[Preparation and Evaluation of Clinical-Grade Human Umbilical Cord-Derived Mesenchymal Stem Cells with High Expression of Hematopoietic Supporting Factors].
Jie Tang, Pei-Lin Li, Xiao-Yu Zhang, Xiao-Tong Li, Fu-Hao Yu, Jia-Yi Tian, Run-Xiang Xu, Bo-Feng Yin, Li Ding, Heng Zhu
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引用次数: 0
Abstract
Objective: To prepare clinical-grade human umbilical cord-derived mesenchymal stem cells (hUC-MSC) with high expression of hematopoietic supporting factors and evaluate their stem cell characteristics.
Methods: Fetal umbilical cord tissues were collected from healthy postpartum women during full-term cesarean section. Wharton's jelly was mechanically separated and hUC-MSCs were obtained by explant culture method and enzyme digestion method in an animal serum-free culture system with addition of human platelet lysate. The phenotypic characteristics of hUC-MSCs obtained by two methods were detected by flow cytometry. The differences in proliferation ability between the two groups of hUC-MSCs were identified through CCK-8 assay and colony forming unit-fibroblast (CFU-F) assay. The differences in multilineage differentiation potential between the two groups of hUC-MSCs were identified through induction of adipogenic, osteogenic, and chondrogenic differentiation. The mRNA expression levels of hematopoietic supporting factors such as SCF, IL-3, CXCL12, VCAM1 and ANGPT1 in the two groups of hUC-MSCs were identified by real-time fluorescence quantiative PCR(RT-qPCR).
Results: The results of flow cytometry showed that hUC-MSCs obtained by the two methods both expressed high levels of CD73, CD90 and CD105, while lowly expressed CD31, CD45 and HLA-DR. The results of CCK-8 and CFU-F assay showed that the proliferation ability of hUC-MSCs obtained by explant culture method was better than those obtained by enzyme digestion method. The results of the triple lineage differentiation experiment showed that there was no significant difference in multilineage differentiation potential between the two grous of hUC-MSCs. The results of RT-qPCR showed that the mRNA expression levels of hematopoietic supporting factors SCF, IL-3, CXCL12, VCAM1 and ANGPT1 in hUC-MSCs obtained by explant cultrue method were higher than those obtained by enzyme digestion method.
Conclusion: Clinical-grade hUC-MSCs with high expression levels of hematopoietic supporting factors were successfully cultured in an animal serum-free culture system.
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