严重烧伤早期大鼠肺泡巨噬细胞CD14的表达及调控

The expression and modulation of CD14 in pulmonary alveolar macrophage of early severe burned rats

目的探讨严重烧伤对大鼠肺泡巨噬细胞(AM)CD14膜蛋白(CD14)表达变化的影响,以及抗CD14抗体对AM产生肿瘤坏死因子α(TNF-α)的调控作用。方法20%Ⅲ度烧伤大鼠检测早期外周血内毒素(LPS)浓度,体外分离培养AM,RT-PCR方法观察膜表面CD14mRNA表达、免疫组化方法观察蛋白含量及ELISA方法观察分泌TNF-α的变化。烧伤血清刺激体外培养的正常大鼠AM,观察培养上清中TNF-α浓度变化及CD14抗体的抑制作用。结果大鼠烧伤后外周血各时相点LPS浓度均明显高于对照组,与此相对应,烧伤组大鼠AM各时相点CD14mRNA表达、蛋白含量均明显增高,AM培养上清中TNF-α浓度亦显著增高(P<0.01)。以烧伤血清与AM培养1h后,烧伤组培养上清中TNF-α浓度增加值明显高于对照组(P<0.01),而在CD14抗体存在时,TNF-α浓度增加值明显低于烧伤组(P<0.01)。结论严重烧伤后外周血LPS浓度增加,AM膜表面CD14受体亦显著增加,使LPS对免疫系统的激活作用显著增大,AM分泌TNF-α明显增加,提示严重烧伤后可以通过调节CD14的作用而减少炎性介质的合成和分泌。

Objective To observe the role of pulmonary alveolar macrophage （AM） CD14 protein on the production of TNF-α after severely burned rats at early stage. Methods SD rats were burned with 20% TBSA Ⅲ injury and the dynamic changes of plasma level of LPS was observed. After AMs of burned rats were isolated and cultured, level of CD14 mRNA expression and protein of AMs produced by AMs were detected with RT-PCR, immunohistochemical and ELISA method respectively. In vitro, level of TNF-α in the supernatant of normal rat AMs cultured with postburn serum with or without CD14 antibody were also detected. Results Level of plasma LPS increased significantly after severe burn injury. Correspondingly, the expression of CD14 mRNA and protein of AMs and level of TNF-α in the supernatant of AMs also increased. When AMs were cultured with postbum serum, the level of TNF-α in the supematant increased significantly, which could be reversed at the present of CD14 antibody. Conclusions After severe burn injury, the role of LPS stimulation to immune system augments because of the increase of plasma I,PS level and AMs CD14 receptor numbers, resulting in the increase of TNF-α secretion. These indicates that the production of proinflammatory cytokines may be inhibited via modulating CD14 signal transduction.