“红色复合体”致病菌裂解液对人牙周膜细胞成骨分化能力的影响

Effects of Extracts from“Red Complex”Periodontal Pathogens on Osteogenic Differentiation Capacity of Human Periodontal Ligament Cells

目的:探讨“红色复合体”牙周致病菌裂解液对人牙周膜细胞成骨分化能力的影响。方法:收集因正畸需要而拔除的健康前磨牙,原代培养人牙周膜干细胞(hPDLCs);采用免疫荧光方法和成骨分化诱导实验对培养的原代hPDLCs进行鉴定;体外联合培养牙龈卟啉单胞菌、齿垢密螺旋体和福赛斯坦纳菌,建立“红色复合体”模型,扫描电镜对其进行鉴定;制备“红色复合体”裂解液,通过碱性磷酸酶试剂盒、实时定量PCR、茜素红染色,检测裂解液对hPDLCs碱性磷酸酶活性,成骨分化相关基因表达的影响;以及对hPDLCs成骨矿化能力的影响。结果:在“红色复合体”裂解液的刺激下,hPDLCs碱性磷酸酶活性,成骨分化相关基因ALP、Runx2、OPN、COL1的表达以及成骨矿化水平显著降低。结论:“红色复合体”牙周致病菌能够抑制hPDLCs的成骨分化能力。

Objective:To evaluate the effects of extracts from“Red Complex”periodontal pathogens on osteogenic differentiation capacity of human periodontal ligament cells(hPDLCs).Methods:hPDLCs were isolated and primary cultured from first premolars extracted from orthodontic patients.The primary cultured cells were verified by immunofluorescence and osteogenic induction methods.The model of“Red Complex”was set by co-culturing Pg,Td,and Tf in vitro,and was verified by SEM.Soluble bacterial extracts obtained from“Red Complex”was used to investigate the effects of“Red Complex”periodontal pathogens on the ALP activities,osteogenic related genes expression,and mineralization of hPDLCs by performing ALP assay,RT-PCR,and Alizarin Red staining.Results:Under the stimulation of 100μg/mL bacterial extracts,the ALP activities,expression of osteogenic related genes ALP,Runx2,OPN,and COL1,and mineralization of hPDLCs were significantly decreased.Conclusion:“Red Complex”periodontal pathogens inhibited the osteogenic differentiation capacity of hPDLCs.