乙二胺四乙酸和透明质酸对牙周膜成纤维细胞活力和细胞因子表达的影响

Effect of EDTA and hyaluronic acid on the activity of periodontal ligament fibroblasts and the expression of cytokines

目的:研究乙二胺四乙酸(ethylenediaminetetraacetic acid,EDTA)和透明质酸(hyaluronic acid,HA)对牙周膜成纤维细胞(periodontal ligament fibroblasts,PDLFs)活力和细胞因子表达的影响。方法:选取因正畸拔除的智牙12颗,制备成5 mm×4 mm大小根片,按照处理方式不同分为EDTA组、HA组、EDTA+HA组和未处理组。将其置于孔板中并在根片上接种PDLFs,于接种24、48 h后,采用MTT法检测各组细胞增殖活力,显微镜下观察PDLFs的黏附情况。利用ELLISA法和Western免疫印迹法测定PDLFs产生的炎性细胞因子IL-6、IL-8、IL-1β和TNF-α水平,采用SPSS 22.0软件包对数据进行统计分析。结果:接种24、48 h后,与未处理组相比,各处理组均能促进PDLFs增殖和黏附,且联合处理促细胞增殖和黏附效果优于单一处理,差异均具有统计学意义(P<0.05);EDTA组和HA组促细胞增殖和黏附效果随接种时间的延长而增加,但组间相比无显著差异(P>0.05)。ELLIS A法和Wes tern免疫印迹检测结果显示,各处理组较未处理组均能抑制炎性因子IL-6、IL-1β和TNF-α表达,促进IL-8表达,且EDTA+HA组的作用效果更明显,差异有统计学意义(P<0.05)。结论:EDTA+HA能显著促进牙周膜成纤维细胞生长和抑制炎性细胞因子表达,可能与其增强成纤维细胞黏附力、提高其生存能力有关。

PURPOSE:To investigate the effect of ethylenediaminetetraacetic acid(EDTA)and hyaluronic acid(HA)on the activity of periodontal ligament fibroblasts(PDLFs)and the expression of cytokines.METHODS:Twelve wisdom teeth extracted due to orthodontics treatment were selected and prepared into 5 mm×4 mm root pieces,then divided into EDTA group,HA group,EDTA+HA group and untreated group according to different treatment methods.They were placed in the well plate and PDLFs inoculated on the root piece.After 24 and 48 h of inoculation,the cell proliferation of each group was detected by MTT assay.The adhesion of PDLFs was observed under microscope.The inflammatory cytokines(IL-6,IL-8 and IL-1βand TNF-α)levels produced by PDLFs were determined by ELLISA and Western Blot.SPSS 22.0 software package was used for statistical analysis.RESULTS:After 24 h and 48 h inoculation,all the treatments promoted cell proliferation and adhesion of PDLFs compared with the untreated group,and the combined treatment promoted cell proliferation and adhesion significantly better than single treatment(P<0.05).The cell proliferation and adhesion effect of EDTA group and HA group increased with the inoculation time,without significant difference between the groups(P>0.05).The results of ELLISA and Western blot showed that compared with the untreated group,the treatment groups inhibited the expression of inflammatory factors IL-6,IL-1βand TNF-α,and promoted the expression of IL-8,and the effect of EDTA+HA group was much more significant(P<0.05).CONCLUSIONS:EDTA+HA can significantly promote the growth of periodontal ligament fibroblasts and inhibit the expression of inflammatory cytokines,which may be related to its ability to enhance the adhesion of fibroblasts and then improve their viability.