白藜芦醇对牙髓卟啉单胞菌脂多糖诱导成骨细胞产生MIP-2的影响

Effect of resveratrol on the expression of MIP-2 in P.e-LPS-induced mouse osteoblasts

目的:研究牙髓卟啉单胞菌(Porphyromonas endodontalis,P.e)脂多糖(lipopolysaccharide,LPS)影响成骨细胞产生巨噬细胞炎性蛋白2(macrophage inflammatory protein-2,MIP-2)mRNA和蛋白分泌的作用,以及白藜芦醇对P.eLPS诱导分泌MIP-2产生的抑制作用。方法:以不同剂量的P.e-LPS (0～50 mg/L)作用于MC3T3-El细胞,并以20 mg/L P.e-LPS作用MC3T3-El细胞不同时间(0～48 h),采用实时反转录PCR (real-time RT-PCR)和酶联免疫吸附试验(ELISA)检测MIP-2 mRNA和蛋白的表达;采用ELISA方法检测白藜芦醇对20 mg/L P.e-LPS刺激MC3T3-El细胞24 h后MIP-2蛋白表达的影响。采用SPSS 13.0软件包对结果进行单因素方差分析和Dunnett t检验。结果:当不同剂量P.e-LPS (0～50 mg/L)刺激MC3T3-El细胞时,MIP-2 mRNA的表达和蛋白的分泌随剂量的提高而升高;其中,蛋白表达从(41.86±2.49)ng/L升高到(3126.74±158.30)ng/L,差异显著(P<0.05)。在观察时间内(0～48 h),20 mg/L P.e-LPS刺激MC3T3-El细胞后,MIP-2 mRNA的表达和蛋白的分泌随时间的延长而增高,48 h时蛋白表达量最高,为(2102.55±123.27)ng/L(P<0.01)。10 mol/L白藜芦醇预处理细胞1 h,可抑制P.e-LPS诱导的MIP-2蛋白的表达,蛋白水平从(1805.33±67.54)ng/L降低到(813.82±47.21)ng/L,差异显著(P<0.01)。结论:P.e-LPS可诱导成骨细胞表达和分泌MIP-2,白藜芦醇对P.e-LPS诱导的MIP-2蛋白分泌发挥抑制作用。

PURPOSE:To investigate the effect of lipopolysaccharides(LPS)extracted from Porphyromonas endodontalis(P.e)on the expression of macrophage inflammatory protein-2(MIP-2)mRNA and protein levels in MC3 T3-E1 cells and the influence of resveratrol on the expression of MIP-2 protein in P.e-LPS induced cells.METHODS:MC3 T3-E1 cells were treated with different concentrations of P.e-LPS(0-50 mg/L)and 20 mg/L P.e-LPS for different time(0-48 h).The expression of MIP-2 mRNA and protein was detected by real-time RT-PCR and enzyme linked immunosorbent assay(ELISA).MC3 T3-E1 cells were pretreated with resveratrol for 1 h in the presence of 20 mg/L P.e-LPS for 24 h,which was detected by ELISA.Statistical analysis was performed using one-way ANOVA and Dunnett t test with SPSS 13.0 software package.RESULTS:Treatment of MC3 T3-El cells with different concentrations of P.e-LPS(0-50 mg/L)caused a significantly increase in MIP-2 mRNA and protein expression in dose-dependent manners.The expression of MIP-2 protein increased from(41.86±2.49)ng/L to(3126.74±158.30)ng/L,and the difference was significant(P<0.05).In the observation time(0-48 h),the impact of 20 mg/L P.e-LPS on induction of MIP-2 in MC3 T3-El cells exhibited a timedependent manner.At 48 h,the maximal induction of MIP-2 protein expression was(2102.55±123.27)ng/L(P<0.01).Incubation of cells with 10μmol/L resveratrol for 1 h significantly decreased the expression of MIP-2 protein from(1805.33±67.54)ng/L to(813.82±47.21)ng/L,and the difference was significant(P<0.05).CONCLUSIONS:The results suggest that P.e-LPS may mediate MIP-2 expression in MC3 T3-E1 cells,and resveratrol has a significant inhibitory effect on this process.