转化生长因子β3对脂多糖诱导的成骨细胞中IL-6表达的影响

doi:10.19439/j.sjos.2017.01.005

摘要/Abstract

摘要： 目的探讨转化生长因子β3（transforming growth factor β3, TGF-β3）对成骨细胞内炎性细胞因子IL-6表达的影响,及其发挥抗炎作用的机制。方法 20 μg/mL牙髓卟啉单胞菌脂多糖（lipopolysaccharide of Porphyromonas endodontalis,P.e-LPS）刺激人成骨肉瘤细胞系MG63,构建成骨细胞炎症模型。取不同浓度（5~20 ng/mL）的人重组蛋白生长因子TGF-β3和TGF-β1,分别与20 μg/mL P.e-LPS共同作用于MG63细胞24 h后,利用实时荧光定量PCR检测细胞内IL-6 mRNA的表达,ELISA法检测培养液上清中IL-6的表达水平。以10 ng/mL TGF-β3预处理细胞30 min后,再与20 μg/mL P.e-LPS共同作用20 min,Western 印迹法检测细胞内ERK1/2蛋白磷酸化的表达。采用SPSS13.0软件包对数据进行统计学分析。结果实时荧光定量PCR结果显示,单独20 μg/mL P.e-LPS作用下,MG63细胞内IL-6的表达显著增高 (P<0.01);TGF-β1在低浓度条件下（5~10 ng/mL）对IL-6的表达无显著作用,仅在20 ng/mL时可显著抑制IL-6的表达（P<0.05）。不同浓度的TGF-β3与P.e-LPS共同作用均可显著抑制IL-6的表达 (P<0.01)。ELISA结果显示,10~20 ng/mL TGF-β3可在蛋白水平上对IL-6有显著抑制作用（P<0.05）。单独P.e-LPS作用时,可使 MG63细胞内ERK1/2蛋白的磷酸化水平升高(P<0.01);而当TGF-β3与P.e-LPS共同作用时,ERK1/2的磷酸化被抑制（P<0.05）。结论相同浓度条件下,TGF-β3比TGF-β1对成骨细胞炎症的抑制作用更为显著,ERK1/2信号机制参与了TGF-β3的抗炎过程。

关键词: 转化生长因子β, 3, 成骨细胞, 炎症, IL-6

Abstract: PURPOSE: To explore the effect of transforming growth factor β3 (TGF-β3) on IL-6 expression in inflammatory MG63, and the mechanism by which TGF-β3 exert its anti-inflammatory effect. METHODS: Cell line MG63 was stimulated by 20 μg/mL lipopolysaccharide of Porphyromonas endodontalis (P.e-LPS) to establish the inflammatory model of osteoblast. TGF-β3 or TGFβ1 varying from 5 to 20 ng/mL was added together with P.e-LPS for 24 h, then the mRNA expression of IL-6 was detected by real-time PCR, the role of TGF-β3 on IL-6 protein was further verified by ELISA. MG63 was pretreated with 10 ng/mL TGF-β3 for 30 min in RPMI 1640 medium without fetal bovine serum (FBS), then the cells were cultured for another 20 min with 20 μg/mL P.e-LPS, the phosphorylation level of ERK1/2 was measured by Western blot. Statistical analysis was performed using one-way ANOVA with SPSS13.0 software package. RESULTS: The results of real-time PCR revealed that, when MG63 was treated with 20 μg/mL P.e-LPS alone, the mRNA expression of IL-6 increased significantly(P<0.01). When TGF-β1 was added with P.e-LPS, it could barely decrease IL-6 prominently at the highest concentration (P<0.05).Whereas, the inhibition effect of TGF-β3 on IL-6 was dramatic (P<0.01), ELISA results showed that 10-20 ng/mL TGF-β3 blocked the IL-6 expression at protein level (P<0.05). 20 μg/mL P.e-LPS promoted the phosphorylation level of ERK1/2 in MG63(P<0.01), while with 10 ng/mL TGF-β3, the effect of P.e-LPS on ERK1/2 was blocked(P<0.05). CONCLUSIONS: TGF-β3 is more potent than TGF-β1 in inhibiting MG63, and ERK1/2 is involved in its anti-inflammatory effect.

Key words: Transforming growth factor β, 3, Osteoblast, Inflammation, IL-6

中图分类号:

R781.3

王贵玲, 于雅琼, 郭佳杰, 仇丽鸿. 转化生长因子β3对脂多糖诱导的成骨细胞中IL-6表达的影响[J]. 上海口腔医学, 2017, 26(1): 21-25.

WANG Gui-ling, YU Ya-qiong, GUO Jia-jie, QIU Li-hong. Inhibiting effect of transforming growth factor β3 on IL-6 expression in MG63 induced by lipopolysaccharide[J]. Shanghai Journal of Stomatology, 2017, 26(1): 21-25.

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