富血小板血浆的三次离心法制备及其对脂肪干细胞增殖、成骨矿化的影响  

Platelet-rich plasma generated via the three centrifugalization method and its impact on adipose tissue-derived stem cells proliferation and mineralization

作　　者：徐燕[1] 叶眉[1] 陈芳[1] 姜大川[1] 孙晓瑜[1] 林潇[2] 沈继龙[3] 

机构地区：[1]安徽医科大学口腔医院牙周科 [2]安徽医科大学卫生管理学院卫生管理学教研室 [3]安徽医科大学人兽共患病省重点实验室,合肥230032

出　　处：《国际口腔医学杂志》2011年第1期14-18,共5页International Journal of Stomatology

基　　金：安徽省自然科学基金资助项目(090413146);安徽省教育厅产学研基金资助项目(KJ2010A159);安徽医科大学博士科研基金资助项目(XJ200703)

摘　　要：目的探索高效制备富血小板血浆(PRP)的方法,并观察不同体积分数的PRP对Beagle犬脂肪干细胞(ADSC)体外增殖及其向成骨细胞分化的影响。方法抽取3只Beagle犬静脉血各40 mL,分别用传统的二次离心法和改良的三次离心法制备PRP,进行血小板计数分析。分别将体积分数为5%、10%、15%、20%的PRP加于Beagle犬ADSC的培养液中,于作用后48、72、96 h检测细胞的增殖状况;将ADSC分别培养在添加了体积分数为5%、10%、15%、20%的PRP和不加PRP的成骨细胞诱导液中,观察ADSC的成骨表达以及PRP对其成骨矿化的影响。结果三次离心法制备的PRP中血小板的含量明显高于二次法制备的。各组PRP促ADSC增殖作用均显著强于阴性对照组(P<0.001);不同体积分数PRP组间差异有统计学意义(P<0.001),细胞增殖在体积分数15%PRP组、96 h时达到峰值。von Kossa染色中矿化结节数量随着浓度上升而增多,体积分数20%PRP组最多。结论三次离心法制备PRP的效率高于二次离心法。PRP对Beagle犬ADSC的体外增殖有促进作用,与PRP体积分数和作用时间呈正相关关系。ADSC矿化结节的数量与PRP的体积分数呈正相关关系。Objective The study investigated an effective method of generating platelet-rich plasma（PRP） and evaluated the impact of different concentrations of PRP on adipose tissue-derived stem cell（ADSC） proliferation and mineralization in vitro.Methods 40 mL whole blood was collected from 3 healthy Beagle dogs and then PRP was separated by the two centrifugalization method and the three centrifugalization method respectively.Then the platelet was counted and analyzed.ADSC were exposed to DMEM medium with various concentrations of PRP（5%,10%,15%,20%） respectively.After incubation of 48,72,96 h,the proliferations of ADSC were measured. ADSC were cultured in osteoblasts induced medium and osteoblasts induced medium with PRP（5%,10%,15%, 20%） respectively,The osteogenetic expression of ADSC and the influence on its osteogenesis mineralization of PRP were observed.Results The number of platelet from the three centrifugalization method was more than that from the two centrifugalization method.The role on various concentrations of PRP on cell proliferation was more significant than that of negative control（P〈0.001）,the differences between various concentrations of PRP groups had a statistical significance（P〈0.001）.The group of 96 h,15% PRP reached to peak value in ADSC proliferation.The quantity of mineralization node in ADSC was maximized under the condition of osteoblasts induced medium with 20% PRP.Conclusion The three centrifugalization method was more efficient than the two centrifugalization method.PRP promoted ADSC proliferation in both the dose-and time-dependent ways while PRP improved ADSC mineralization in the dose-dependent way only.

关 键 词：富血小板血浆 脂肪干细胞 细胞增殖 成骨矿化 

分 类 号：R329[医药卫生—人体解剖和组织胚胎学]