脂多糖和白细胞介素-1β对人牙周膜细胞中诱导型一氧化氮合酶表达的影响

Effect of lipopolysaccharide and interleukin-1β on the expression of inducible nitric oxide synthase gene in human periodontal ligament cells

目的观察脂多糖(LPS)和白细胞介素-1β(IL-1β)对人牙周膜细胞(hPDLCs)表达诱导型一氧化氮合酶(iNOS)和一氧化氮(NO)的影响。方法应用LPS和IL-1β刺激hPDLCs后,通过实时定量PCR检测iNOS基因的表达情况,收集细胞上清液,酶联免疫吸附试验(ELISA)测定诱导后细胞中iNOS的含量变化,采用硝酸还原酶法检测NO的表达水平。结果未受刺激的hPDLCs仅能产生微量的iNOS和NO;LPS和IL-1β刺激hPDLCs后,检测到iNOS、iNOS mRNA及NO的含量随着时间和质量浓度的增加而显著增加(P<0.05),在相同时间或相同质量浓度的条件下,IL-1β单独刺激或与LPS联合刺激hPDLCs产生iNOS及NO的能力强于LPS单独刺激(P<0.05)。结论 LPS和IL-1β刺激hPDLCs可以增加iNOS和NO的表达,为动物实验中牙周局部注射LPS和IL-1β诱导iNOS和NO表达奠定实验基础。

Objective To observe the effect of lipopolysaccharide（LPS） and interleukin-1β（IL-1β） on the expression of inducible nitric oxide synthase（iNOS） gene and nitric oxide（NO） in human periodontal ligament cells（hPDLCs）. Methods After stimulating hPDLCs by LPS and IL-1β,RT-PCR had been used to identify the expression of iNOS gene.The activity of iNOS in the culture was quantitated by enzyme-linked immune sorbent assay（ELISA）.And the level of NO was determined by nitrate reductase method.Results Slight amount of iNOS and NO had been detected in hPDLCs without sitimulation,but when stimulating with LPS and IL-1β,the amount of the iNOS mRNA increased significantly in the cells in the time and dose dependent way（P0.05）.Under the stimulation with the same time or same dose,the productions of iNOS and NO stimulated by IL-1β and in combination with LPS were larger than that stimulated with LPS（P0.05）.Conclusion The expression NO and iNOS could be increased by stimulating hPDLCs with LPS and IL-1β,which may contribute to the research on injecting LPS and IL-1β at periodontal tissue of animal models.