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Scaffold topography and culture medium conditions for human wharton's jelly mesenchymal stem cells (hWJ-MSC) are critical components of the approach to nucleus pulposus (NP) tissue engineering. Aim: To evaluate the silk fibroin (SF) scaffold topography analysis (optimal thickness and pore diameter) and to determine culture medium conditions for the growth and differentiation of hWJ-MSC. Method: hWJ-MSCs were seeded into different thicknesses and pore size diameters and grown in different concentrations of glucose, platelet rich plasma (PRP) and oxygen. The cell-seeded scaffold was evaluated for cell attachment, growth and differentiation potency. Results & discussion: The results indicated that SF scaffold with a minimum thickness 3.5 mm and pore diameter of 500 μm with cells cultured under low glucose, 10% PRP and normoxia conditions induced the growth and differentiation of hWJ-MSCs, indicated by the accumulation of glycosaminoglycans content and the presence of type II collagen, as markers of NP-like cells.