Hydrogels based on Fmoc-diphenylalanine and Fmoc-diglycine for nucleus pulposus disc tissue engineering

摘要

With a view to repair the degenerated nucleus pulposus (NP) of intervertebral discs, we grew bovine NP cells on fluorenylmethoxycarbonyl-diphenylalanine (Fmoc-FF)/ Fmoc-diglycine (Fmoc-GG) hydrogels. The gels prepared in two different molar ratios of Fmoc-FF to Fmoc-GG, 1:0 and 1:1. These gels were characterized in terms of their physical and biological properties including fibre structure, cell attachment, collagen and sulphated glycosaminoglycan (s-GAG) productions. Fmoc-FF and the composition of 1:1 Fmoc-FF/Fmoc-GG hydrogels (20 mmol/L) were synthesized by utilizing a sequential change in pH (Jayawarna et al, 2007). Fibre structure of hydrogels was determined by using CryoSEM. In vitro studies of hydrogels seeded by NP cells were performed using red fluorescent cell membrane dye PKH-26, confocal microscopy, Sircol collagen and DMMBTM assays. Cryo-SEM showed a dense network of fine fibres in Fmoc-FF, whereas an overlapping mesh of flat ribbons was seen in the composition of 1:1 Fmoc-FF/Fmoc-GG. Confocal microscopy showed the majority of the NP cells stained with PKH-26 remained in a rounded morphology within both Fmoc-FF and the composition of 1:1 Fmoc-FF/Fmoc-GG hydrogels after 5 days of culture. SircolTM and DMMB assays showed deposition of collagen and sulphate-glycosaminoglycan by NP cells cultured within both Fmoc-FF and the composition of 1:1 Fmoc-FF/Fmoc-GG over 3 weeks of culture. Taken together, the results of present study provide preliminary evidence for the use of a self-assembling peptide hydrogel as a scaffold for the synthesis and accumulation of NP-like extracellular matrix for intervertebral disc tissue repair.