中华口腔医学杂志
中華口腔醫學雜誌
중화구강의학잡지
Chinese Journal of Stomatology
2011年
9期
531-536
,共6页
贾舸%仇丽鸿%李任%吕游%于雅琼%钟鸣
賈舸%仇麗鴻%李任%呂遊%于雅瓊%鐘鳴
가가%구려홍%리임%려유%우아경%종명
卟啉单胞菌,牙髓%脂多糖类%白细胞介素6%Toll样受体
卟啉單胞菌,牙髓%脂多糖類%白細胞介素6%Toll樣受體
계람단포균,아수%지다당류%백세포개소6%Toll양수체
Porphyromonas endodontalis%Lipopolysaccharides%Interleukin-6%Tool like receptors
目的 探讨CD-14和Tool样受体（Toll like receptors，TLR）在牙髓卟啉单胞菌（Porphyromonas endodontalis,Pe）脂多糖诱导小鼠成骨细胞白细胞介素6(IL-6)表达中的作用。方法用10 mg/L的Pe-脂多糖分别作用于小鼠成骨细胞MC3T3-E1不同时间，未加Pe-脂多糖的MC3T3-E1为空白对照组。反转录聚合酶链反应(RT-PCR)和酶联免疫吸附试验检测细胞IL-6基因和蛋白的表达，RT-PCR和流式细胞术检测细胞表面CD-14、TLR-2及TLR-4基因和蛋白的表达变化。抗小鼠CD-14、TLR-2及TLR-4抗体预处理细胞后，采用RT-PCR法检测Pe-脂多糖诱导细胞IL-6mRNA表达的变化。应用SPSS 11.0统计软件对结果进行单因素方差分析Dunnett-t检验。结果Pe-脂多糖作用于细胞后，与空白对照组比较细胞IL-6 mRNA和蛋白的表达明显增加，6h时IL-6表达[(36.534±0.574) ng/L]显著高于空白对照组[(11.696±0.672)ng/L)]，P＜O.01；空白对照组中CD-14和TLR-4 mRNA呈弱表达,CD-14和TLR-4的阳性细胞率分别为(39.038±3.131)％和( 11.438±0.385)％,加入Pe-脂多糖后CD-14和TLR-4 mRNA表达明显增加，CD-14和TLR-4的阳性细胞率分别增加至(62.407±1.800)％和(21.367±2.271)％，但TLR-2的表达未见明显变化；中和抗体实验证明CD-14或TLR-4抗体均可部分抑制细胞IL-6 mRNA的表达，TLR-2抗体则无此作用。结论Pe-脂多糖作用于成骨细胞MC3T3-E1诱导其炎症因子IL-6的产生依赖于CD-14和TLR-4受体，与TLR-2无关。
目的 探討CD-14和Tool樣受體（Toll like receptors，TLR）在牙髓卟啉單胞菌（Porphyromonas endodontalis,Pe）脂多糖誘導小鼠成骨細胞白細胞介素6(IL-6)錶達中的作用。方法用10 mg/L的Pe-脂多糖分彆作用于小鼠成骨細胞MC3T3-E1不同時間，未加Pe-脂多糖的MC3T3-E1為空白對照組。反轉錄聚閤酶鏈反應(RT-PCR)和酶聯免疫吸附試驗檢測細胞IL-6基因和蛋白的錶達，RT-PCR和流式細胞術檢測細胞錶麵CD-14、TLR-2及TLR-4基因和蛋白的錶達變化。抗小鼠CD-14、TLR-2及TLR-4抗體預處理細胞後，採用RT-PCR法檢測Pe-脂多糖誘導細胞IL-6mRNA錶達的變化。應用SPSS 11.0統計軟件對結果進行單因素方差分析Dunnett-t檢驗。結果Pe-脂多糖作用于細胞後，與空白對照組比較細胞IL-6 mRNA和蛋白的錶達明顯增加，6h時IL-6錶達[(36.534±0.574) ng/L]顯著高于空白對照組[(11.696±0.672)ng/L)]，P＜O.01；空白對照組中CD-14和TLR-4 mRNA呈弱錶達,CD-14和TLR-4的暘性細胞率分彆為(39.038±3.131)％和( 11.438±0.385)％,加入Pe-脂多糖後CD-14和TLR-4 mRNA錶達明顯增加，CD-14和TLR-4的暘性細胞率分彆增加至(62.407±1.800)％和(21.367±2.271)％，但TLR-2的錶達未見明顯變化；中和抗體實驗證明CD-14或TLR-4抗體均可部分抑製細胞IL-6 mRNA的錶達，TLR-2抗體則無此作用。結論Pe-脂多糖作用于成骨細胞MC3T3-E1誘導其炎癥因子IL-6的產生依賴于CD-14和TLR-4受體，與TLR-2無關。
목적 탐토CD-14화Tool양수체（Toll like receptors，TLR）재아수계람단포균（Porphyromonas endodontalis,Pe）지다당유도소서성골세포백세포개소6(IL-6)표체중적작용。방법용10 mg/L적Pe-지다당분별작용우소서성골세포MC3T3-E1불동시간，미가Pe-지다당적MC3T3-E1위공백대조조。반전록취합매련반응(RT-PCR)화매련면역흡부시험검측세포IL-6기인화단백적표체，RT-PCR화류식세포술검측세포표면CD-14、TLR-2급TLR-4기인화단백적표체변화。항소서CD-14、TLR-2급TLR-4항체예처리세포후，채용RT-PCR법검측Pe-지다당유도세포IL-6mRNA표체적변화。응용SPSS 11.0통계연건대결과진행단인소방차분석Dunnett-t검험。