Department of Oral and Maxillofacial Surgery, College of Medicine, The Catholic University of Korea, Seoul 06591, Korea.
Department of Periodontics, College of Medicine, The Catholic University of Korea, Seoul 06591, Korea.
Medicina (Kaunas). 2021 Jun 8;57(6):586. doi: 10.3390/medicina57060586.
NELL-1 is a competent growth factor and it reported to target cells committed to the osteochondral lineage. The secreted, osteoinductive glycoproteins are reported to rheostatically control skeletal ossification. This study was performed to determine the effects of NELL-1 on spheroid morphology and cell viability and the promotion of osteogenic differentiation of stem cell spheroids. Cultures of stem cell spheroids of gingiva-derived stem cells were grown in the presence of NELL-1 at concentrations of 1, 10, 100, and 500 ng/mL. Evaluations of cell morphology were performed using a microscope, and cell viability was assessed using a two-color assay and Cell Counting Kit-8. Evaluation of the activity of alkaline phosphatase and calcium deposition assays involved anthraquinone dye assay to determine the level of osteogenic differentiation of cell spheroids treated with NELL-1. Real-time quantitative polymerase chain reaction (qPCR) was used to evaluate the expressions of , , , , and mRNAs. The applied stem cells produced well-formed spheroids, and the addition of NELL-1 at tested concentrations did not show any apparent changes in spheroid shape. There were no significant changes in diameter with addition of NELL-1 at 0, 1, 10, 100, and 500 ng/mL concentrations. The quantitative cell viability results derived on Days 1, 3, and 7 did not show significant disparities among groups ( > 0.05). There was statistically higher alkaline phosphatase activity in the 10 ng/mL group compared with the unloaded control on Day 7 ( < 0.05). A significant increase in anthraquinone dye staining was observed with the addition of NELL-1, and the highest value was noted at 10 ng/mL ( < 0.05). qPCR results demonstrated that the mRNA expression levels of RUNX2 and BSP were significantly increased when NELL-1 was added to the culture. Based on these findings, we conclude that NELL-1 can be applied for increased osteogenic differentiation of stem cell spheroids.
NELL-1 是一种有能力的生长因子，据报道，它可以靶向已经向成骨软骨谱系分化的细胞。分泌的、具有成骨诱导性的糖蛋白被报道可以对骨骼矿化进行静态控制。本研究旨在确定 NELL-1 对干细胞球体形态和细胞活力的影响，以及促进干细胞球体的成骨分化。将牙龈来源的干细胞的干细胞球体培养物在 1、10、100 和 500ng/ml 的浓度下添加 NELL-1 进行培养。使用显微镜评估细胞形态，使用双色测定法和细胞计数试剂盒-8 评估细胞活力。碱性磷酸酶活性和钙沉积测定的评估涉及蒽醌染料测定，以确定用 NELL-1 处理的细胞球体的成骨分化水平。实时定量聚合酶链反应 (qPCR) 用于评估 、 、 、 和 基因的表达。所应用的干细胞产生了形态良好的球体，并且在测试浓度下添加 NELL-1 并没有显示出球体形状的任何明显变化。在添加浓度为 0、1、10、100 和 500ng/ml 的 NELL-1 时，直径没有明显变化。第 1、3 和 7 天的定量细胞活力结果在组间没有显著差异（>0.05）。与未加载对照相比，第 7 天 10ng/ml 组的碱性磷酸酶活性显著升高（<0.05）。添加 NELL-1 后观察到蒽醌染料染色显著增加，在 10ng/ml 时达到最高值（<0.05）。qPCR 结果表明，添加 NELL-1 后，RUNX2 和 BSP 的 mRNA 表达水平显著增加。基于这些发现，我们得出结论，NELL-1 可用于增加干细胞球体的成骨分化。
