Impact of Culture Platform Transition on Adipose-derived Mesenchymal Stromal Cells Molecular Signatures: A Multi-Omics Analysis of Small-Scale versus Large-Scale Production

Mesenchymal stromal cells (MSCs) are being tested in numerous clinical trials, yet the limited progression of these trials to advanced stages indicates unresolved translational challenges. Expanding MSCs is a critical step in most therapeutic applications and bioreactor-based culture offers large-scale production compared to monolayer cultures. Nevertheless, since MSCs sense their microenvironment, it is crucial to understand how these systems affect their properties. We expanded human adipose-derived mesenchymal stromal cells (AD-MSCs) from the same donors in both small- and large-scale platforms. Bulk-RNA sequencing and mass-spectrometry analysis demonstrate that small-scale culture had a broader range of differentially expressed genes and proteins within immunomodulatory, cell migration and cell adhesion pathways. In contrast, the large-scale culture showed a lower amount of differentially expressed genes - and proteins were proteinsparticularly associated mainly with extracellular matrix synthesis. Our findings demonstrate that expansion platforms significantly impact MSCs’ properties of MSCs, highlighting the need with optimise expansion conditions to obtain high cell yield without compromising MSCs’ their attributes.