Screening for Growth-Factor Combinations Enabling Synergistic Differentiation of Human MSC to Nucleus Pulposus Cell-Like Cells

Kosuke Morita, Jordy Schol, Tibo N. E. Volleman, Daisuke Sakai, Masato Sato, Masahiko Watanabe
2021 Applied Sciences  
Multiple studies have examined the potential of growth factors (GF) to enable mesenchymal stromal cells (MSC) to nucleus pulposus (NP) cell-like cell differentiation. Here we screened a wide range of GF and GF combinations for supporting NP cell-like cell differentiation. Methods: Human MSC were stimulated using 86 different GF combinations of TGF-β1, -2, -3, GDF5, -6, Wnt3a, -5a, -11, and Shh. Differentiation potency was assessed by alcian blue assay and NP cell marker expression (e.g.,
more » ... ssion (e.g., COL2A1, CD24, etc.). The top four combinations and GDF5/TGF-β1 were further analyzed in 3D pellet cultures, on their ability to similarly induce NP cell differentiation. Results: Almost all 86 GF combinations showed their ability to enhance proteoglycan production in alcian blue assay. Subsequent qPCR analysis revealed TGF-β2/Wnt3a, TGF-β1/Wnt3a, TGF-β1/Wnt3a/GDF6, and Wnt3a/GDF6 as the most potent combinations. Although in pellet cultures, these combinations supported NP marker expression, none showed the ability to significantly induce chondrogenic NP matrix production. Only GDF5/TGF-β1 resulted in chondrogenic pellets with significantly enhanced glycosaminoglycan content. Conclusion: GDF5/TGF-β1 was suggested as an optimal GF combination for MSC to NP cell induction, although further assessment using a 3D and in vivo environment is required. Wnt3a proved promising for monolayer-based NP cell differentiation, although further validation is required.
doi:10.3390/app11083673 fatcat:mfeyk6ow2rbgpdldvesmwgb7um