Toll样受体4对人牙周膜成纤维细胞表达p-IRAK1和p-IкB-α的影响

Effect of TLR4 on expressions of p-IRAK1 and p-IκB-α in HPDLCs stimulated by lipopolysaccharide

目的:观察Toll样受体4(toll-like receptors 4,TLR4)对人牙周膜成纤维细胞(human periodontal ligamentcells,HPDLCs)在内毒素脂多糖(lipopolysaccharides,LPS)刺激下表达磷酸化IRAK1(phospho-IRAK1,p-IRAK1)和磷酸化IкB-α(phospho-IкB-α,p-IкB-α)的影响。方法:采用Western印迹和图像分析技术,检测HPDLCs受大肠杆菌LPS(1μg/ml)刺激2.5、5、10和15min后表达p-IRAK1和p-IkB-α的水平;同时检测1∶100滴度的抗TLR4单克隆抗体对HPDLCs在1μg/mlLPS刺激下表达p-IRAK1和p-IкB-α的影响。采用SPSS10.0软件包对结果进行单因素方差分析。结果:LPS刺激HPDLCs 5min后,HPDLCs表达p-IRAK1和p-IкB-α的水平最高,条带灰度与3-磷酸甘油醛脱氢酶(GAPDH)条带的比值分别由0.054、0.19增加到0.785、0.809(P<0.05)。抗TLR4单克隆抗体预处理的HPDLCs在LPS刺激下表达p-IRAK1和p-IкB-α的水平均降低,条带灰度与GAPDH条带的比值分别由0.82、0.874降低到0.099、0.201(P<0.05)。结论:TLR4参与了HPDLCs受LPS刺激后的信号传导过程,可能介导了牙周病的发生和发展。

PURPOSE： To evaluate the effect of TLR4 on expressions of p-IRAK1 and p-IκB-α in HPDLCs stimulated by lipopolysaccharide. METHODS： Western blot and image analysis were used to detect the levels of p-IRAK1 and p- IIκB-α in HPDLCs stimulated by 1μg/ml LPS after 2.5,5,10 and 15 minutes, and to detect the levels of these proteins in HPDLCs stimulated by 1μg/ml LPS after pretreatment with monoclonal antibody of TLR4. Statistical analysis was performed with SPSS10.0 software package for one-way ANOVA. RESULTS： The expressions of p-IRAK1 and p-IκB-α in HPDLCs were enhanced after stimulation of LPS.The gray scale values of protein band at 0 minute were 0.054, 0.19 respectively, significantly increased to 0.785, 0.809 after 5 minutes （P〈0.05）. After pretreatment with monoclonal antibody of TLR4, the expressions of these proteins weakened compared with those without pretreatment. The gray scale values of protein band significantly decreased from 0.82,0.874 to 0.099,0.201, respectively （P〈0.05）. COCLUSIONS： TLR4 is involved in the signaling transduction in HPDLCs stimulated with LPS, and might be involved in the progression of periodontis.