INTRODUCTION An understanding of the extracellular matrix characteristics which stimulate and guide stem cell differentiation in the dental pulp is fundamental for the development of enhanced dental regenerative therapies. Our objectives, in this study, were to determine whether stem cells from the apical papilla (SCAP) responded to substrate stiffness, whether hydrogels providing micropatterned topographical cues stimulate SCAP self-alignment, and whether the resulting alignment could influence their differentiation towards an odontogenic lineage in-vitro. METHODS Experiments utilized gelatin methacryloyl (GelMA) hydrogels of increasing concentrations (5, 10 and 15%). We determined their compressive modulus via unconfined compression and analyzed cell spreading via F-actin/DAPI immunostaining. GelMA hydrogels were micropatterned using photolithography, in order to generate microgrooves and ridges of 60 and 120μm, onto which SCAP were seeded and analyzed for self-alignment via fluorescence microscopy. Lastly, we analyzed the odontogenic differentiation of SCAP using alkaline phosphatase protein expression (ANOVA/Tukey α=0.05). RESULTS SCAP appeared to proliferate better on stiffer hydrogels. Both 60 and 120μm micropatterned hydrogels guided the self-alignment of SCAP with no significant difference between them. Similarly, both 60 and 120μm micropattern aligned cells promoted higher odontogenic differentiation than non-patterned controls. SIGNIFICANCE In summary, both substrate mechanics and geometry have a statistically significant influence on SCAP response, and may assist in the odontogenic differentiation of dental stem cells. These results may point toward the fabrication of cell-guiding scaffolds for regenerative endodontics, and may provide cues regarding the development of the pulp-dentin interface during tooth formation.

中文翻译：

---

微图案化的水凝胶和细胞排列增强了体外培养的根尖乳头干细胞的牙源性潜力。

引言对刺激和引导牙髓中干细胞分化的细胞外基质特性的了解是开发增强的牙齿再生疗法的基础。在这项研究中，我们的目标是确定来自根尖乳头（SCAP）的干细胞是否对基质刚度作出反应，提供微图案形貌线索的水凝胶是否刺激SCAP自对准，以及所产生的对准是否会影响它们向牙源性谱系的分化体外。方法实验使用浓度增加（5％，10％和15％）的明胶甲基丙烯酰基（GelMA）水凝胶。我们通过无限制压缩确定了它们的压缩模​​量，并通过F-肌动蛋白/ DAPI免疫染色分析了细胞的扩散。使用光刻对GelMA水凝胶进行微图案化，以产生60和120μm的微沟槽和脊，在其上接种SCAP并通过荧光显微镜分析自对准。最后，我们使用碱性磷酸酶蛋白表达（ANOVA / Tukeyα= 0.05）分析了SCAP的牙源性分化。结果在较硬的水凝胶上，SCAP的增殖似乎更好。60和120μm的微图案水凝胶均能指导SCAP的自对准，两者之间无显着差异。同样，与未图案对照相比，60和120μm微米图案对齐的细胞均可促进更高的牙源性分化。意义总而言之，基材的力学性能和几何形状对SCAP反应均具有统计学上的显着影响，并且可能有助于牙齿干细胞的牙源性分化。