Abstract
Despite the wide use of mesenchymal stromal cells (MSCs) for paracrine support in clinical trials, their variable and heterogeneous supporting activity pose major challenges. While three-dimensional (3D) MSC cultures are emerging as alternative approaches, key changes in cellular characteristics during 3D-spheroid formation remain unclear. Here, we show that MSCs in 3D spheroids undergo further progression towards the epithelial-mesenchymal transition (EMT), driven by upregulation of EMT-promoting microRNAs and suppression of EMT-inhibitory miRNAs. The shift of EMT in MSCs is associated with widespread histone modifications mimicking the epigenetic reprogramming towards enhanced chromatin dynamics and stem cell-like properties, but without changes in their surface phenotype. Notably, these molecular shifts towards EMT in 3D MSCs caused enhanced stem cell niche activity, resulting in higher stimulation of hematopoietic progenitor self-renewal and cancer stem cell metastasis. Moreover, miRNA-mediated induction of EMT in 2D MSCs were sufficient to mimic the enhanced niche activity of 3D spheroid MSCs. Thus, the molecular hierarchy in the EMT gradient among phenotypically indistinguishable MSCs revealed the previously unrecognized functional parameters in MSCs, and the EMT-enhanced “naïve” mesenchymal state represents an ‘activated mesenchymal niche’ in 3D spheroid MSCs.
Highlights
Mesenchymal stromal cells (MSCs) are nonhematopoietic adherent cell populations derived from various organs, including the bone marrow (BM), adipose tissue, and placental tissue
To achieve 3D spheroid formation, mesenchymal stromal cells (MSCs) were cultured on PDMS microchips, a platform for 3D culture by ultra-low attachment[19], and compared with those grown by 2D adherent culture in plastic dishes
MSCs within the spheroids were smaller in size than MSCs in 2D culture (Supplementary Fig. S1a,b) and became more quiescent in cell cycling, as determined by Ki67 staining for detection of proliferation (Supplementary Fig. S1c,d)
Summary
Mesenchymal stromal cells (MSCs) are nonhematopoietic adherent cell populations derived from various organs, including the bone marrow (BM), adipose tissue, and placental tissue. MSCs derived from pericytes in various organs[4] can give rise to self-renewing mesenchymal progenitors, which contribute to stem cell niche in a heterotrophic BM model[5]. These MSCs are frequently expanded by in-vitro culture and these ex vivo cultured MSCs have been widely used for cell therapy in a variety of clinical trials. Our recent study showed that the cellular characteristics of MSCs to support stem cells exhibit dynamic changes according to the pathological changes in bone marrow microenvironment or intensity of the canonical Wnt signaling pathways[10, 11] and can exhibit reversible changes in niche activity by switching of culture conditions[12], pointing possible heterogeneity in the supporting function of MSCs with respect to the culture conditions
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