Promotion of neurite outgrowth and synapse formation is a key step for nervous tissue regeneration. It is important for finding a new biomaterial to guide neuron growth to target neurons. Aminated graphene oxide (NH2-GO) displays electrical properties and dispersibility, which may change the surface charge of neurons and further activate neuronal excitement. However, the molecular guidance mechanism of NH2-GO on neurite outgrowth is seldom reported. In this study, we compared the role of NH2-GO on the spinal cord neurons and cortical neurons. Results indicated that the proper concentrations were at 2 and 4 μg/mL as determined by the CCK-8 assay. Notably, NH2-GO (2 and 4 μg/mL) improved the dispersibility and strengthened the effect of the composite material. In addition, it enables biocompatibility and efficient guidance of growth performance, which is not neurotoxic for neuronal outgrowth under these two concentrations. More interestingly, NH2-GO at 2 μg/mL induced both marked neurite elongation and increased branches in cortical neurons, but there is no significant change of neurite length and branches in spinal cord neurons. Further, the fluorescence intensity and mRNA level of Netrin-1 and DCC (Deleted in Colorectal Cancer) were both enhanced by NH2-GO at 2 μg/mL. Moreover, the function of Netrin-1 and DCC were activated more significantly by NH2-GO at 2 μg/mL in cortical neurons than that of spinal cord neurons. When RhoA was inhibited by the C3 exoenzyme, phosphorylated Rac1 and Cdc42 expression decreased significantly. Thus, NH2-GO at 2 μg/mL could influence Netrin-1/DCC signaling and the downstream RhoGTPase pathway, which may be preferred to guide the neurite growth in cortical neurons. It will provide a promising approach for the development of novel therapeutic methods of nerve regeneration.
Keywords: DCC; RhoGTPase; aminated graphene; nerve regeneration; netrin-1; neurite outgrowth.