Departamento de Clínicas Veterinárias, Instituto de Ciências Biomédicas de Abel Salazar (ICBAS), Universidade do Porto (UP), Rua de Jorge Viterbo Ferreira, n° 228, Porto, Portugal.
Centro de Estudos de Ciência Animal (CECA), Instituto de Ciências, Tecnologias e Agroambiente da Universidade do Porto (ICETA), Rua D. Manuel II, Apartado 55142, Porto, Portugal.
PLoS One. 2018 Oct 10;13(10):e0203936. doi: 10.1371/journal.pone.0203936. eCollection 2018.
Mesenchymal Stromal cells (MSCs) have a potential role in cell-based therapies. Foetal bovine serum (FBS) is used to supplement the basal cell culture medium but presents several disadvantages and risks. Other alternatives have been studied, including human umbilical cord blood plasma (hUCBP), aiming at the development of xeno-free culturing protocols. A comparative characterization of multicomponent metabolic composition of hUCBP and commercial FBS based on Nuclear Magnetic Resonance (NMR) spectroscopy and multivariate statistical analysis was performed. The analysis of 1H-NMR spectra revealed both similarities and differences between the two proposed supplements. Similar metabolites (amino acids, glucose, lipids and nucleotides) were found in the hUCBP and FBS NMR spectra. The results show that the major difference between the metabolic profiles of the two proposed supplements are due to the significantly higher levels of glucose and lower levels of lactate, glutamate, alanine and branched chain amino acids in hUCBP. Similar or slightly different levels of important proteinogenic amino acids, as well as of nucleotides, lipids were found in the hUCBP and FBS. In order to validate it's suitability for cell culture, umbilical cord-MSCs (UC-MSCs) and dental pulp stem cells (DPSCs) were expanded using hUCBP. In both hMSCs, in vitro culture with hUCBP supplementation presented similar to improved metabolic performances when compared to FBS. The two cell types tested expressed different optimum hUCBP percentage content. For DPSCs, the optimum hUCBP content was 6% and for UC-MSCs, 4%. Cultured hMSCs displayed no changes in senescence indicators, as well as maintained characteristic surface marker's expression. FBS substitution was associated with an increase in early apoptosis events, in a dose dependent manner, as well as to slight up- and down-regulation of targeted gene's expression. Tri-lineage differentiation capacity was also influenced by the substitution of FBS by hUCBP.
间充质基质细胞（MSCs）在基于细胞的治疗中具有潜在作用。胎牛血清（FBS）被用于补充基础细胞培养基，但存在多种缺点和风险。已经研究了其他替代物，包括人脐带血血浆（hUCBP），旨在开发无动物成分的培养方案。本研究通过核磁共振（NMR）光谱和多变量统计分析，对 hUCBP 和商业 FBS 的多成分代谢组成进行了比较分析。1H-NMR 谱分析揭示了两种拟议补充剂之间的相似性和差异。在 hUCBP 和 FBS NMR 谱中发现了相似的代谢物（氨基酸、葡萄糖、脂质和核苷酸）。结果表明，两种拟议补充剂代谢谱之间的主要差异是由于 hUCBP 中葡萄糖水平显著较高，而乳酸盐、谷氨酸盐、丙氨酸和支链氨基酸水平较低。在 hUCBP 和 FBS 中发现了相似或略有不同水平的重要蛋白质氨基酸以及核苷酸和脂质。为了验证其用于细胞培养的适用性，使用 hUCBP 扩增脐带-MSCs（UC-MSCs）和牙髓干细胞（DPSCs）。在两种 hMSCs 中，与 FBS 相比，使用 hUCBP 补充物进行体外培养具有相似或改善的代谢性能。两种测试的细胞类型表达了不同的最佳 hUCBP 含量。对于 DPSCs，最佳 hUCBP 含量为 6%，而对于 UC-MSCs，最佳 hUCBP 含量为 4%。培养的 hMSCs 衰老标志物的表达没有变化，并且保持了特征性表面标志物的表达。FBS 替代物与早期凋亡事件的增加有关，呈剂量依赖性，并且靶向基因的表达也略有上调和下调。FBS 替代物还影响三系分化能力。
