| LBP、sCD14与LPS相互作用对巨噬细胞释放TNF-α的影响 |
| 投稿时间：2010-08-13 修订日期：2010-12-23 点此下载全文 |
| 引用本文：张颖,舒旷怡,王金文,赵飞,刘洋,李向阳.LBP、sCD14与LPS相互作用对巨噬细胞释放TNF-α的影响[J].医学研究杂志,2011,40(2):26-30 |
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| 基金项目:浙江省自然科学基金资助项目（Y2080596）;温州市科技计划项目（Y20100094） |
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| 中文摘要:目的研究脂多糖结合蛋白（LBP）、可溶性CD14（sCD14）与脂多糖（LPS）相互作用后对巨噬细胞产生TNF-α的影响。方法用PMA诱导THP-1细胞为巨噬细胞；设定LPS浓度为1ng/ml，以100、10、1、0.1、0.01的比例，将LBP-LPS混合37℃孵育15min后分别加入细胞培养板中；以及混合后不孵育直接加入；同时设立LBP对照组。在LPS浓度为10ng/ml和1ng/ml两组中，分别加入1、0.5、0.1、0.01μg/ml的sCD14，混合37℃孵育15min后分别加入两组细胞培养板中；以及混合后不孵育直接加入；同时设立sCD14对照组。用免疫化学发光法检测TNF-α的释放情况。最后对数据进行统计学分析。结果LPS浓度为1ng/ml分泌TNF-α最低（143±9pg/ml）,与10、100、1000ng/ml间比较有统计学差异，P均<0.05；10、100、1000ng/ml 三组间无统计学差异（P>0.05）；TNF-α的释放量随着LPS的浓度增高会出现饱和现象。浓度为0.01、0.1、1、10、100ng/ml的LBP刺激巨噬细胞TNF-α的释放量分别为39.4±1.2、107±5、109±3、102±1、119±9pg/ml。LBP/LPS浓度比值≤10时,TNF-α释放量随LBP/LPS浓度比值增大而增高，在LBP/LPS浓度比值为10时释放量最高；未孵育组结果为231±10pg/ml，预孵育组结果为164±7pg/ml；两组TNF-α释放量有显著性差异（P<0.01）。在LBP/LPS比值为100时，TNF-α释放量下降；未孵育组结果为149±7pg/ml，预孵育组结果为138±2pg/ml。LPS浓度为0时，随着sCD14浓度增加，THP-1细胞释放TNF-α未见增加。LPS浓度为1ng/ml，sCD14浓度为0.01μg/ml和0.1μg/ml时，TNF-α释放量无显著性差异（P=0.515和0.566）；sCD14浓度为0.5和1μg/ml,结果有显著性差异（P=0.017和P<0.01）。在LPS浓度为10ng/ml，sCD14浓度为0.5μg/ml和1μg/ml时TNF-α释放量结果均有显著性差异（P<0.01）。未预孵育组与预孵育组TNF-α的释放量结果有显著性差异（P=0.014）。结论LBP能活化巨噬细胞，低浓度LBP可促进LPS刺激巨噬细胞释放TNF-α，且释放量随LBP浓度的增加而增加；高浓度LBP对LPS刺激巨噬细胞有负性调节作用。sCD14能促进LPS刺激巨噬细胞释放TNF-α，并表现出一定的剂量效应关系。 |
| 中文关键词:LBP sCD14 TNF-α |
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| The Effect of LBP,sCD14 Interacted with LPS on Macrophage Secreting TNF-α |
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| Abstract:ObjectiveTo investigate the effect of LBP,sCD14 interacted with LPS on macrophage secreting TNF-α. MethodsTHP-1 was induced to macrophage by PMA firstly.When the concentration of LPS was 10ng/ml,LBP-LPS pre-incubated (incubated at 37℃ for 15min after mixed together) or without pre-incubated stimulated macrophage according to the ratio of LBP/LPS (100,10,1,01,0.01).And macrophage treated with LBP was as a contrast.After that, they were divided into two groups and both of LPS were 10ng/ml and 1ng/ml,and the concentrations of sCD14 were 1,0.5,0.1,0.01μg/ml.The secretion of TNF-α was detected by immune chemical radiation assay and a statisticcal analysis was made. ResultsTNF-α secretion was the lowest(143±9pg/ml) when LPS was 1ng/ml,and had a significant difference from 10,100,1000ng/ml(P<0.05).There was no difference among the three groups(P>0.05). The secretion of TNF-α had a dose-response relationship with LPS.TNF-α were 39.4±1.2,107±5,109±3,102±1,119±9pg/ml at corresponding concentration of LBP (0.01,0.1,1,10,100ng/ml).When LBP/LPS ≤10,the secretion of TNF-α increased with the enhance ratio of LBP/LPS,and reached a peak at 10.LBP-LPS in without pre-incubated group was 231±10pg/ml，and the other was 164±7pg/ml.There was significant difference between the two groups（P<0.01）.TNF-α was decreased at 100 of LBP/LPS and without pre-incubation was 149±7pg/ml, pre-incubation was 138±2pg/ml.sCD14 itself could not stimulate macrophage to secrete TNF-α.When the concentration of LPS was 1ng/ml，sCD14 was 0.01μg/ml or 0.1μg/ml,TNF-α secretion had no visible diversity（P=0.515 and 0566）.However,When the concentration of sCD14 were 0.5 and 1μg/ml,it had a obvious diversity（P=0.017 and P<0.01）.When the concentration of LPS was 10ng/ml，sCD14 was 0.5μg/ml or 1μg/ml,TNF-α secretion had significant difference（P<0.01）,and pre-incubation and without pre-incubation also appeared significant difference（P=0.014）.ConclusionLBP itself could activate macrophage,and low concentration of LBP promoted LPS to stimulate macrophage to secrete TNF-α with the concentration of LBP increasing.But it had a down-regulation effect on macrophage when high concentration.sCD14 also promoted the secretion of TNF-α and showed a dose-response relationship with sCD14. |
| keywords:Lipoolysaccharide-binding protein Soluble CD14 Tumor necrosis factor-α |
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