Optimization of seeding density of OP9 cells to improve hematopoietic differentiation efficiency.

IF 4.6 Q2 MATERIALS SCIENCE, BIOMATERIALS
Xin-Xing Jiang, Meng-Yi Song, Qi Li, Yun-Jian Wei, Yuan-Hua Huang, Yan-Lin Ma
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Abstract

Background: OP9 mouse stromal cell line has been widely used to induce differentiation of human embryonic stem cells (hESCs) into hematopoietic stem/progenitor cells (HSPCs). However, the whole co-culture procedure usually needs 14-18 days, including preparing OP9 cells at least 4 days. Therefore, the inefficient differentiation system is not appreciated. We aimed to optimize the culture conditions to improve differentiation efficiency.

Methods: In the experimental group, we set six different densities of OP9 cells and just cultured them for 24 h before co-culture, and in the control group, OP9 cells were cultured for 4 days to reach an overgrown state before co-culture. Then we compared the hematopoietic differentiation efficiency among them.

Results: OP9 cells were randomly assigned into two groups. In the experimental group, six different plated numbers of OP9 cells were cultured for 1 day before co-culture with hESCs. In contrast, in the control group, OP9 cells were cultured for 4 days at a total number of 3.1 × 104 cells/cm2 in a 6-well plate to reach an overgrown state before co-culture. Hematopoietic differentiation was evaluated with CD34 immunostaining, and compared between these two groups. We could not influence the differentiation efficiency of OP9 cells with a total number of 10.4 × 104 cells/cm2 in a 6-well plate which was cultured just for 1 day, followed by co-culture with hESCs. It reached the same differentiation efficiency 5 days earlier than the control group.

Conclusion: The peak of CD34 + cells appeared 2 days earlier compared to the control group. A total number of 1.0 × 106 cells in a 6-well plate for OP9 cells was appropriate to have high differentiation efficiency.

优化 OP9 细胞的播种密度,提高造血分化效率。
背景:OP9 小鼠基质细胞系已被广泛用于诱导人类胚胎干细胞(hESCs)分化为造血干细胞/祖细胞(HSPCs)。然而,整个共培养过程通常需要 14-18 天,其中包括至少 4 天的 OP9 细胞准备时间。因此,这种低效的分化系统并未得到重视。我们旨在优化培养条件,提高分化效率:在实验组中,我们设置了六种不同密度的 OP9 细胞,在共培养前仅培养 24 小时;在对照组中,OP9 细胞培养 4 天达到过度生长状态后再进行共培养。然后比较它们的造血分化效率:将 OP9 细胞随机分为两组。结果:OP9 细胞被随机分为两组,实验组中,6 个不同数量的 OP9 细胞经 1 天培养后与 hESCs 共同培养。而在对照组中,OP9 细胞以 3.1 × 104 cells/cm2 的总数量在 6 孔板中培养 4 天,达到过度生长状态后再与 hESCs 共同培养。用 CD34 免疫染色法评估造血分化情况,并对两组细胞进行比较。我们无法影响 OP9 细胞的分化效率,其总数量为 10.4 × 104 cells/cm2,在 6 孔板中仅培养 1 天,然后与 hESCs 共培养。结论:与对照组相比,CD34 + 细胞的分化效率提前了 5 天:结论:与对照组相比,CD34 + 细胞的高峰期提前了 2 天。OP9 细胞在 6 孔板中的总数量为 1.0 × 106 个细胞时,分化效率较高。
本文章由计算机程序翻译,如有差异,请以英文原文为准。
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来源期刊
ACS Applied Bio Materials
ACS Applied Bio Materials Chemistry-Chemistry (all)
CiteScore
9.40
自引率
2.10%
发文量
464
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