富血小板纤维蛋白对人牙周膜成纤维细胞迁移及成骨分化的影响

摘要

目的：观察Choukroun’s富血小板纤维蛋白（ PRF）对自体人牙周膜成纤维细胞（ hPDLCs）迁移、成骨分化的影响，探讨PRF在牙周组织再生治疗中的潜能。方法：原代培养hPDLCs；制备PRF，实验分为P1组（1片PRF浸出液）、P2组（2片PRF浸出液）和对照组，划痕实验观察记录各组第24、48、72 h细胞迁移的距离；Transwell制备迁移模型，按实验分组培养24 h后结晶紫染色观察；成骨矿化诱导液制备不同浓度的PRF浸出液（ P1组、P2组），碱性磷酸酶（ ALP）试剂盒检测3、5、7 d细胞破碎液中ALP活性；成骨矿化诱导液连续培养21 d，茜素红染色后测定矿化结节面积。结果：划痕实验中P1组、P2组细胞迁移距离大于对照组（ F＝316．248、32．846和1169．847， P均＜0．001），P1组、P2组比较差异无统计学意义（P均＞0．05）。 Transwell 迁移实验中P1组、P2组与对照组比较，迁移细胞数增加（F＝742．729，P＜0．001），P1组、P2组比较差异无统计学意义（P＞0．05）。 ALP活性P1组、P2组与对照组比较差异均有统计学意义（ F＝474．202、1383．521、2317．965， P均＜0．001）， P1组、P2组比较差异无统计学意义（P均＞0．05）。 P1组、P2组与对照组矿化结节面积比较差异有统计学意义（F＝332．280，P＜0．001），P1组与P2组比较差异无统计学意义（ P＞0．05）。结论：PRF对hPDLCs具有促进其迁移和成骨分化的作用，提示PRF在牙周组织再生工程中具有很大的临床应用潜能。%Aim:To investigate the effects of platelet-rich fibrin( PRF) on the migration and osteogenetic differentia-tion of human periodontal ligament cells (hPDLCs), and to evaluate the potential value of PRF used in periodontal tissue regeneration .Methods:The hPDLCs were primarily cultured and the PRF was prepared .There were three groups:1 piece of PRF group (P1 group), 2 pieces of PRF group (P2 group), and control group.The cell migration distances at the 24th, 48th, 72th h were recorded under microscope in scratches experiment .Transwell was used to prepare migration mod-el, and the results were observed by crystal violet staining after 24 h.Alkaline phosphatase ( ALP) activity was detected by the kits of ALP at three time points .hPDLCs were induced differentiation by osteogenesis mineralization fluid and to ob-serve mineralized nodules after being cultured for 21 days.Results:Scratch experimental results showed that the cell mi-gration distances of the experimental groups were significantly greater than that of the control group (F=316.248,32.846, and 1 169.847, P<0.001), but there was no significant difference between two experimental groups (P>0.05).Tran-swell migrating experiment showed that compared with control group , the cells of the experimental groups migrating to the outside of the Transwell chambers were significantly more than that of control group (F=742.729,P<0.001), but there was no significant difference between the two experimental groups (P>0.05).The ALP activity of P1 group and P2 group were significantly more than that of control group (F=474.202,1 383.521,2 317.965,P<0.001), but there was no sig-nificant difference between P1 group and P2 group (P>0.05).There were significant differences in mineralized nodule ar-ea between experimental groups and control group (F=332.280,P<0.001), but there was no significant difference be-tween P1 group and P2 group(P>0.05).Conclusion:PRF could promote hPDLCs migration and osteogenetic differentia-tion, and PRF may have great potential value in clinical application of periodontal tissue engineering .