Determining the Notch1 Expression in Chondrogenically Differentiated Rat Amniotic Fluid Stem Cells in Alginate Beads Using Conditioned Media from Chondrocytes Culture

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Abstract
The molecular mechanisms involved in the progression of chondrogenic differentiation require considerable studies, despite significant advancement in recent years. Chondrocytes are known for their paracrine effect yet there is little understanding about the impact of the paracrine factors released by chondrocytes in chondrogenic differentiation of mesenchymal stem cells. In this study, it was aimed to investigate the role of Notch signaling pathway during chondrogenesis of rat amniotic fluid stem cells (RAFSCs) in 3D culture in alginate beads using various induction media including the conditioned media from chondrocytes culture as well, for a period of 21 days. Our results indicated that Notch1 and VEGF genes were expressed during chondrogenic differentiation of RAFSCs along with SOX9 and COL2A1 upregulation in all different groups. However, SOX9 was significantly upregulated in group containing both conditioned media and standard chondrogenic induction medium (CM + IM group). The results suggest that conditioned medium can initiate chondrogenic differentiation, but it does not sufficiently contain all the factors potent for strong upregulation of SOX9. However, conditioned medium maintains the Notch1 gene expression at a sufficient level during the chondrogenesis stage. We propose that studies in this area are needed to understand the exact mechanisms involved in progression of differentiation under the influence of conditioned media.

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