Biosynthesis and characterization of a neuroprotective injectable copolymeric scaffold impregnated with GOQD-infused HA-collagen electrospun nanofibers: A strategy to prevent oxidative damage, plaque formation, and neurodegeneration.

In this study, an injectable matrix was fabricated using electrospun collagen/Hyaluronic acid/GOQD nanofiber (Collagen-HA-GOQD) and thermoresponsive N-succinyl chitosan-grafted-carboxyl-terminated poly-N-isopropyl acrylamide (SCS-g-PNIPAAm-COOH) hydrogel. Different characterization tools authenticated the synthesis of SCS-g-PNIPAAm-COOH, GOQD (size ∼5 nm), and Collagen-HA-GOQD nanofibers with average diameter of -153 nm. The nanocomposite exhibited a 3D honeycomb-shaped porous structure, and its sol-gel transition occurred at 33.4 °C with G' of 186 ± 0.5 Pa at body temperature. The scaffold's components affected the bio-interfacial interaction, as presence of nanofiber ameliorated the properties of the copolymer. Scaffold 3 containing collagen-HA-GOQD nanofibers exhibited significantly higher MTT metabolic activity than the other groups, as well as it promoted significantly higher cell adherence and proliferation of SH-S5Y5 cells compared to the other two scaffolds and the control group. Scaffold 3 also exhibited the highest fluorescence intensity at each time interval, followed by Scaffold 2 and Scaffold 1. Increased mRNA expression of the proliferation-associated markers PCNA and Ki67, together with the neural progenitor-associated marker Nestin, in Scaffold 3-treated SH-SY5Y cells were observed compared with the other groups. Further, the upregulated expression of talin, integrin β1, phospho-ERK, and NeuN in the Scaffold 3-treated group suggests a correlation in their involvement in integrin-mediated cell-matrix interaction, and neuronal-like cellular responses, which requires further mechanistic studies to confirm. The enhanced mRNA expression of NeuN in Scaffold 3 further postulated its crucial role in promoting cellular differentiation and maturation. The inhibitory effect of Scaffold 3 on Aβ-peptide aggregation was established, and it also disrupted the preformed amyloid plaque in cells and mitigated the induced oxidative stress. Thus, scaffold 3 can be projected as a suitable matrix to trigger neuronal-like cellular responses. Unlike most Alzheimer's Disease therapies that target a single pathology, this scaffold can provide a regenerative niche, inhibit amyloid aggregation, and scavenge ROS - a multi-pronged in vitro platform that warrants further validation in differentiated neuronal models and in vivo studies.