a Medical Biotechnology and Nanotechnology Department , Zanjan University of Medical Sciences , Zanjan , Iran.
b Department of Pathology, Cancer Gene Therapy Research Center , Zanjan University of Medical Sciences , Zanjan , Iran.
Artif Cells Nanomed Biotechnol. 2018 Aug;46(5):1025-1033. doi: 10.1080/21691401.2017.1358733. Epub 2017 Aug 6.
Due to their renewal and potency, umbilical cord blood (UCB) stem cells have the ability to proliferate and serve as an attractive alternative source for bone marrow transplantation. However, insufficient number of haematopoietic stem cells (HSCs) in UCB is still a major constraint in clinical applications.
In vitro expansion of stem cells on fibronectin (Fn)-coated poly-L-lactic acid (PLLA) scaffold can be a proper way to overcome this limitation.
UCB CD133 + cells were isolated by magnetic cell sorting (MACS), and then the flowcytometry method was used for analysing CD133 + cells. Confirmed cells were seeded on the Fn-coated PLLA scaffold; also, collagen-coated PLLA scaffold, PLLA scaffold and two-dimensional (2D) culture system were expanded for 7 days. During this time, we used the flowcytometric method for analysing CD133 + cells and real-time PCR for the expression level of CXCR4 gene. The number of total cells and CD133 + cells, as well as MTT assay and colony-forming unit (CFU) assay were evaluated.
Flowcytometry data indicated that the purity of CD133 + before expansion was 93%. After 7 days, there was higher number of CD133 + cells on the Fn-coated PLLA scaffold compared to other groups. Moreover, results of MTT and colony assays showed higher viability and proliferation of CD133 + on the Fn-coated PLLA scaffold. Also, the quantity of CXCR4 gene expression increased compared to other groups.
The Fn-coated scaffold was the most effective scaffold for proliferation and improved the adhesiveness to the scaffold.
The Fn-coated PLLA scaffold could be a suitable in vitro mimicry niche over a 2D system.
由于脐带血（UCB）干细胞具有更新和活力，它们能够增殖，并成为骨髓移植的有吸引力的替代来源。然而，UCB 中的造血干细胞（HSCs）数量不足仍然是临床应用的主要限制。
在纤维连接蛋白（Fn）涂层的聚-L-乳酸（PLLA）支架上体外扩增干细胞可以是克服这一限制的适当方法。
通过磁细胞分选（MACS）分离 UCB CD133+细胞，然后使用流式细胞术分析 CD133+细胞。确认的细胞接种在 Fn 涂层 PLLA 支架上；此外，胶原蛋白涂层 PLLA 支架、PLLA 支架和二维（2D）培养系统也进行了 7 天的扩增。在此期间，我们使用流式细胞术分析 CD133+细胞，实时 PCR 分析 CXCR4 基因的表达水平。评估总细胞和 CD133+细胞的数量、MTT 测定和集落形成单位（CFU）测定。
流式细胞术数据表明，扩增前 CD133+的纯度为 93%。7 天后，Fn 涂层 PLLA 支架上的 CD133+细胞数量明显高于其他组。此外，MTT 和集落测定结果表明，Fn 涂层 PLLA 支架上 CD133+的活力和增殖更高。此外，与其他组相比，CXCR4 基因表达量增加。
Fn 涂层支架是增殖最有效的支架，可提高对支架的粘附性。
Fn 涂层 PLLA 支架可以作为 2D 系统的体外模拟龛。
