Self-Assembling Peptide Scaffold Carrying Neural-Cell Adhesion Molecule-Derived Mimetic-Peptide Transplantation Promotes Proliferation and Stimulates Neurite Extension by Modulating Tau Phosphorylation and Calpain/Glycogen Synthase Kinase 3 beta (GSK-3β) in Neurons

BACKGROUND: Self-assembling peptide scaffolds have been extensively applied in tissue engineering. Many investigations have modified self-assembling peptide scaffolds by integrating functional motifs, with promising applications. This study aimed to generate a novel RADA16 self-assembling peptide scaffold integrating a neural-cell adhesion molecule-derived mimetic-peptide (SIDRVEPYSSTAQ) and evaluated the effects on neuron proliferation. MATERIAL/METHODS: A 37-amino-acids peptide of RADA16-activation motif containing neural-cell adhesion molecule-derived mimetic-peptide (SIDRVEPYSSTAQ) was synthesized and self-assembled into a scaffold. Dorsal root ganglion (DRG) and spinal cord motor neurons (SCMN) were primarily isolated and identified. Neurons (DRG and SCMN) were divided into FRM, FRM-MP, and FRM-MP-LiCl groups. The adherence ability of neurons was evaluated using toluidine blue staining. Proliferation and apoptosis of neurons were assessed using CCK-8 and flow cytometry assay, respectively. Immunofluorescence assay was used to measure neurite extension. Western blot assay was used to assess GSK-3β/p-GSK-3β, Tau/p-Tau, and calpain expression in neurons. RESULTS: FRM-MP-LiCl released multiple-peptide with higher efficiency. FRM-MP-LiCl significantly enhanced proliferation and inhibited apoptosis compared to FRM and FRM-MP groups (p<0.05). FRM-MP-LiCl incubation increased adherent ability compared to the FRM and FRM-MP groups (p<0.05). FRM-MP-LiCl remarkably increased neurite length (p<0.05), numbers (p<0.05), and branches. FRM-MP-LiCl significantly reduced GSK-3β phosphorylation (p-GSK-3β/GSK-3β) and Tau phosphorylation (p-Tau/Tau) compared to the FRM and FRM-MP groups (p<0.05). FRM-MP-LiCl significantly downregulated calpain compared to the FRM and FRM-MP groups (p<0.05). CONCLUSIONS: The generated self-assembling peptide scaffold carrying FRM motif (SIDRVEPYSSTAQ) promoted neuron proliferation and adherent ability, inhibited apoptosis, and stimulated neurite extension, by reducing Tau protein phosphorylation through the calpain/GSK-3β signaling pathway.