右美托咪定通过α7nAChR介导的TLR4/NF-κB通路减轻脂多糖诱导的急性肺损伤

Dexmedetomidine alleviates LPS-induced acute lung injury byα7nAChR mediated TLR4/NF-κB pathway

目的探讨右美托咪定是否通过α7烟碱乙酰胆碱受体(α7nAChR)介导的Tol l样受体4(TLR4)/核因子-κB(NF-κB)通路减轻脂多糖(LPS)诱导的急性肺损伤(ALI)。方法24只Wistar大鼠随机分为对照组、急性肺损伤组、右美托咪定治疗组与α7nAChR抑制剂α-BGT组,每组6只。麻醉后,对照组腹腔注射生理盐水;急性肺损伤组股静脉注射LPS(10 mg/kg)诱导ALI模型;右美托咪定治疗组给予LPS后即刻股静脉持续输注右美托咪定[5μg/(kg.h)]至实验结束;α7nAChR抑制剂α-BGT组在输注右美托咪定前半小时腹腔注射1μg/kgα-BGT,其余处理同右美托咪定治疗组。LPS注射后12 h处死大鼠,收集血液和肺组织。HE染色观察肺组织病理学变化并进行损伤评分。抽取颈动脉血检测氧分压(PaO_(2))、碳酸氢根(HCO_(3)^(–))及乳酸(Lac)水平;测定肺组织湿/干重比(W/D)和髓过氧化物酶(MPO)活性;计数支气管肺泡灌洗液(BALF)中总细胞、中性粒细胞及巨噬细胞数;ELISA法检测血液中肿瘤坏死因子-α(TNF-α)、白细胞介素-6(IL-6)、IL-10水平;Western blotting检测肺组织中α7nAChR、TLR4、p-NF-κB蛋白表达水平。结果肺组织病理学观察见右美托咪定治疗可明显减轻LPS诱导的肺泡壁和肺组织间隔增厚以及炎性细胞浸润,降低肺损伤病理评分(P<0.01)。与对照组比较,急性肺损伤组PaO_(2)、HCO_(3)^(–)水平降低,Lac、W/D、TNF-α、IL-6、IL-10水平及MPO活性升高,总细胞、中性粒细胞及巨噬细胞计数增多,肺组织中α7nAChR、TLR4、p-NF-κB蛋白表达水平升高(P<0.01);与急性肺损伤组比较,右美托咪定治疗组PaO_(2)、HCO_(3)^(–)、IL-10水平升高,Lac、W/D、TNF-α、IL-6水平及MPO活性降低,总细胞、中性粒细胞及巨噬细胞计数减少,肺组织中α7nAChR蛋白表达水平升高,TLR4、p-NF-κB蛋白表达水平降低(P<0.01)。而右美托咪定的作用可被α7nAChR抑制剂α-BGT部分逆转。结论右美托咪定可能通过α7nAChR介导的TLR4/NF-κB通路减轻LPS诱导的ALI。

Objective To investigate whether dexmedetomidine alleviates lipopolysaccharide(LPS)-induced acute lung injury(ALI)through alpha-7 nicotinic acetylcholine receptor(α7nAChR)mediated TLR4/NF-κB pathways.Methods Twenty-four Wistar rats were randomly divided into 4 groups:control group(n=6),acute lung injury group(n=6),dexmedetomidine treatment group(n=6),andα7nAChR inhibitorα-BGT group(n=6).After anesthesia,rats in the control group were intraperitoneally injected with normal saline to serve as normal control;the rest rats all received LPS(10 mg/kg)via a femoral vein to induce the ALI model.For the dexmedetomidine treatment group,rats were continuously injected with dexmedetomidine(5μg/kg per hour)via the femoral vein immediately after LPS administration.For theα7nAChR inhibitorα-BGT group,rats received 1μg/kgα-BGT half an hour before dexmedetomidine administration as the dexmedetomidine treatment group.The rats were sacrificed 12 hours after LPS injection to collect the blood and lung tissues.Histology of the lungs was examined with HE staining.The lung injury score was calculated.In the blood sample,PaO_(2),HCO_(3)^(–),Lac,W/D,MPO activity were measured.The number of total cells,neutrophils,and macrophages in bronchoalveolar lavage fluid(BALF)were measured,and the concentrations of tumor necrosis factor-α(TNF-α),interleukin-6(IL-6),IL-10 in serum were measured by ELISA.The protein expression ofα7nAChR,TLR4,p-NF-κB were determined by Western blotting.Results Compared with the control group,LPS induced marked lung histological injury,which was less pronounced in the dexmedetomidine treatment group.Compared with the control group,the levels of PaO_(2) and HCO_(3)^(–)were decreased,Lac,W/D,TNF-α,IL-6,IL-10,MPO,the total number of cells,neutrophils,and macrophages were increased in the acute lung injury group(P<0.01).Compared with the acute lung injury group,PaO_(2),HCO_(3)^(–)and IL-10 were increased,and Lac,W/D,TNF-α,IL-6,MPO,the total number of cells,neutrophils,and macrophages were decreased in the dexmedetomidine treatment group(P<0.01).Compared with the control group,the protein levels ofα7nAChR,TLR4,p-NF-κB were increased in the acute lung injury group(P<0.01).Compared with the acute lung injury group,the protein levels ofα7nAChR was increased,and TLR4,p-NF-κB were decreased in the dexmedetomidine treatment group(P<0.01).However,the effect of dexmedetomidine was reversed by theα7nAChR inhibitorα-BGT.Conclusion Dexmedetomidine may reduce LPS-induced ALI throughα7nAChR mediated TLR4/NF-κB pathways.