Quantitative Analysis of SSEA3+Cells from Human Umbilical Cord after Magnetic Sorting
Quantitative Analysis of SSEA3+Cells from Human Umbilical Cord after Magnetic Sorting
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磁选后对人脐带 SSEA3 细胞进行定量分析
DOI：
10.1177/0963689719844260
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发表时间：
2019-07-01
影响因子：
3.3
通讯作者：
Young, Wise
中科院分区：
文献类型：
--
作者：
Leng, Zikuan;Sun, Dongming;Young, Wise
多系分化的耐应激（Muse）细胞是一群多能阶段特异性胚胎抗原3（SSEA 3）+间充质干细胞，由Mari Dezawa在2010年首次描述。尽管一些研究者已经报道了脐带组织中的SSEA 3+间充质细胞，但是没有人定量地比较从沃顿氏胶（WJ）和人脐带（HUCs）的脐带衬里（CL）分离的SSEA 3+细胞。我们从HUCs中分离出WJ和CL，用胶原酶培养从这两种组织中分离的间充质基质细胞（MSC），并在三次传代中定量SSEA 3+细胞的百分比。第一代分别有5.0% +/- 4.3%和5.3% +/-5.1%SSEA 3+细胞来自WJ和CL，但CL组中P0和P2之间以及WJ组中P0和P1之间SSEA 3+细胞的百分比显著降低（P < 0.05）。磁激活细胞分选（MACS）显著富集SSEA 3+细胞至91.4% +/-3.2%。在培养分选的群体时，我们发现SSEA 3+百分比在P2-P5中的范围为62.5%至76.0%，然后在P6和P9之间下降至42.0%至54.7%。在P10时，培养物含有37.4%的SSEA 3+细胞。在P10后，我们重新分选细胞，并在培养物中获得89.4%的SSEA 3+细胞。基于MACS富集SSEA 3+细胞的程序，随后在培养物中扩增和再富集步骤，允许在相对纯的培养物中分离数百万个SSEA 3+细胞。当培养时，分选的SSEA 3+细胞分化成胚状体球，并在移植到挫伤的Sprague-Dawley大鼠脊髓中后存活4周。移植的SSEA 3+细胞从挫伤部位周围的四个注射点迁移到损伤区域，并且没有产生任何肿瘤。脐带是胎儿Muse细胞的极好来源，并且我们的方法允许实用且有效地分离和扩增相对纯的SSEA 3 + Muse细胞群体，所述相对纯的SSEA 3 + Muse细胞群体可以与人类白细胞抗原匹配以用于在人体试验中移植。
Multilineage-differentiating stress-enduring (Muse) cells are a population of pluripotent stage-specific embryonic antigen 3 (SSEA3)+ mesenchymal stem cells first described by Mari Dezawa in 2010. Although some investigators have reported SSEA3+ mesenchymal cells in umbilical cord tissues, none have quantitatively compared SSEA3+ cells isolated from Wharton's jelly (WJ) and the cord lining (CL) of human umbilical cords (HUCs). We separated WJ and the CL from HUCs, cultured mesenchymal stromal cells (MSCs) isolated from these two tissues with collagenase, and quantified the percentage of SSEA3+ cells over three passages. The first passage had 5.0% +/- 4.3% and 5.3% +/- 5.1% SSEA3+ cells from WJ and the CL, respectively, but the percentage of SSEA3+ cells decreased significantly (P < 0.05) between P0 and P2 in the CL group and between P0 and P1 in the WJ group. Magnetic-activated cell sorting (MACS) markedly enriched SSEA3+ cells to 91.4% +/- 3.2%. Upon culture of the sorted population, we found that the SSEA3+ percentage ranged from 62.5% to 76.0% in P2-P5 and then declined to 42.0%-54.7% between P6 and P9. At P10, the cultures contained 37.4% SSEA3+ cells. After P10, we resorted the cells and achieved 89.4% SSEA3+ cells in culture. The procedure for MACS-based enrichment of SSEA3+ cells, followed by expansion in culture and a re-enrichment step, allows the isolation of many millions of SSEA3+ cells in relatively pure culture. When cultured, the sorted SSEA3+ cells differentiated into embryoid spheres and survived 4 weeks after transplant into a contused Sprague-Dawley rat spinal cord. The transplanted SSEA3+ cells migrated into the injury area from four injection points around the contusion site and did not produce any tumors. The umbilical cord is an excellent source of fetal Muse cells, and our method allows the practical and efficient isolation and expansion of relatively pure populations of SSEA3+ Muse cells that can be matched by human leukocyte antigen for transplantation in human trials.