右美托咪定对急性肺损伤小鼠NLRP3炎性小体介导炎症反应的作用研究

Effect of dexmedetomidine on inflammatory response mediated by NLRP3 inflammasome in mice with acute lung injury

目的 研究右美托咪定对急性肺损伤(acute lung injury, ALI)小鼠的保护作用和机制。方法 将25只C57BL/6雄性小鼠按照随机数表法分为对照组、模型组、右美托咪定12.5μg/kg(低剂量)组、右美托咪定25μg/kg(中剂量)组和右美托咪定50μg/kg(高剂量)组。使用鼻腔给予10μg脂多糖(LPS)的方法构建急性肺损伤模型。采用酶联免疫吸附试验(ELISA)测量肺泡灌洗液(bronchoalveolar lavage fluid, BALF)中肿瘤坏死因子-α(TNF-α)、白介素-1β(IL-1β)、白介素-6(IL-6)和巨噬细胞趋化蛋白-1(MCP-1)含量。测量小鼠肺组织湿质量/干质量(wet-dry weight, W/D)值、BALF总蛋白、总细胞数、中性粒细胞数和巨噬细胞数。采用苏木精-伊红(HE)染色评价肺组织病理损伤,并评分。采用蛋白印迹评价肺组织核苷酸结合寡聚化结构域样受体蛋白3(NOD-like receptor protein 3, NLRP3)、凋亡相关斑点样蛋白(apoptosis associated speck like protein containing a CARD, ASC)、半胱氨酸蛋白酶-1(Caspase-1)p20、IL-1β p17蛋白表达水平。结果 与对照组比较,模型组小鼠BALF中TNF-α、IL-1β、IL-6、MCP-1、W/D值、BALF总蛋白、总细胞数、中性粒细胞数和巨噬细胞数均显著升高,NLRP3、ASC、Caspase-1 p20、IL-1β p17蛋白表达水平升高,肺组织表现严重的炎症反应(水肿、炎性细胞浸润),差异具有统计学意义(P<0.05)。与模型组比较,右美托咪定治疗组小鼠BALF中TNF-α、IL-1β、IL-6、MCP-1、W/D值、BALF总蛋白、总细胞数、中性粒细胞数和巨噬细胞数均显著降低,以及NLRP3、ASC、Caspase-1 p20、IL-1β p17蛋白表达水平显著降低,肺组织炎症反应明显缓解,差异具有统计学意义(P<0.05)。结论 右美托咪定对LPS诱导的ALI小鼠具有保护作用,能缓解肺损伤、肺水肿和炎症反应,并与抑制NLRP3炎性小体活化有关。

Objective To study the protective effect and mechanism of dexmedetomidine on acute lung injury in mice. Methods According to the random number table method, 25 C57BL/6 male mice were divided into the negative control group, the model group, and low, middle and high dose of dexmedetomidine groups with 12.5, 25and 50 μg/kg, respectively. The model of acute lung injury was induced by nasal administration of 10 μg LPS.ELISA was used to measure the content of TNF-α, IL-1β, IL-6 and MCP-1 in BALF. The W/D value of lung tissue, the total BALF protein, and the number of total cells, neutrophils and macrophages were measured. HE staining was used to evaluate the pathological damage of lung tissue. Western blot was used to measure the expression of NLRP3, ASC, Caspase-1 p20 and IL-1β p17 in lung tissue. Results Compared with the negative control group, the levels of TNF-α, IL-1β, IL-6 and MCP-1, W/D value, total BALF protein, and the number of total cells, neutrophils and macrophages in BALF were increased, the expression levels of NLRP3, ASC, Caspase-1p20 and IL-1β p17 in lung tissues were enhanced, and the severe inflammatory reaction such as edema and inflammatory cell infiltration in lung tissues was observed in the model group(P < 0.05). Compared with the model group, the levels of TNF-α, IL-1β, IL-6 and MCP-1, W/D value, total BALF protein, and the number of total cells, neutrophils and macrophages in BALF were decreased, the expression levels of NLRP3, ASC, Caspase-1p20 and IL-1β p17 in lung tissues were reduced, and the severe inflammatory reaction such as edema and inflammatory cell infiltration in lung tissues was alleviated in the dexmedetomidine group(P < 0.05). Conclusion Dexmedetomidine has a protective effect on LPS-induced ALI mice, which can alleviate lung injury, pulmonary edema and inflammatory reaction, and is related to inhibiting the activation of NLRP3 inflammasome.