乏白细胞富血小板血浆与富白细胞富血小板血浆对兔牙髓间充质干细胞增殖和成骨分化的影响

Different effects of leukocyte-poor platelet-rich plasma and leukocyte-rich platelet-rich plasma on proliferation and osteogenic differentiation of rabbit dental pulp mesenchymal stem cells

目的比较乏白细胞富血小板血浆(以下称乏白细胞血浆)与富白细胞富血小板血浆(以下称富白细胞血浆)对兔牙髓间充质干细胞增殖和成骨分化的影响。方法从新西兰大白免门牙分离、培养并鉴定牙髓间充质干细胞,经鉴定所提取的牙髓细胞纯度较高,符合间充质干细胞要求。采集大白免耳中动脉血,制备兔源性乏白细胞血浆与富白细胞血浆。经鉴定乏白细胞血浆中白细胞平均密度为<100/μL、血小板平均密度为9. 46×105/μL,富白细胞血浆中白细胞平均密度为9. 89×103/μL、血小板平均密度为9. 42×105/μL。分别采用乏白细胞血浆、富白细胞血浆、乏血小板血浆培养牙髓间充质干细胞(分别设为乏白细胞组、富白细胞组、对照组),观察各组细胞增殖率及牙髓间充质干细胞标志物NS、成骨分化标志物Runx-2相对表达量。结果两组同浓度乏白细胞或富白细胞血浆培养时细胞增殖率比较无统计学差异(P均> 0. 05);当乏白细胞或富白细胞血浆浓度为0~10%时,两组细胞增殖率随血浆浓度增加而升高;当乏白细胞或富白细胞血浆浓度为15%时,两组细胞增殖率均进入平台期。乏白细胞组和富白细胞组NS、Runx-2相对表达量均高于对照组,且乏白细胞组Runx-2相对表达量高于富白细胞组(P均<0. 05)。结论乏白细胞血浆较富白细胞血浆在促牙髓干细胞成骨分化方面更具优势,临床修复骨缺损时建议使用乏白细胞血浆。

Objective To compare the effects of leukocyte-rich platelet-rich plasma and leukocyte-rich platelet-rich plasma on proliferation and osteogenic differentiation of rabbit dental pulp mesenchymal stem cells.Methods The dental pulp mesenchymal stem cells were isolated,cultured,and identified from New Zealand white teeth.The extracted dental pulp cells were highly purified and met the requirements of mesenchymal stem cells.The middle ear artery blood was taken,and rabbit-derived leukocyte-poor platelet-rich plasma and leukocyte-rich platelet-rich plasma were prepared,and the white blood cell and platelet density were identified.Two different platelet-rich plasmas were used to treat dental pulp mesenchymal stem cells,and their proliferative capacity and osteogenic differentiation were observed.Results The aver-age density of leukocytes from whole blood was 4.67×103/μL,the average density of platelets was 3.05×105/μL,the average concentration of leukocytes in leukocyte-poor platelet-rich plasma was<100/μL,and the average density of plate-lets was 9.46×105/μL.The average white blood cell density in leukocyte-rich platelet-rich plasma was 9.89×103/μL,and the average platelet density was 9.42×105/μL.The proliferation rate was positively correlated with platelet-rich plas-ma concentration,when the concentration was between 0 and 10%,while it did not increase significant when the concentra-tion was over 10%.The relative expression of NS in the leukocyte-poor group and the leukocyte-rich group was higher than that in the blank control group,while the relative expression of Runx-2 was also higher than that in the blank control group.The relative expression of Runx-2 in the leukocyte-poor group was higher than that in the leukocyte-rich group(P<0.05).Conclusion Leukocytic-poor platelet-rich plasma is superior to leukocyte-rich platelet-rich plasma in promoting osteogenic differentiation of dental pulp stem cells,which is recommended to use leukocyte plasma in clinical repair of bone defects.