Sox9 modulates cell survival and adipogenic differentiation of multipotent adult rat mesenchymal stem cells.

Sox9 is a key transcription factor in early chondrogenesis with distinct roles in differentiation processes and during embryonic development. Here, we report that Sox9 modulates cell survival and contributes to the commitment of mesenchymal stem cells (MSC) to adipogenic or osteogenic differentiation lineages. We found that the Sox9 activity level affects the expression of the key transcription factor in adipogenic differentiation, C/EBPβ, and that cyclin D1 mediates the expression of the osteogenic marker osteocalcin in undifferentiated adult bone-marrow-derived rat MSC. Introducing a stable Sox9 knockdown into undifferentiated rat MSC resulted in a marked decrease in proliferation rate and an increase in apoptotic activity. This was linked to a profound upregulation of p21 and cyclin D1 gene and protein expression accompanied by an induction of caspase 3/7 activity and an inhibition of Bcl-2. We observed that Sox9 silencing provoked a delayed S-phase progression and an increased nuclear localization of p21. The protein stability of cyclin D1 was induced in the absence of Sox9 presumably as a function of altered p38 signalling. In addition, the major transcription factor for adipogenic differentiation, C/EBPβ, was repressed after silencing Sox9. The nearly complete absence of C/EBPβ protein as a result of increased destabilization of the C/EBPβ mRNA and the impact on osteocalcin gene expression and protein synthesis, suggests that a delicate balance of Sox9 level is not only imperative for proper chondrogenic differentiation of progenitor cells, but also affects the adipogenic and probably osteogenic differentiation pathways of MSC. Our results identified Sox9 as an important link between differentiation, proliferation and apoptosis in undifferentiated adult rat mesenchymal stem cells, emphasizing the importance of the delicate balance of a precisely regulated Sox9 activity in MSC not only for proper skeletal development during embryogenesis but probably also for successful repair and regeneration of tissues and organs in adults.