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Abstract
Mesenchymal stem/stromal cells (MSCs) have acquired considerable attention in regenerative medicine and cell-based therapies due to their remarkable regenerative and immunomodulatory properties. Understanding the molecular mechanisms underpinning MSC function is essential for harnessing their therapeutic potential effectively. This study uses the multifaceted approach to comprehensively characterize the MSC functional activity. First, we employed a high-resolution mass spec technique to interpret the alteration of lipids during MSC aging and established connections between alterations in cellular lipid composition and overall functionality to determine the number of passages MSCs can go for biomanufacturing processes. Next, we employed label-free imaging techniques to establish a direct correlation between single-cell MSC lipidomic profiles and their morphological characteristics to uncover novel insights into the heterogeneity within MSC populations and their implications for cell therapy outcomes. Finally, we explored the high-content imaging technique to perform live imaging of MSC donors seeded in a microfluidic device that can closely mimic the in vivo culture conditions. Our finding helped in assessing the high-potency donor based on migration and locomotion behavior. Overall, this study provides a comprehensive understanding of morphological features, lipid metabolic networks, and functional behavior of MSCs and offers insights into optimizing MSCs efficacy to enhance clinical therapeutics.