결과Pe-지다당작용우세포후，여공백대조조비교세포IL-6 mRNA화단백적표체명현증가，6h시IL-6표체[(36.534±0.574) ng/L]현저고우공백대조조[(11.696±0.672)ng/L)]，P＜O.01；공백대조조중CD-14화TLR-4 mRNA정약표체,CD-14화TLR-4적양성세포솔분별위(39.038±3.131)％화( 11.438±0.385)％,가입Pe-지다당후CD-14화TLR-4 mRNA표체명현증가，CD-14화TLR-4적양성세포솔분별증가지(62.407±1.800)％화(21.367±2.271)％，단TLR-2적표체미견명현변화；중화항체실험증명CD-14혹TLR-4항체균가부분억제세포IL-6 mRNA적표체，TLR-2항체칙무차작용。결론Pe-지다당작용우성골세포MC3T3-E1유도기염증인자IL-6적산생의뢰우CD-14화TLR-4수체，여TLR-2무관。
Objective To evaluate the effect of cluster of differentiation 14(CD-14) and Toll like receptors(TLR) on the expression of interleukin-6( IL-6 ) mRNA induced by Porphyromonas endodontalis (Pc) lipopolysaccharides (LPS). Methods MC3T3-E1 cells were treated with 10 mg/L Pe-LPS for different hours, and the cells uninvolved by anything as the blank group. The expression of IL-6 was detected by reverse transcription polymerse chain reaction (RT-PCR) and enzyme-liked immunosorbent assay ( ELISA ). The expression of CD-14, TLR-2 and TLR-4 mRNA was observed at different time point (0-24 h)by RT-PCR. The protein of CD-14, TLR-2 and TLR-4 was analyzed with a flow cytometer. MC3T3-E1 cells were pretreated with anti-CD-14, anti-TLR-2 and anti-TLR-4 antibody for 1 h, and then cells were stimulated with 10 mg/L Pe-LPS for 6 h. The expression of IL-6 mRNA was examined by RT-PCR.Statistical analysis was performed using one-way ANOVA Dunnett-t test with SPSS 11.0 software package.Results The IL-6 mRNA and proteins increased significantly after treatment with Pe-LPS. When MC3T3-E1 cells treated by Pe-LPS for 6 h, the expression of proteins soared from ( 11. 696 ±0. 672) ng/L to(36.534 ± 0.574) ng/L(P ＜0.01); In the control group, the CD-14 and TLR-4 mRNA are ambly-expression, and the ratios of CD-14 and TLR-4 positive cells were (39.038 ± 3. 131 )％ and ( 11. 438 ±0. 385)％ respectively in MC3T3-E1. After treatment by Pe-LPS, the expression of CD-14 and TLR-4 mRNA increased significantly, and the ratios of CD-14 and TLR-4 positive cells markedly increased to (62. 407 ± 1. 800 ) ％ and ( 21. 367 ± 2. 271 ) ％. TLR-2 expression did not change apparently after Pe-LPS treatment. The expression of IL-6 mRNA was partly inhibited by anti-CD-14 or anti-TLR-4 antibody,but not by TLR-2. Conclusions Pe-LPS can induce the expression of IL-6 in osteoblast MC3T3-E1 through CD-14 and TLR-4, but not TLR-2.
