Molecular dissection of somatic NF1 pseudarthrosis

Neurofibromatosis Type 1 (NF1) is a tumor predisposition syndrome with pleiotropic somatic manifestations, including formation of bone pseudarthrosis after fracture. The pathogenesis of NF1 pseudarthroses remains unclear, though defects in osteogenesis have been posited. Here, we applied time-series single-cell RNA-sequencing (scRNAseq) to patient-matched control and pseudarthrosis-derived primary bone mesenchymal stromal cells (MSCs). We show that osteogenesis occurs in NF1-/- MSCs. In contrast, expression of genetic pathways associated with skeletal mineralization were reduced in NF1-/- cells compared to co-cultured NF1+/- fracture-derived cells. Overall design: Control bone-derived primarry MSCs underwent time-series scRNAseq prior to differentiation and after 3-days and 9-days of osteogenesis. Patient-matched pseudarthrosis-derived primary MSCs underwent time-series scRNAseq prior to differentiation and following 3-, 6-, and 9-days of osteogenesis. All experiments were performed using the Fluidigm C1 instrument.