富血小板血浆用于牙周组织再生的研究 Ⅰ、PRP的提取及对PDLFs增殖的影响

Autologous platelet-rich plasma (PRP) for local application in periodontal regeneration.Ⅰ、the isolation of PRP and its effects on the proliferation of periodontal ligament fibroblasts(PDLFs).

目的 :建立富血小板血浆 (PRP)提取的方法 ,以探讨作为自身复合生长因子来源的可行性 ,并研究其对牙周膜成纤维细胞 (PDLFs)增殖的影响。方法 :利用二次离心 ( 10 0 0rpm× 15min ;3 0 0 0rpm× 8min)分离全血 ,获得PRP。将PRP设 5个浓度组 ( 5 % ,10 % ,2 0 % ,3 0 % ,5 0 % ) ,与对照组 (不含PRP)比较 ,观察它们对PDLFs增殖的影响。结果 :所获得的血小板浓度均超过全血的 4倍 ,实验组各浓度的PRP均可促进PDLFs增殖 ,实验组间及与对照组相比差异均具有显著性 (p <0 .0 1)。当PRP浓度在 5 %～ 3 0 %时促增殖作用呈剂量依赖性。结论 :二次离心法是一种简单易行的提取PRP的方法。PRP可促进PDLFs的增殖。

Objective:To establish a new method of isolating platelet-rich plasma (PRP) and the observation of its effects on the proliferation of periodontal ligament fibroblasts(PDLFs). Method:Platelet-rich plasma was isolated by two-step centrifugation (1000rpm,15min;3000rpm,8min) from 10 adult dogs. Primary cell cultures of PDLFs were obtained from healthy third incision teeth.2×10 4 cells/well were incubated in a 96-well plate in Alpha-MEM medium containing 15% calf serum for 24 h. The medium was then removed and replaced with Alpha-MEM medium without CS and incubated for a further 48h to quiesce the cells. Then the medium was dicarded and cells were exposed to 5,10,20,30 and 50% PRP in 100μl Alpha-MEM medium for 24h. Control group was cells in serum-free Alpha-MEM medium .Cell proliferation analysis was performed utilizing MTT assay .Result:This two-step centrifugation method sequestered and concentrated platelets to a level over four times baseline whole blood . Cell proliferation was enhanced by addition of 5,10,20,30,50% PRP supernatant to the cell cultures in comparison to cells in basal medium.At the dose of 5,10,20,30% PRP promoted PDLFs growth in a dose-dependant manner.Conclusion:A convenient two-step centrifugation method of concentrating platelets was established. PRP produced by this technique can augment PDLFs proliferation.