Abstract
Here we report three new nanofibrous, self-assembling multidomain peptide (MDP) sequences and examine the effect of sequence on the morphology and expansion of encapsulated Stem cells from Human Exfoliated Deciduous teeth (SHED). We modified our previously reported set of serine-based MDPs, changing the serine residues in the amphiphilic region to threonine. The three new threonine-based sequences self-assemble into antiparallel β-sheet nanofibers, confirmed by CD and IR. AFM and negative-stained TEM show that the nanofibers formed by the new sequences are more curved than their serine-containing predecessors. Despite this change in nanofiber morphology, SEM illustrates that all three new sequences still form porous hydrogels. K(TL)2SLRG(TL)3KGRGDS, with a designed cleavage site, is able to be degraded by Matrix Metalloprotease 2. We then examine SHED cell response to these new sequences as well as their serine-based predecessors. We observe faster cell attachment and spreading in hydrogels formed by K2(SL)6K2GRGDS and K(SL)3RG(SL)3KGRGDS. By day 3, the SHEDs in all of the serine-based sequences exhibit a fibroblast-like morphology. Additionally, the SHED cells expand more rapidly in the serine-based gels while the cell number remains relatively constant in the threonine-based peptides. In hydrogels formed by K2(TL)6K2GRGDS and K(TL)2SLRG(TL)3KGRGDS, this low expansion rate is accompanied by changes in morphology where SHEDs exhibit a stellate morphology after 3 days in culture; however, by day 7 they appear more fibroblast-shaped. Throughout the duration of the experiment, the SHED cells encapsulated in the K2(TL)6K2 hydrogels remain rounded. These results suggest that the basic MDP structure easily accommodates modifications in sequence and, for SHED cells, the threonine-containing gels require the integrin-binding RGDS sequence for cell attachment to occur, while the serine-based gels are less selective and support an increase in cell number, regardless of the presence or absence of RGDS.