Regulation Of Cyclic Tensile Stress Combined With Elasticity Of PDMS Substrates On The Rabbit Annulus Fibrosus-derived Stem Cells

Objective: Planted the rabbit fibrosus-derived stem cells(AFSCs)on polydimethylsiloxane(PDMS)substrates with different elastic modulus,cyclic tensile stress(CTS)was applied at the same time,to investigate the differentiation effect of cyclic tensile stress combined with different elastic PDMS substrates.Methods: Different elastic PDMS substrates were prepared by different compound ratios of 1: 40,1: 20,1: 10,respectively.Isolated and cultured rabbit fibrosus-derived stem cells and seeded on silicone dishes with different elastic PDMS substrates.Treated with static culture and appropriate CTS(0.5Hz frequency,5% strain,4 hours / day)respectively for 3 days.Mechanical and water contact angle test for elastic modulus,hydrophilicity and hydrophobicity of the PDMS substrates.RT-q PCR and Elisa assay were used to detect the expression of genes and related proteins after culturing under different conditions.Cell proliferation and skeletal staining test cell growth,morphological changes.Results: The results of mechanical test indicated that the elastic modulus of three kinds PDMS substrates were 5.95 Mpa,10.33 Mpa and 15.41 Mpa respectively.Water contact angle test showed no significant difference of hydrophily and hydropobicity between the three kindssubstrates.The expression of Col-I gene increased with the increase of substrate modulus,while the expression of Col-II and aggrecan genes decreased.In the low-elasticity substrate group,the expression of Col-II and aggrecan genes of cells with CTS-stimulated was obviously higher.In the high-elasticity substrate group,the expression of Col-I genes of cells with CTS-stimulated cells was obviously higher.Medium elastic substrate group did not show significant difference.Related protein levels results are similar to gene expression.Conclusion: In our study,PDMS materials were used to fabricate different elastic modulus substrates that simulate the three regions from the inner to outer of the annulus fibrosus.At the same time,the character of PDMS materials which can meet the requirements of tensile stress was also utilized.We detected that there was certain regulating effect of suitable CTS combined with different elastic PDMS substrates to the expression of genes and related proteins of AFSCs.Our results can provide important reference for the regulation and differentiation of annulus fibrosus tissue engineering from two important aspects of annulus fibrosus biomechanics.