槲皮素对牙龈卟啉单胞菌脂多糖刺激的人牙龈成纤维细胞生物学行为的影响研究

目的研究槲皮素对牙龈卟啉单胞菌脂多糖(Porphyromonas gingivalis lipopolysaccharide,P.g LPS)刺激的人牙龈成纤维细胞(human gingival fibroblasts,HGFs)生物学行为的影响。方法采用DNA片段凝胶电泳、CCK-8法、细胞划痕法观察不同浓度槲皮素(10、20、50、100μmol/L)对体外培养HGFs的毒性作用,以及对HGFs增殖与迁移的影响。采用P.g LPS刺激HGFs来建立体外炎症刺激模型,通过流式细胞术、细胞活性氧(reactive oxygen species,ROS)检测、酶联免疫吸附试验(ELISA)和实时荧光定量PCR(qRT-PCR)进一步观察槲皮素对HGFs凋亡、ROS的产生及肿瘤坏死因子-α(tumor necrosis factorα,TNF-α)和前列腺素E2(prostaglandin E2,PGE2)表达的影响。结果槲皮素对HGFs无毒性作用,且对HGFs的增殖无影响(P>0.05)。各组细胞迁移率总的比较,差异有统计学意义(F=9.973,P<0.05),在处理48 h后,20μmol/L槲皮素处理组细胞迁移率大于10、50μmol/L槲皮素处理组以及对照组,差异均有统计学意义(均P<0.05)。槲皮素对P.g LPS刺激的HGFs凋亡具有抑制作用,且其能够抑制并预防P.g LPS刺激的HGFs中ROS相对产生量增加现象(均P<0.05)。槲皮素处理显著抑制了P.g LPS诱导的TNF-α表达增加现象(P<0.05),而槲皮素处理组PGE2的表达水平与对照组比较,差异无统计学意义(P>0.05)。结论浓度为20μmol/L的槲皮素能够促进体外培养HGFs的迁移,且具有抗氧化、抗炎保护作用。

Effects of quercetin on biological behavior of human gingival fibroblasts stimulated by Porphyromonas gingivalis lipopolysaccharide

Objective　To study the effects of quercetin on the biological behavior of human gingival fibroblasts（HGFs）stimulated by Porphyromonas gingivalis lipopolysaccharide（P.g LPS）. Methods　DNA fragment electrophoresis，CCK-8 and wound healing method were used to observe the effect of different concentrations（10，20，50，100 μmol/L）of quercetin on cell toxicity，cell proliferation，and cell migration of HGFs. Then we further stimulated HGFs with P.g LPS to create an in vitro inflammatory model；by applying flow cytometry，reactive oxygen species（ROS）test，enzyme-linked immunosorbent assay（ELISA）and realtime quantitative PCR（qRT-PCR），we tested the influence of quercetin on cell apoptosis and ROS production，as well as on TNF-α and PGE2 expression. Results　No toxic effect or proliferation influence of quercetin was observed on HGFs（P > 0.05）. There was a significant difference in overall cell migration rate among each group（F = 9.973，P < 0.05）. After 48 h of treatment，the group with quercetin at 20 μmol/L showed higher cell migration rate than the 10，50 μmol/L-quercetin treatment group and control group（P < 0.05）. Quercetin inhibited HGFs apoptosis stimulated by P.g LPS. Quercetin treatment could prevent and also inhibit the relative increase of ROS in HGFs induced by P.g LPS（P < 0.05）. Quercetin significantly inhibited the increased expression of TNF-α stimulated by P.g LPS（P < 0.05）；however，no significant difference in the expression of PGE2 was found between quercetin treatment group and control group（P > 0.05）. Conclusion　Quercetin at 20 μmol/L can promote in vitro HGFs cell migration and has anti-oxidative and anti-inflammatory protective effects.