Three-dimensional self-assembling peptide matrix enhances the formation of embryoid bodies and their neuronal differentiation

Abstract

We have tailored the properties of a self-assembling peptide (SAP) matrix to direct embryonic stem cells towards neuronal differentiation by adopting a three-dimensional (3D) culture, matching mechanical strength with that of neural tissue, and incorporating fixed laminin-derived pentapeptide signals (IKVAV). We report here that such a matrix alone can induce mouse embryonic stem (ES) cells to first develop into embryoid body (EB) and increase their propensity for subsequent neuronal differentiation. Embryoid bodies were observed by day 5 of culture in SAP matrix. βIII-tubulin as an early neuronal commitment marker was more prominently detected in cells cultured in the matrix containing IKVAV signals. Interestingly, ES-derived cells did not display distinct neuron morphology within the 3D culture; however, 55 ± 10% of those cells within IKVAV conjugated matrix and 38 ± 6% of those within base matrix displayed higher potential towards neuronal differentiation after 7 days. When retrieved and recultured on a tissue culture plate, they exhibited extended neurite outgrowths and networks in the absence of any additional neuronal differentiation growth factor. The up-regulated expression of neuronal development markers (MAP2 and MeCP2) and the down-regulation of glial marker (GFAP) support that further neuronal differentiation takes place upon reculture. The results showed that an artificial matrix composed of designer SAPs could prompt the formation of EB and provides the cues favoring neuronal differentiation of ES cells. © 2013 Wiley Periodicals, Inc.