The Comparative Study Of Modulation On The Differentiation Of Rabbit Annulus Fibrosus-derived Stem Cells And Rabbit Bone Marrow Mesenchymal Stem Cells Using Scaffolds Of Various Elastic Moduli

Objective: To isolate and culture rabbit annulus fibrosus-derived stem cells(AFSCs) and bone marrow mesenchymal stem cells(BMSCs) and manufacture PECUU scaffolds with different moduli. To make comparisons of differentiation ability of AFSCs and BMSCs into AF-like cells of radial regions of native annulus fibrosus using electrospun scaffolds of various moduli.Methods: We harvested the annulus fibrosus and bone marrow of the same six-month-old New Zealand white rabbit and isolated and cultured AFSCs and BMSCs. PECUU scaffolds of various moduli were fabricated using electrospinning technology. We planted AFSCs and TGF-β3-mediated BMSCs(t BMSCs) into scaffolds. The proliferation ability of both two types of stem cells in the scaffolds with MTS assay and the differentiation ability into AF-like cells of radial regions of native annulus fibrosus through RT-q PCR and CTFM technology were explored.Results: We successfully isolated and cultured AFSCs and BMSCs which expressed stem cell markers OCT-4, NS and SSEA-4 and were capable of induced adipogenesis, osteogenesis and chondrogenesis. PECUU scaffolds of various moduli with high porosity were fabricated. AFSCs and t BMSCs on the substrate appeared to proliferate well during culture. The expression of collagen-I in both t BMSCs and AFSCs increased with the increase of scaffold modulus, whereas the expression of collagen-II and aggrecan gene expression showed an opposite trend. Interestingly, the expression of collagen-I, collagen-II and aggrecan genes in t BMSCs on PECUU scaffolds were consistently higher than those in AFSCs regardless of scaffold modulus. In addition, the cell traction forces(CTFs) of both t BMSCs and AFSCs gradually decreased with the increase of scaffold modulus, which is similar to the CTF change of cells from inner to outer regions of native AF tissue.Conclusion: Both two kinds of stem cells were modulated by substrate modulus in terms of gene expression and cell traction forces and had the similar differentiation ability into AF-like cells of radial regions of native rabbit annulus fibrosus.