Lee Hyunjin, Son Jihwan, Min Sae Kyung, Na Chae-Bin, Yi Gawon, Koo Heebeom, Park Jun-Beom
Department of Periodontics, College of Medicine, The Catholic University of Korea, Seoul 06591, Korea.
Department of Medical Life Sciences and Department of Biomedicine & Health Sciences, College of Medicine, The Catholic University of Korea, Seoul 06591, Korea.
Materials (Basel). 2019 Aug 23;12(17):2693. doi: 10.3390/ma12172693.
The objective of the present investigation is to determine the effects of neutral, anionic, and cationic liposomes loaded with doxorubicin with thin-lipid-film-hydration method on the cellular viability and osteogenesis of stem cell spheroids. Spheroid formation and morphology of the three-dimensional spheroid were noted with an inverted microscope. Quantitative cellular viability was assessed using a commercially available kit. Osteogenic potential was evaluated by applying alkaline phosphatase activity and anthraquinone dye of Alizarin Red S. Western blot analysis was performed using collagen I expression. Spheroids were formed in each silicon elastomer-based concave microwell on Day 1. Noticeable changes of the spheroid were seen with a higher concentration of doxorubicin, especially in the cationic liposome group at Days 5 and 7. We found that the application of doxorubicin for 5 days significantly reduced the cellular viability. A higher concentration of doxorubicin produced a significant decrease in alkaline phosphatase activity. Alizarin Red S staining showed that extracellular calcium deposits were evenly noted in each group. An increase of calcium deposits was noted on Day 14 when compared to Day 7. The morphology of the groups with higher concentrations of doxorubicin showed to be more dispersed. We noticed that doxorubicin-loaded cationic liposomes resulted in the highest uptake of the examined cell spheroids and that doxorubicin-loaded liposomes affected the osteogenic differentiation. The implication of this study is that the type of liposome should be selected based on the purpose of the application.
本研究的目的是确定采用薄膜水化法制备的负载阿霉素的中性、阴离子和阳离子脂质体对干细胞球的细胞活力和成骨作用的影响。使用倒置显微镜观察三维球体的球体形成和形态。使用市售试剂盒评估细胞活力的定量。通过应用碱性磷酸酶活性和茜素红S的蒽醌染料来评估成骨潜力。使用胶原蛋白I表达进行蛋白质免疫印迹分析。第1天在每个基于硅弹性体的凹形微孔中形成球体。在较高浓度的阿霉素作用下，尤其是在第5天和第7天的阳离子脂质体组中，观察到球体有明显变化。我们发现阿霉素作用5天会显著降低细胞活力。较高浓度的阿霉素会使碱性磷酸酶活性显著降低。茜素红S染色显示，每组中细胞外钙沉积均匀可见。与第7天相比，第14天钙沉积增加。较高浓度阿霉素组的形态显示更分散。我们注意到，负载阿霉素的阳离子脂质体导致所检测的细胞球摄取量最高，并且负载阿霉素的脂质体影响成骨分化。本研究的意义在于应根据应用目的选择脂质体类型。
