Controlling Neuronal Cell Growth through Composite Laminin Supramolecular Hydrogels

Abstract

Designing an extracellular matrix mimic by biofunctionalization of polymeric scaffolds is a popular strategy and extremely crucial for facilitating the interactions between cells and the matrix. To this direction, supramolecular gels are gaining exponential attention over the last few years, owing to their potential biocompatibility and biodegradability. In spite of diverse biological roles of native laminin, the bioactivities of self-assembling laminin-derived short peptides were less explored. In this work, we have explored the minimalist design to develop hydrogel scaffolds based on IKVAV and YIGSR peptides individually and their composite matrix, which can provide structurally and functionally relevant materials for tissue engineering. Till date, composite supramolecular gels solely made up of self-assembling IKVAV and YIGSR peptides have never been reported. Such composite gels can be a closer mimic of natural laminin protein, which could mimic the essential functions of the short peptide fragments present on different chains of the extracellular matrix protein, laminin. Interestingly, we used a unique strategy of simple mixing of the two laminin mimetic peptides, which tend to induce coassembly with a self-sorted nanofibrous network with relatively enhanced mechanical strength. The physicochemical properties of the biofunctional hydrogels were studied using different microscopic, spectroscopic, and rheology techniques. To assess the bioactivity of laminin-derived scaffolds in controlling neuronal cell growth, its biocompatibility, cellular growth, and proliferation were quantified using C6 glial cells and SHSY5Y neuroblastoma cells. The live/dead staining further confirmed the adhesion and proliferation of the cells. A significant increase in neurite length provides clear evidence on mimicking the neurite extension function of native laminin protein by its short derivatives. Interestingly, similar β-III tubulin expression and cell cycle phases were observed, in comparison to control, which indicated normal cellular functioning of the cells cultured over short laminin hydrogel scaffolds. All bioassays suggested that Fmoc YIGSR promotes growth of neural cells to a greater extent and maintains healthier morphology, in comparison to hydrophobic Fmoc IKVAV, owing to the entangled longer fibrous network formed by YIGSR peptide. It is expected that thinner long fibers provide a more uniform surface and are more supportive for cell adhesion in comparison to hydrophobic, shorter fibers IKVAV peptide. However, in composite gels, the detrimental effect of hydrophobic IKVAV peptide could be reduced and better adhesion and proliferation could be achieved along with enhanced cell survival. These observations demonstrate the high potential of the laminin-derived hydrogels in tissue engineering and neuronal stem cell differentiation in future.