changes were evaluated by hematoxylin eosin staining,the expression of surfactant associated protein D(SP-D)in lung tissue were detected by immunofluorescence assay;total protein concentration in BALF were detected by BCA method,and the levels of interleukin-6(IL-6)and tumor necrosis factorα(TNF-α)levels in BALF were measured by enzyme-linked immunosorbent assay(ELISA),the protein expression of SP-D in BALF and lung tissue were determined by Western blot.Statistical analysis was performed by SPSS software,and continuous variables were compared with one-way analysis of variance among the groups.Results Compared with the control group,the mice in ALI group had higher lung histopathology injury score,higher total protein concentration,higher IL-6 and TNF-αlevels in BALF,and had less SP-D positive cells in lung tissue;and also had lower expression of SP-D in both BALF and lung tissue,with statistical significance(all P<0.05).Compared with ALI group,the mice in ALI+LIRA group had lower lung histopathology injury score,lower total protein concentration,lower IL-6 and TNF-αlevels in BALF,and had more SP-D positive cells in lung tissue;and also had higher expression of SP-D in both BALF and lung tissue,with statistical significance(all P<0.05).Conclusions Liraglutide attenuates the severity of acute lung injury in septic mice,and its protective mechanism may be associated with the promotion of SP-D secretion.