兔牙髓干细胞与Pluronic F-127嵌段共聚物的体外相容性  

Biocompatibility of dental pulp progenitor cells and Pluronic F-127 hydrogel

作　　者：帕尔哈提.阿布肚热合曼 白尔娜.吾守尔 木合塔尔.霍加 刘晓文[1] 

机构地区：[1]新疆维吾尔自治区人民医院口腔科,乌鲁木齐830001

出　　处：《中华口腔医学研究杂志（电子版）》2016年第2期97-103,共7页Chinese Journal of Stomatological Research(Electronic Edition)

基　　金：国家自然科学基金(地区科学基金项目;81560180)

摘　　要：目的探讨牙髓干细胞(DPSC)与Pluronic F-127水凝胶生物相容性。方法酶解组织块法获得DPSC,镜下观察细胞形态。制备20%、25%、30%(w/v)的Pluronic F-127水凝胶,扫描电镜(SEM)观察水凝胶支架空间形态,检测支架材料的溶胀率,凝胶化时间。用不同浓度Pluronic F-127水凝胶包封DPSC进行三维培养,并以普通培养皿的二维培养作为对照,镜下观察细胞生长情况。培养第1、3、5、7天通过噻唑蓝(MTT)法检测支架对DPSC增殖的影响。将最佳浓度的Pluronic F-127用于包封DPSC,常规矿化液诱导14 d进行茜素红、甲苯胺蓝染色、实时荧光定量聚合酶链反应(PCR)检测成骨相关骨桥蛋白(OPN)、Ⅱ型胶原,以评价Pluronic F-127对DPSC成骨及成软骨分化的影响。数据分析采用单因素方差分析。结果成功分离并纯化DPSC,细胞在支架内生长良好;MTT结果显示:第1天各组间差异无统计学意义;第3天,20%Pluronic F-127组细胞增殖A标准值(0.774±0.095)显著高于其他培养组(A_(对照)=0.353±0.096、A_(25%PF)=0.559±0.053、A_(30%PF)=0.532±0.093),差异有统计学意义(F=15.993,P=0.000);第5天时,20%Pluronic F-127组细胞增殖A标准值(0.554±0.510)显著高于其他培养组(A_(对照)=0.260±0.097、A_(25%PF)=0.353±0.049、A_(30%PF)=0.212±0.049),差异有统计学意义(F=20.821,P=0.000)。成骨及成软骨诱导后茜素红及甲苯胺蓝染色呈阳性,对照组呈阴性。实时荧光定量PCR显示,三维诱导组m RNA相对表达水平显著高于其他培养组,差异有统计学意义(F_(OPN)=65.576,P_(OPN)=0.000;F_(COL-2)=38.382,P_(COL-2)=0.000)。结论 20%Pluronic F-127适合DPSC的培养,并可支持其生长及分化。Pluronic F-127可作为DPSC的生物载体,有望成为理想的组织工程支架材料。Objective To assess the biocompatibility of nonionic triblock copolymer Pluronic F-127 with dental pulp stem cells（DPSCs）. Methods DPSCs were harvested with enzymatic digestion method.Following the preparation of 20%,25%,30%（w/v）Pluronic F-127,Scanning electron microscopy（SEM）was utilized to observe the structure of the hydrogel,and swelling kinetics and gelling properties were examined. After the encapsulation of cells of third passage in Pluronic F-127,MTT tests were performed to asses the effect of hydrogels on the proliferation of DPSCs on the 1,3,5,7（th）day of incubation. On day 14,Osteogenic and chondrogenic differentiation ability of DPSCs encapsulated in the hydrogel were estimated by means of alizarin red S,toluidine blue staining,Real-time q-PCR. Data were analyzed using one-way ANOVA procedure. Results DPSCs were isolated and encapsulated in the hydrogel scaffold successfully,MTT test shows that There was no significant difference of cell proliferation in the 1st day. 20% PF group（0.774 ± 0.095）enhanced cell proliferation significantly higher than other groups（A_（control）= 0.353 ± 0.096,A_（25% PF）= 0.559 ± 0.053,A_（30% PF）= 0.532 ± 0.093）in the 3（rd） day（F = 15.993,P = 0.000）,in the 5（th） day,20%PF group（0.554 ± 0.510）also significantly enhanced cell proliferation than other groups（A_（control）= 0.260 ±0.097,A_（25% PF）= 0.353 ± 0.049,A_（30% PF）= 0.212 ± 0.049;F = 20.821,P = 0.000）. Alizarin Red S and toluidine staining showed positive results. RT-q PCR shows that the relative expression of osteoblast and chondroid markers were increased compared to the control and 2D induction groups（F_（OPN）=65.576,P_（OPN）=0.000;F_（COL-2）=38.382,P_（COL- 2）= 0.000）. Conclusion Pluronic F-127 of 20%（w/v）could be a promising cell carrier material for bone and cartilage tissue engineering.

关 键 词：牙髓 干细胞 生物相容性 水凝胶 Pluronic F-127 组织工程 

分 类 号：R783.1[医药卫生—口腔医学]