Polyethylene glycol hydrogel (PEG) conjugated with arginyl glycyl aspartic acid (RGD) (PEG-RGD) has been considered to be a scaffold in three-dimensional (3D) culture that improves neurite outgrowth; on the other hand, tenascin C controls neural growth and differentiation. In this study, the effect of a combined RGD and tenascin C mixture in 3D culture (3D-PEG-RGD-TnC) on the survival, growth and differentiation of neural stem cells. The viability of the culture has been evaluated by live/dead assay and the results show that the viability of NSCs in 3D-PEG-RGD-TnC is significantly higher than its value in 3D-PEG-RGD. The proliferation was evaluated by MTS test and was found to be slightly improved but statistically not significant. Accordingly, the differentiation was evaluated by immunoreactivity to nestin, neurofilament 68, neurofilament 160, neurofilament 200 and GFAP; and the expression of nestin, neuro D, musashi1, β-tubulin III, GFAP, MBP and Oct4 was studied using RT-PCR. The results showed enhancement of the differentiation of NSCs into the neuronal phenotype in 3D-PEG-RGD-TnC. The morphology of NSCs cultured in 3D-PEG-RGD-TnC showed neurite outgrowths and increase in the contact between the differentiated cells' extensions. The conclusion of this study was that NSC survival, proliferation and differentiation are enhanced when the cells are cultured in 3D-PEG-RGD-TnC.
Keywords: 3D culture; RGD peptide; extracellular matrix; neural stem cells; polyethylene glycol; tenascin.
Copyright © 2014 John Wiley & Sons, Ltd.