Bispo Daniela S C, Graça Inês C R, Jesus Catarina S H, Rodrigues João E, Correia Marlene C, Atella Sabrina, Duarte Iola F, Goodfellow Brian J, Oliveira Mariana B, Mano João F, Gil Ana M
Department of Chemistry, CICECO - Aveiro Institute of Materials, University of Aveiro, Campus Universitario de Santiago, Aveiro, 3810-193, Portugal.
Stem Cell Res Ther. 2025 Jun 7;16(1):294. doi: 10.1186/s13287-025-04419-x.
Mesenchymal stem cells (MSC) are pivotal bioengineering tools, offering significant promise for applications in bone regeneration. However, their therapeutic potential is limited by inter-donor variability and experimental issues. This study aimed to identify robust metabolic markers of osteodifferentiation applicable across multiple donors, while providing insight into the metabolic pathways actively involved in the process.
Untargeted nuclear magnetic resonance (NMR) metabolomics was applied to characterize the intra- and extracellular metabolic adaptations of human adipose-derived MSC (hAMSC) undergoing osteogenic differentiation, compared to proliferation alone. Multivariate and univariate statistical analysis was carried out on data from three independent donors, and cross-validation was employed to evaluate the predictive capacity of the proposed markers.
Variations in the levels of selected (nine) intracellular and (seventeen) extracellular metabolites detect osteodifferentiation by day 7 (out of 21), with nearly 100% accuracy. These signatures suggest a metabolic shift from glycolysis/OxPhos to lactic fermentation, fatty acid β-oxidation and phosphocreatine hydrolysis. Intracellular glucose, lactate, citrate and specific amino acids are redirected towards protein synthesis and glycosylation, with some of the secreted metabolites (e.g., citrate) seemingly involved in biomineralization and other extracellular roles. Membrane metabolism, antioxidant mechanisms and adenosine metabolism are also impacted by osteodifferentiation.
These findings reveal effective donor-independent markers of hAMSC osteodifferentiation, with a robust extracellular signature standing out for potential rapid and non-invasive detection of osteocommitted cells.
间充质干细胞（MSC）是关键的生物工程工具，在骨再生应用方面具有巨大潜力。然而，其治疗潜力受到供体间变异性和实验问题的限制。本研究旨在确定适用于多个供体的强大的成骨分化代谢标志物，同时深入了解该过程中积极参与的代谢途径。
应用非靶向核磁共振（NMR）代谢组学来表征人脂肪来源的MSC（hAMSC）在成骨分化过程中的细胞内和细胞外代谢适应性变化，并与单纯增殖情况进行比较。对来自三个独立供体的数据进行多变量和单变量统计分析，并采用交叉验证来评估所提出标志物的预测能力。
在第7天（共21天）时，通过检测选定的（9种）细胞内和（17种）细胞外代谢物水平的变化可检测到成骨分化，准确率近100%。这些特征表明代谢从糖酵解/氧化磷酸化转变为乳酸发酵、脂肪酸β-氧化和磷酸肌酸水解。细胞内葡萄糖、乳酸、柠檬酸和特定氨基酸被重新导向蛋白质合成和糖基化，一些分泌的代谢物（如柠檬酸）似乎参与生物矿化和其他细胞外作用。膜代谢、抗氧化机制和腺苷代谢也受到成骨分化的影响。
这些发现揭示了hAMSC成骨分化有效的不依赖供体的标志物，其中强大的细胞外特征在潜在快速、无创检测成骨细胞方面尤为突出。
