低浓度牙龈卟啉单胞菌脂多糖刺激骨唾液酸蛋白的基因表达和转录

Low concentration of Porphyromonas gingivalis lipopolysaccharide stimulates bone sialoprotein gene transcription

目的研究低浓度牙龈卟啉单胞菌(Porphyromonas gingivalis,P.g)脂多糖(Lipopolysaccharide,LPS)对成骨细胞系(ROS17/2.8)中骨唾液酸蛋白(Bonesialoprotein,BSP)基因表达和转录的调节作用,以及细胞外信号调节蛋白激酶(ERK1/2)转导途径阻断剂(U0126)对此调节作用的影响。方法0.01mg/LP.g·LPS作用ROS17/2.8细胞0h、3h、6h和12h后,用Northern杂交观察BSP和骨桥蛋白(Osteopontin,OPN)mRNA的表达;再将ROS17/2.8细胞随机分为四组:空白对照组、LPS(0.01mg/LP.g·LPS)组、U0126(5μmol/L)组、U0126+LPS组(U0126预刺激细胞30min后,U0126与P.g·LPS共同作用),各组持续作用12h后,用瞬时转染法分析BSP基因启动子的转录活性。结果0.01mg/LP.g·LPS作用ROS17/2.8细胞12小时后BSP和OPN的mRNA杂交条带增强;0.01mg/LP.g·LPS使BSP基因启动子(pLUC3)转录活性值与空白载体活性值的比值升高1.582(F=5.734,P<0.05),U0126使其降低2.693(F=11.500,P<0.01),U0126使LPS对比值的升高变化降低2.242(F=6.204,P<0.05)。结论低浓度(0.01mg/L)P.g·LPS增强ROS17/2.8细胞BSP基因表达和转录,而且其对BSP基因转录活性的上调作用是经由ERK1/2信号转导路径介导的。

Objective To investigate the effects of lower concentration of Porphyromonas gingivalis lipopolysaccharide （ P.g. LPS） on expression of bone sialoprotein （BSP） in ROS17/2.8 cells. Methods ROS17/2.8 cells stimulated by 0.01mg/L P.g. LPS for Oh, 3h, 6h, 12h, BSP and osteopontin （OPN） mRNA expressions were examined by Northern blot. ROS17/2.8 ceils were divided into four groups randomly： control group, LPS （0.01 mg/L P. g. LPS） group, U0126 （5μmol/L） group and U0126 ＋ LPS group （pretreated with U0126 for 30min before being stimulated by P. g. LPS together）. All of the cells were treated for 12h, and the transcription activities of BSP gene promoter were measured by luciferase assay. Results BSP and OPN mRNA expression was increased after stimulated by 0.01mg/L P. g. LPS for 12h. In addition, the ratio of transcription activity of BSP gene （pLUC3） to the empty vector was increased 1.582 times （ P 〈0.05） by 0. 01mg/L P.g. LPS, decreased 2.693 times （ P 〈0.01） by U0126. The increase of the ratio by LPS was decreased 2.242 （ P 〈 0.05） by U0126. Conclusion The stimulatory effects of lower concentration （0.01mg/L） of P.g. LPS on the BSP gene transcription might be mediated through ERKI/2 signal - transduction pathway.