手术创伤后Toll样受体4/髓样分化蛋白-2的表达

Expression of Toll-like receptor 4/myeoid differentiation protein-2 after surgical trauma

目的研究手术创伤对Toll样受体4(TLR4)/髓样分化蛋白2(MD2)表达的影响及其临床意义。方法设计自身配对实验,采集12例手术患者外周静脉血;用密度梯度离心法和黏附洗脱法分离单核细胞,用流式细胞仪检测单核细胞TLR4和MD2的表达;血浆脂多糖(LPS)、血浆TNFα和IL10,以及血浆TLR4和MD2含量分别用鲎试剂法、放免法和ELISA法检测。结果单核细胞TLR4和MD2在术后第1～3天显著上调表达(P<0.01),两者具有显著相关性;围手术期血浆LPS在正常范围波动(<50μg/L),血浆TNFα在各时间点差异无统计学意义(P>0.05);血浆IL10于术后第3天显著升高(P<0.01);血浆MD2在术后第3～5天显著升高(P<0.01),而TLR4仅在第3天轻度升高(P>0.05)。结论手术创伤早期可导致TLR4/MD2上调表达,致机体敏感性增强;此后血浆抑制性介质IL10和MD2升高并产生免疫抑制效应。这种双向性变化可能是大手术后全身炎症反应综合征(SIRS)和脓毒症发生的分子机制之一。

Objective To study the expression of endotoxin receptor complex,Toll-like receptor 4 (TLR4)/myeloid differentiation protein-2 (MD-2),on peripheral blood monocytes and in serum of perioperative patients and to discuss its clinical implications.Methods A self pair-matching experiment was designed and 12 cardiothoracic surgery patients without endotoxemia or infective complications were chosen.Monocytes were separated from 10 ml heparinized vein blood of patients by using the density centrifugation and adhesion-washing way,the activity and purity of cells were identified by Trypan-blue and Giemsa staining,respectively.The expression of TLR4 and MD-2 was detected by FCM.The levels of TNF-alpha and IL-10 in serum were examined by radioimmunoassay.The plasma LPS was detected with limulus lysate test.The concentration of TLR4 and MD-2 was determined by ELISA.Results[WT5”BZ] The activity and purity of monocytes was (97.5± 5.2)% and (96.5± 4.7)%,respectively.TLR4 and MD-2 proteins were significantly up-regulated at day 1 to day 3 after surgical trauma (P< 0.01) and their expression tendency showed a transparent relativity.The levels of plasma LPS varied in normal range in perioperative patients (<0.5 EU/mL).The changes of serum TNF-alpha were not significant (P> 0.05) while a marked increased level of IL-10 was found at day 3 after operation (P< 0.01).The increased serum level of MD-2 was found at 3 to 5 day after operation,but TLR4 just was slightly elevated at the third day after operation.Conclusions[WT5”BZ] TLR4/MD-2 on monocytes was up-regulated after surgical trauma so that host's sensitivity to inflammatory factors was enhanced firstly.But host immune function was then suppressed because of the increased suppressive medium such as MD-2 and IL-10 in serum.Taking together,our data suggested that this kind of dysfunction of immune system might be one molecular pathogenesis of systemic inflammatory response syndrome (SIRS)/sepsis after major surgery operation.