前B细胞克隆增强因子在急性呼吸窘迫综合征大鼠肺组织中表达及对炎症因子TNF-α、IL-1β、NF-κB p65表达影响的研究

Expression of pre-B-cell colony enhancing factor and its influence on the expression of PBEF,TNF-α,IL-1β and NF-κB p65 in pulmonary tissues of rats with ARDS

目的:观察前B细胞克隆增强因子(pre-B cell colony enhancing factor,PBEF)在急性呼吸窘迫综合征(acute respiratory distress syndrome,ARDS)大鼠中肺组织的表达,以及对ARDS大鼠肺组织炎症因子肿瘤坏死因子-α(tumor necrosis factor-α,TNF-α)、白细胞介素-1β(interleukin-1β,IL-1β)mRNA以及核因子-κB(nuclear factor-κB,NF-κB)p65蛋白表达的影响,通过以上指标的变化,结合已有体外实验所观察到的现象试图证实前B细胞克隆增强因子在ARDS中的促炎作用。方法:将40只成年SD大鼠随机分为空白对照组(C组)、模型组(OA组)、药物干预组(D组)、溶媒对照组(S组),OA组、D组及S组大鼠用油酸尾静脉注射复制ARDS模型,D组造模前腹腔内注射PBEF抑制剂FK866,S组造模前注射等体积FK866溶媒二甲基亚砜。造模成功6 h后取材,比较各组大鼠肺组织PBEF、TNF-α、IL-1βmRNA以及NF-κB p65蛋白表达情况。结果:C组大鼠肺组织中几乎见不到PBEF表达,而在其他各组大鼠肺组织肺泡壁及肺泡水肿液中、支气管黏膜上皮及血管内皮均可发现PBEF蛋白分布;OA组、D组和S组大鼠PBEF及炎症因子mRNA和NF-κB p65蛋白表达增加(PBEF:P=0.000,P=0.000,P=0.000;TNF-α:P=0.035,P=0.000,P=0.000;IL-1β:P=0.000,P=0.000,P=0.000;NF-κB p65:P=0.000,P=0.000,P=0.000),D组大鼠肺组织PBEF、炎症因子mRNA和NF-κB p65蛋白表达较OA组和S组低(PBEF:P=0.002,P=0.006;TNF-α:P=0.004,P=0.001;IL-1β:P=0.001,P=0.015;NF-κB p65:P=0.001,P=0.000)。结论:PBEF可能通过激活炎症反应、促进炎症因子表达等途径造成肺组织的炎性损伤,进而在ARDS发生发展中起重要作用。

Objective：To investigate the influence of pre-B cell colony enhancing factor（PBEF）on the mRNA expression of PBEF,tumor necrosis factor-α（TNF-α）,interleukin-1β（IL-1β）as well as the protein expression of nuclear factor-κB（NF-κB）p65 in lung tissue in rats with acute respiratory distress syndrome（ARDS）induced by oleic acid tail vein injection and drug intervention;to potentially verify the pro-inflammation role of PBEF in ARDS and to provide new ideas and evidences for further study of the pathogenesis,pathophysiological processes and treatment of ARDS by referring to the phenomena observed by other scholars in vitro.Methods：Forty healthy adult male SD rats were randomly divided into control group（group C）,model group（group OA）,drug intervention group（group D）,and solvent control group（group S）. Rats in group OA,group D and group S were used to induce ARDS models by oleic acid tail vein injection at a dose of 0.15 ml/kg. Rats in group D received PBEF inhibitor FK866 10 mg/kg and group S also received the same volume of its solvent dimethyl sulfoxide by intraperitoneal injection. Whether the models were successfully made was preliminary determined by rats＇ respiratory rates,color of skin and lips as well as their activity. Rats were anesthetized by intraperitoneal injection of 3.5% chloral hydrate（10 ml/kg）six hours after successfully modeling. The right upper lobe of lung was immediately doused in formalin for immunohistochemical detection and the right lower lobe was immediately frozen at-80 ℃ refrigerator or liquid nitrogen for detecting the expression of PBEF,TNF-α,IL-1β mRNA by RT-qPCR,as well as the protein expression of NF-κB p65 by Western blot. Results：The protein expression of PBEF could be found in the bronchial epithelia,vascular endothelia,alveolar wall and alveolar edema fluid of rats with ARDS by immunohistochemical detection. The expression of PBEF,TNF-α,IL-1β mRNA and NF-κB p65 protein significantly increased in all groups except group C（PBEF：P=0.000,P=0.000,P=0.000;TNF-α：P=0.035,P=0.000,P=0.000;IL-1β：P=0.000,P=0.000,P=0.000;NF-κB p65：P=0.000,P=0.000,P=0.000）. Rats in group D had an ameliorated change in the expression of inflammatory factors mRNA and NF-κB p65 protein compared with those in group OA and group S（PBEF：P=0.002,P=0.006;TNF-α：P=0.004,P=0.001;IL-1β：P=0.001,P=0.015;NF-κB p65：P=0.001,P=0.000）. Conclusion：PBEF may induce the inflammatory injury of pulmonary tissues by activating the inflammatory response and promoting the expression of inflammatory cytokines.Thus it may play an important role in the development of ARDS.