Department of Periodontics, College of Medicine, The Catholic University of Korea, Seoul 06591, Republic of Korea.
Department of Medicine, Graduate School, The Catholic University of Korea, Seoul 06591, Republic of Korea.
Biomed Res Int. 2022 Jul 14;2022:9818299. doi: 10.1155/2022/9818299. eCollection 2022.
This study was aimed at evaluating the effects of transforming growth factor- on the differentiation and mRNA expression of organoids made out of human mesenchymal stem cells. Cell organoids composed of gingiva-derived stem cells were cultured in the presence of transforming growth factor-1 at concentrations ranging from 0, 1, 10, to 20 ng/ml. Evaluations of the cell morphology of the organoids were performed on days 7, 9, 11, and 14. Quantitative cellular viability was completed on day 14. Alkaline phosphatase activity assays were performed to evaluate the differentiation of stem cells on day 14. Real-time polymerase chain reactions were used to determine the expression levels of TGF-1, RUNX2, OCN, SOX9, and COL1A1 mRNA on day 14. The stem cells produced well-formed organoids on day 7, and the addition of transforming growth factor-1 did not result in relevant changes in their shape. The organoids grew in size and became more intact with longer incubation times. On day 14, the diameters were 222.2 ± 9.6, 186.1 ± 4.8, 197.2 ± 9.6, and 211.1 ± 19.2 m for transforming growth factor-1 at final concentrations of 0, 1, 10, and 20 ng/ml, respectively. Quantitative cell viability results from day 14 exhibited no significant difference between the groups ( > 0.05). There was significantly higher alkaline phosphatase activity with the addition of transforming growth factor-1 with the highest value for the 1 ng/ml group ( < 0.05). Real-time polymerase chain reaction results demonstrated that the mRNA expression levels of RUNX2, OCN, and SOX were higher in 1 ng/ml but did not reach statistical significance. Treatment with 1 ng/ml of transforming growth factor-1 significantly increased COL1A1 mRNA expression at day 14. The application of transforming growth factor-1 increased differentiation, which was confirmed by alkaline phosphatase activity and mRNA expression while maintaining cell viability.
本研究旨在评估转化生长因子-β对人间质干细胞来源的类器官分化和 mRNA 表达的影响。将由牙龈干细胞组成的细胞类器官在浓度范围为 0、1、10 和 20ng/ml 的转化生长因子-β存在下培养。在第 7、9、11 和 14 天评估类器官的细胞形态。在第 14 天完成定量细胞活力测定。在第 14 天进行碱性磷酸酶活性测定以评估干细胞的分化。使用实时聚合酶链反应确定第 14 天 TGF-β1、RUNX2、OCN、SOX9 和 COL1A1 mRNA 的表达水平。第 7 天,干细胞产生了形态良好的类器官,添加转化生长因子-β不会导致其形态发生相关变化。类器官随孵育时间的延长而增大且变得更加完整。第 14 天,终浓度为 0、1、10 和 20ng/ml 的转化生长因子-β的类器官直径分别为 222.2±9.6、186.1±4.8、197.2±9.6 和 211.1±19.2μm。第 14 天的定量细胞活力结果显示各组间无显著差异(>0.05)。加入转化生长因子-β后碱性磷酸酶活性显著升高,以 1ng/ml 组最高(<0.05)。实时聚合酶链反应结果表明,RUNX2、OCN 和 SOX 的 mRNA 表达水平在 1ng/ml 时较高,但未达到统计学意义。第 14 天,1ng/ml 转化生长因子-β处理显著增加 COL1A1 mRNA 表达。转化生长因子-β的应用增加了分化,这通过碱性磷酸酶活性和 mRNA 表达得到证实,同时保持细胞活力。