Single-Cell RNAseq of Out-of-Thaw Mesenchymal Stromal Cells Shows Striking Tissue-of-Origin Differences and Inter-donor Cell-Cycle Variations

Medrano-Trochez, Camila; Chatterjee, Paramita; Pradhan, Pallab; Ogle, Molly E; Botchwey, Edward A; Kurtzberg, Joanne; Yeago, Carolyn; Gibson, Greg; Roy, Krishnendu

Mesenchymal stromal cells (MSCs) from a variety of tissue sources are widely investigated in clinical trials, and the MSCs are often administered immediately after thawing the cryopreserved product. While previous reports have examined the transcriptome of freshly-cultured MSCs from some tissues, little is known about the single-cell transcriptomic profiles of out-of-thaw MSCs from different tissue sources. Such understanding could help determine which tissue origins and delivery methods are best suited for specific indications. Here, we characterized cryopreserved MSCs, immediately post-thaw, from bone marrow (BM) and cord tissue (CT), using single-cell RNA sequencing (scRNA-seq). We show that out-of-thaw BM-vs. CT-MSCs have significant differences in gene expression. Gene-set enrichment analyses implied divergent functional potential. In addition, we show that MSC-batches can vary significantly in cell cycle status, suggesting different proliferative vs. immunomodulatory potentials. Our results provide a comprehensive single-cell transcriptomic landscape of clinically and industrially relevant MSC products. Highlights Single cell gene expression comparison between Bone-marrow derived MSCs and Cord-tissue derived MSCs Donor effects and cell heterogeneity on tissue-specific MSC gene expression Single Cell Pooling Enhances Differential Expression Analysis for Bone marrow and Cord tissue MSC samples Gene ontology reveals tissue specific unique molecular function and pathways