Repairing osteochondral defects (OCD) remains a formidable challenge due to the high complexity of native osteochondral tissue and the limited self-repair capability of cartilage. In this regard, the development of osteochondral tissue engineering with scaffolds seeded with stem cells along with mechanical stimulation has been considered by the researchers as a new proposed technique for the repair of this tissue. In this study, at first we fabricated an integrated and biomimetic trilayered Silk Fibroin (SF) scaffold containing SF nano fibers in each layer. Then fluid wall shear stress in different areas of the scaffold was predicted in dynamic cell culture condition under the inlet velocity of 0.4 ml/min in a perfusion bioreactor using finite elements and fluid-structure interactions methods. Finally, using the simulation results, osteogenesis and chondrogenesis of rabbit adipose derived stem cells (RADSCs) were analyzed. The results showed that this novel osteochondral graft has a seamlessly integrated layer structure and a high degree of pore interconnectivity. The average size of the pores in the bone layer, middle layer, and cartilage were 76, 152, and 102 microns, respectively. In addition, this biomimetic scaffold presented compressive moduli of 0.4 MPa and uitimate tensile strength of 10 MPa in the wet state. Also, based on the simulation analyses, the shear stress distribution is more uniform if the bone layer is exposed to the fluid inlet path which facilitates bone differentiation. Good adhesion and infiltration of cells were observed after 14 days dynamic culture. The results of expression analysis of differentiated genes in bone and cartilage layer containing RADSc after 21 days of culture under static and dynamic conditions showed that perfusion flow significantly upregulated the expression of bone and cartilage genes in the respective layers and downregulated the hypertrophy gene expression in intermediate layer of scaffold.