脂联素对人牙周膜成纤维细胞的作用

The effect of adiponectin on human periodontal ligament fibroblasts in vitro

目的:原代培养人牙周膜成纤维细胞(human periodontal ligament cell,hPDLC),研究脂联素受体(adiponectinreceptor,Adipo R)在hPDLC中的表达情况,以及脂联素(adiponectin,ADP)对hPDLC增殖的作用。方法:收集因正畸需要而拔除的健康前磨牙10颗,玻片覆盖组织块法原代培养hPDLC,RT-PCR法检测分析AdipoR1和AdipoR2在hPDLC中的表达。使用不同浓度ADP(0,5,10,20ng/μL)刺激体外培养的hPDLC,采用四甲基偶氮唑盐(MTT)法,分别测定第1、4、7天的细胞增殖活性。采用SSPS16.0软件包对数据进行统计学分析。结果:免疫组化显示,培养的细胞波形丝蛋白染色阳性,角蛋白染色阴性,证明体外原代培养细胞及传代细胞均为中胚层组织来源的hPDLC,且无上皮来源细胞混杂。琼脂糖凝胶电泳检测显示,Adipo R2在人牙周膜成纤维细胞中表达阳性。MTT法检测的生长曲线显示,随着ADP浓度的增加,细胞增殖活性增加。培养24h后,C、D组与对照组相比差异显著(P<O.05);培养96h及168h后,各组与对照组相比差异显著(P<O.05)。结论:脂联素受体2在人牙周膜成纤维细胞中表达阳性,脂联素在一定浓度范围内可提高hPDLC的增殖活性。

PURPOSE： To investigate the expression of adiponectin receptor in periodontal tissues and the effect of adiponectin with different concentrations on the proliferation of human periodontal ligament fibroblast cells.METHODS： Ten human healthy premolars extracted for orthodontic reasons were collected.The primary cells were isolated from human periodontal ligament and cultured by tissue block method,then passaged when the cells completely covered the bottom of petri dish.The expression of AdipoR1 and AdipoR2 in human periodontal ligament fibroblasts was detected by RT-PCR.Adiponectin（0,5,10,20ng/μL）was added to petri dish in which the cells were incubated for 24 hours.The cells were then incubated with adiponectin for 7 days,and determined by MTT assay.SPSS16.0 software package was used for statistical analysis.RESULTS： By immunocytochemical method,the cells were stained positively to antibodies against vimentin,and negatively to antibodies against cytokeratin,indicating that they were external embryo mesenchymal cells,without epithelial cell mixation.Agarose gel electrophoresis showed Adipo R2 was positively expressed in human periodontal ligament fibroblasts.MTT assay showed that proper doses of adiponectin could promote proliferation of human periodontal ligament fibroblasts（P〈0.05）.The number of adherent cells of group C or group D was significantly greater than that of group A after 24 hours（P〈0.05）;Group B,group C,and group D had significantly more adherent cells than group A after 96 hours and 168 hours（P〈0.05）.CONCLUSIONS： Adiponectin receptor 2 is expressed in the periodontal tissues.The proper doses of adiponectin could promote proliferation of human periodontal ligament fibroblasts.