乳铁蛋白对脂多糖激活的U937细胞TLR4信号通路相关膜分子表达的影响

Effects of lactoferrin on the expression of TLR4 signaling pathway-associated membrane molecule in LPS-activated U937 macrophages

目的:研究重组人乳铁蛋白(recombinant human lactoferrin,rh LF)对脂多糖(lipopolysaccharide,LPS)激活的U937细胞Toll样受体4(toll-like receptors 4,TLR4)信号通路相关膜分子TLR4、CD14、MD2表达的影响。方法:经佛波酯(phorbol 12-myristate 13-acetate,PMA)诱导成熟的U937细胞随机分为6组:对照组、LPS组(给予100 ng/ml LPS)、LPS+50μg/ml rh LF组、LPS+100μg/ml rh LF组、LPS+200μg/ml rh LF组、LPS+500μg/ml rh LF组,采用RT-PCR和Western blot检测各组细胞TLR4、CD14、MD2 m RNA和蛋白的表达,ELISA检测各组细胞上清液中肿瘤坏死因子-α(tumor necrosis factor-α,TNF-α)的含量。结果:LPS组U937细胞TLR4、CD14、MD2 m RNA和蛋白的表达均明显高于对照组(P<0.05),LPS+50μg/ml rh LF组三者m RNA和蛋白的表达与LPS组差异不明显(P>0.05),LPS+100μg/ml rh LF组、LPS+200μg/ml rh LF组、LPS+500μg/ml rh LF组均明显降低了LPS诱导的TLR4、CD14、MD2 m RNA和蛋白的表达(P<0.05);LPS组TNF-α的含量明显高于对照组(P<0.05),LPS+50μg/ml rh LF组、LPS+100μg/ml rh LF组、LPS+200μg/ml rh LF组、LPS+500μg/ml rh LF组均明显降低了LPS诱导的TNF-α的产生(P<0.05)。结论:rh LF通过抑制TLR4介导的LPS跨膜信号转导和炎症因子TNF-α的分泌,对LPS所致的脓毒血症具有潜在的治疗作用。

Objective：To investigate the effects of recombinant human lactoferrin（rh LF）on the expression of toll-like receptors 4（TLR4）signaling pathway-associated membrane molecule（TLR4,CD14 and MD2）in lipopolysaccharide（LPS）-activated U937 macrophages. Methods ：U937 was induced to maturation by phorbol 12- myristate 13- acetate（PMA） and then divided into six groups：control group,LPS group（100 ng/ml LPS）,LPS＋50 μg/ml rh LF group,LPS＋100 μg/ml rh LFgroup,LPS＋200 μg/ml rh LF group and LPS＋500 μg/ml rh LF group. The m RNA and protein expressions of TLR4,CD14 and MD2 were determined by RT-PCR and Western blot. The concentration of tumor necrosis factor-α（TNF-α）was detected by ELISA. Results：The m RNA and protein expression of TLR4,CD14 and MD2 increased significantly in LPS group than in control group（P0.05）. The m RNA and protein expression of TLR4,CD14 and MD2 had no significant change in LPS＋50 μg/ml rh LF group（P 0.05）,but reduced significantly in LPS＋100 μg/ml rh LF group,LPS＋200 μg/ml rh LF group and LPS＋500 μg/ml rh LF group than in LPS group（P0.05）. The concentration of TNF-αincreased significantly in the LPS group than in the control group（P0.05）. The levels of TNF-α reduced significantly in LPS＋50 μg/ml rh LF group,LPS＋100 μg/ml rh LF group,LPS＋200 μg/ml rh LF group and LPS＋500 μg/ml rh LF group than in LPS group（P0.05）.Conclusion：rh LF might have a potential protective effect on LPS induced sepsis through suppressing TLR4 transmembrane signal transduction pathway and reducing the secretion of TNF-α.