Abstract
Background: 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., 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.
Highlights
The primary causes of disability worldwide are low back and neck pain [1]
All growth factors except single factor supplementation of Sonic hedgehog (Shh) and Wnt5a resulted in enhanced proteoglycan production (Figure 1)
We aimed to identify growth factors or factor combinations that showed the highest potency in enhancing mesenchymal stromal cells (MSC) to nucleus pulposus (NP) cell-like cell differentiation
Summary
The primary causes of disability worldwide are low back and neck pain [1] Despite their prevalence and socioeconomic impact, long-term curative treatments for these disorders are lacking. In reaction to these degenerative changes, the native IVD cells secrete inflammatory factors, potentially stimulating nerve ingrowth, vascularization, and immunogenic cell influx in the normally avascular and un-innervated discs or sensitizing nearby nerve structures [8,9,10]. These changes can trigger nociception and in severe cases, might result in debilitating pain and disability in patients [11,12]. We screened a wide range of GF and GF combinations for supporting NP cell-like cell differentiation
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