Although mesenchymal stromal cells (MSCs) have demonstrated great therapeutic potential, the heterogeneity of MSCs may be responsible for the incongruent data obtained in MSC-based preclinical studies and clinical trials. Here, four mouse clonal MSC lines, termed MSC1, MSC2, MSC3, and MSC4, were isolated and extensively characterized. MSC4 cells grew most rapidly and formed colonies of the largest size, whereas MSC3 cells exhibited the slowest growth and formed only a few tiny clusters. MSC4 cells could differentiate into adipocytes, osteoblasts, and chondrocytes in vitro, and more importantly, establish hematopoietic microenvironment in vivo; whereas the other lines displayed uni-adipogenic, osteo-chondrogenic, or non-differentiation potential. All lines were positive for Sca-1, CD106, and CD44; MSC4 was also positive for CD90.2. In terms of immunosuppressive capacity, MSC2, MSC3, and MSC4 cells exerted clear inhibitory effects on lymphocyte proliferation, whereas MSC1 did not. Further investigation revealed that the NO and not the PGE2 pathway may play a role in the different immunomodulatory effects of the cell lines. To clarify the molecular basis of this heterogeneity, we employed RNA sequencing to compare the gene expression profiles of the four subtypes, revealing a relationship between gene expression and variability in subtype function. This study provides novel information about the heterogeneity of MSCs and insight into the selection of optimal cell sources for therapeutic applications.
Keywords: Biological properties;Immunosuppression; CFSE; CFU-F; FCS; Gene expression; IDO; MSCs; Mesenchymal stromal cells; NO; PGE2; TGF-β3; carboxyfluorescein diacetate succinimidyl ester; colony forming unit-fibroblast; fetal calf serum; indoleamine 2,3 dioxygenase; mesenchymal stromal cells; nitric oxide; prostaglandin E2; transforming growth factor-β3.
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