Transcriptional plasticity of stromal cells amplifies their differentiation efficiency in vitro

Human bone marrow-derived stromal cells (also termed mesenchymal stem cell -MSCs) are progenitors capable of differentiating into bone forming osteoblasts and fat storing adipocytes. Due to the loss of bone mass being associated with increased marrow fat, trans-differentiation of osteoblasts into adipocytes been hypothesized as a contributor to osteoporotic bone loss and fragility. Reprogramming of transcriptional networks is a prerequisite for cellular differentiation, however, to which extent cell-type specific transcriptional networks modulate cellular plasticity within stromal cells remains unknown. In this study we performed gene expression analysis at bulk and single cell level in stromal cells being repeatedly exposed to osteogenic and adipogenic inducers in vitro. Surprisingly, cell type specific gene networks are not suppressive but instead promoting to obtain an opposing phenotype, e.g., enhanced osteoblast differentiation of adipogenic pre-stimulated stromal cells compared to undifferentiated ones. Mechanistically, lineage-selective genes with enhanced response upon interconversion are primed in the stem cell state and obtain modest activity levels during exposure to opposing lineage conditions. Finally, the presence of cells simultaneously showing an osteogenic and adipogenic phenotype highlighting a strong molecular plasticity of transcriptional networks in stromal cells. These observations provide a strong molecular support for the notion of not only progenitor specification but also the plasticity of differentiated cells contributing to the balance of bone mass and marrow fat content. SummaryWe provide mechanistic evidence, that cells with established lineage-selective gene programs are more vulnerable to obtain gene signatures and phenotypes of opposing lineages, building ground for trans-differentiation between osteoblasts and adipocytes being a coordinated process rather than random and misdirected.