Modulation Of Adhesion And Proliferation Of Mesenchymal Stem Cells Cultured On Silk Biomaterial Modified With RGD Peptide

Object: In order to promote the seed cell's proliferation, adhesion on the tendon tissue engineering scaffold and to promote the synthesis and the secretion of the extracellular matrix. So the silk matrix modified with RGD peptide was prepared, the proliferation and adhesion of MSC on the modified scaffold was investigated.Methods: The GRGDS sequence was covalent on the surface of silk material by the cross linking agent (EDC: NHS) and detected the contents of the Carbon,Nitrogen and Oxygen element by the X-ray photoelectron spectroscopy (XPS). The third generation of mesenchymal stem cells (MSC) was seeded onto the RGD modified silk film, the silk film and the tissue culture plate, and incubated for 4h and 12h at standard condition, in each experiment the number of adhesive cells was counted to evaluate the efficiency of cell adhesion, incubated for 1d, 2d, 3d and 4d, the cell density was calculated and was used as index of cell proliferation. MSC was seeded onto the RGD modified silk film and the silk film, using the inverted phase contrast microscope to observe the cells adherent appearance and cells spreading morphous, using the immunofluorescene technique labeling the cell skeleton after 24h, and observed using the Laser Confocal Microscopy.Results: The XPS detection showed that the contents of the Carbon,Nitrogen and Oxygen element were significant different in the surface between the modified and the non-modified. We found after 4h and 12h culture the efficiency of cell adhesion in the modified silk scaffold was higher than the unmodified silk scaffold, no significant difference between the modified silk scaffold and the tissue culture plate .No significant difference in cell density in each group for culture of 1d, 2d, 3d and 4d. After 24h culture the cells'adherent appearance and cells spreading morphous on the modified silk film were better than the unmodified. Observing undering the Laser Confocal Microscopy the quality and the quantity of cell adhesion on the modified silk film were better than the unmodified and their cytoskeleton was more robust.Conclusion: The GRGDS sequence can be covalent on the surface of silk material by the cross linking agent (EDC: NHS) successfully. RGD peptide enhanced cell adhesion to silk scaffold, but no obvious effects were observed in cell proliferation.