MBL抑制LPS诱导DC成熟机制的研究

Mechanism of MBL Inhibiting the LPS-induced DC Maturation

本研究探讨甘露聚糖结合凝集素(MBL)对细菌脂多糖(LPS)诱导的人外周血单核细胞来源树突状细胞(DC)成熟的机制。从健康成人外周血分离单核细胞(Mo),用常规方法体外诱导未成熟DC(imDC),FACS分析MBL与imDC的结合,并进一步分析MBL对LPS结合imDC的影响;ELISA和Western blot分析MBL与可溶性的TLR4胞外段蛋白(sTLR4)结合;Western blot分析NF-κB的细胞核移位。结果表明,在钙离子存在下,MBL能够直接与imDC结合,sTLR4和LPS可竞争性抑制MBL与imDC的结合。Western blot与ELISA分析结果显示,MBL能够以浓度依赖的方式结合sTLR4蛋白。FACS分析结果进一步表明,MBL通过结合imDC竞争性抑制LPS与imDC的结合。Western blot分析结果亦显示,MBL对LPS诱导的NF-κB细胞核移位有显著的抑制作用。结论:MBL可通过结合表达在imDC的TLR4分子竞争性抑制LPS结合imDC,从而抑制LPS诱导imDC成熟,提示MBL能够通过配体结合调控DC分化成熟。本研究为阐明MBL抑制LPS诱导DC成熟的机制提供了较好的实验依据。

The study was aimed to investigate the mechanism of mannan-binding lectin （MBL） on bacterial lipopolysaccharide （LPS）-induced human peripheral blood monocyte-derived dendritic cell （DC） maturation. The monocytes were prepared from the peripheral blood of healthy adult volunteers. The immature dendritic cells （imDC） were induced by 5-day-culture in medium supplemented with rhGM-CSF and rhlL-4. FACS was used to investigate the interaction of MBL with imDC and the impact of MBL on LPS binding to imDC. ELISA and Western blot was used to analyze the interaction of MBL with soluble TLR4 ectodomain protein （sTLR4） ; Western blot was used to detect LPS-induced NF-KB translocation in imDC. The results showed that MBL could directly bind to imDC in the presence of calcium, sTLR4 protein or LPS could competitively inhibit the binding of MBL to imDC. ELISA and Western blot showed that MBL could evidently bind to sTLR4 protein in a concentrationdependent manner. FACS showed that MBL could competitively inhibit the binding of LPS to imDC by binding to imDC directly. Western blot showed that MBL decreased LPS-induced NF-KB translocation in imDC. It is concluded that MBL may competitively inhibit the binding of LPS to imDC by binding to TLR4 expressed on imDC, resulted in inhibition of LPS-induced DC maturation, suggesting that MBL can regulate DC maturation through ligand-binding. This study provides the good foundation to clarify the mechanism of MBL inhibiting the LPS-induced DC maturation.