Department of Endodontics, Faculty of Dentistry, Hacettepe University, Ankara, Turkey.
Bioengineering Division, Institute for Graduate Studies in Science and Engineering, Hacettepe University, Ankara, Turkey.
Int Endod J. 2018 Apr;51(4):420-430. doi: 10.1111/iej.12869. Epub 2017 Nov 14.
To investigate the proliferation and differentiation potential of human dental pulp stem cells (DPSCs) in a three-dimensional culture model (TDM) by incorporation of VEGF and BMP-2.
TDM was established using fibrin gel (fg) as a soft tissue matrix and demineralized dentine disc (dd) as a hard tissue matrix. DPSCs and vascular endothelial growth factor (VEGF) were encapsulated in fibrin gel (fg-VEGF) and then inserted into bone morphogenetic protein (BMP-2)-coated demineralized dentine discs (dd-BMP-2). DPSCs were incubated for 28 days in various fg/dd combinations in the absence or presence of VEGF and BMP-2. Proliferation and morphology of DPSCs in fibrin gel were analysed using MTT and Live&Dead assays. Release profiles of VEGF and BMP-2 from fibrin gel and dentine discs were quantified using ELISA, and the expressions of angiogenic and odontogenic differentiation markers were determined with RT-qPCR analysis. Data were analysed statistically using Wilcoxon signed rank tests, Kruskal-Wallis tests with Mann-Whitney U tests and Bonferroni adjustment. The level of significance was set at P < 0.05.
DPSCs were able to proliferate and showed interconnected cellular elongations in fibrin gel depending on fibrinogen concentration whilst monolayer control group showed typical fibroblast-like cell morphology. Encapsulating of VEGF in fibrin gel and BMP-2 in gelatin that was used to coat dentine discs allowed the controlled releases of growth factors, which induced angiogenic and odontogenic gene expressions by DPSCs. Higher expressions of PECAM as an angiogenic factor, and BSP, DMP-1, OCN and CBFA as odontogenic factors, were observed in TDM as compared to the other fg/dd combinations and the monolayer control group (P < 0.05).
TDM consisting of fibrin gel and dentine matrix allowed cell-cell interactions. TDM was highly effective in delivering both VEGF and BMP-2 that enhanced the angiogenic and odontogenic potential of DPSCs.
通过将血管内皮生长因子（VEGF）和骨形态发生蛋白-2（BMP-2）纳入三维培养模型（TDM）中，研究人牙髓干细胞（DPSCs）的增殖和分化潜能。
使用纤维蛋白凝胶（fg）作为软组织基质和脱矿牙本质盘（dd）作为硬组织基质建立 TDM。将 DPSCs 和血管内皮生长因子（VEGF）包封在纤维蛋白凝胶（fg-VEGF）中，然后插入到骨形态发生蛋白-2（BMP-2）包被的脱矿牙本质盘中（dd-BMP-2）。在没有或存在 VEGF 和 BMP-2 的情况下，将 DPSCs 在各种 fg/dd 组合中孵育 28 天。使用 MTT 和 Live&Dead 测定法分析纤维蛋白凝胶中 DPSCs 的增殖和形态。使用 ELISA 定量测定纤维蛋白凝胶和牙本质盘中 VEGF 和 BMP-2 的释放曲线，并通过 RT-qPCR 分析确定血管生成和牙源性分化标志物的表达。使用 Wilcoxon 符号秩检验、Kruskal-Wallis 检验和 Mann-Whitney U 检验以及 Bonferroni 调整进行统计数据分析。显著性水平设定为 P < 0.05。
DPSCs 能够增殖，并根据纤维蛋白原浓度在纤维蛋白凝胶中显示出相互连接的细胞伸长，而单层对照组显示出典型的成纤维细胞样细胞形态。将 VEGF 包封在纤维蛋白凝胶中，将 BMP-2 包封在用于涂覆牙本质盘的明胶中，允许生长因子的受控释放，这诱导了 DPSCs 的血管生成和牙源性基因表达。与其他 fg/dd 组合和单层对照组相比，TDM 中观察到更高的血管生成因子 PECAM 以及牙源性因子 BSP、DMP-1、OCN 和 CBFA 的表达（P < 0.05）。
由纤维蛋白凝胶和牙本质基质组成的 TDM 允许细胞-细胞相互作用。TDM 非常有效地输送 VEGF 和 BMP-2，增强了 DPSCs 的血管生成和牙源性潜能。
