Electrically Stimulated MXene-Cryogel Scaffolds for Mineralization in Bone Defects

Introduction:: The field of bone regeneration focuses on the fabrication of biological substitutes to support cellular infiltration, matrix deposition, and mineral formation in native tissue. Previous work has demonstrated that the natural piezoelectric properties of the collagen matrix and overall electrokinetic effects aid in bone formation and resorption; however, due to high cost, efficacy, and challenges mimicking in-vivo properties, techniques aimed at providing electrical stimulation are not commonly used. Further, studies by our group and others have demonstrated cryogel scaffolds’ potential for supporting bone growth due to their macroporous, durable, and biocompatible structure; however, no work has incorporated MXene within cryogels. To harness the potential for bone formation via electrical stimulation and aid in mineralization, cryogels were incorporated with varying concentrations of MXene. We hypothesize that an optimized MXene cryogel will initiate mineralization and enhance osteogenic differentiation when combined with externally applied electrical stimulation.

Materials and Methods:: Chitosan-gelatin (1:4) cryogels were fabricated with various concentrations of MXene (30%, 50%, 70%) in solution or hydrothermally coated following formation. 2D MXene solution was synthesized from a ceramic ‘MAX’ phase and then chemically processed to etch out ‘A’ atoms. Cryogel solutions containing appropriate concentrations of chitosan, gelatin, and MXene were crosslinked, poured into 3-mL syringes, and immediately frozen at sub-zero temperatures (-20°C). Following 18 hours of freezing, cryogels were allowed to thaw at room temperature. Finally, all cryogel constructs were re-frozen at -80°C and lyophilized for 12 hours. Cryogels fabricated without MXene solution were hydrothermally coated in MXene and re-lyophilized. Cryogels fabricated without MXene served as a control. Porosity, conductivity, swelling kinetics, and mechanical integrity were evaluated with SEM, EIS, and compressive testing to determine the effects of MXene additives.

Results, Conclusions, and Discussions:: This study has demonstrated that MXene can be homogenously incorporated into the framework of cryogels without disrupting the advantageous material properties novel to these scaffolds (i.e., macroporous and spongy structure). Specifically, the highest concentration (70% (v/v) MXene) possessed swell potential and pore sizes required for cellular infiltration without significant deviation from the control (Figure 1A, B); these results were also noted among the coated and lower MXene concentrations. Interestingly, as MXene concentration increased, swelling potential also increased. Additionally, it was demonstrated that the methodology for MXene incorporation had no significant impact on the compressive properties after 50% strain, where macro-structural integrity was observed without crack propagation (P< 0.05; Figure 1C). Finally, preliminary analysis supports that the increase of MXene concentration within cryogels increases the conductivity of the scaffold indicated by the decrease in resistance (Figure 1D). Overall, incorporation of MXene did not hinder cryogel pore formation, sizing, and compressive properties vital for incorporation in bone defects. Rather, it enhanced the swelling kinetics of the cryogels while introducing a conductive nature to the cryogel that can aid in mineralization.

Previously recognized for its attractive properties, MXene, in this work, continues to show promise as a novel dopant in the realm of scaffold fabrication. Pore analysis, swelling, and compressive testing demonstrated that MXene incorporation in a cryogel does not impede on critical architectural traits necessary for cryogel scaffolds to target bone regeneration. Ongoing and future work is seeking to identify the differences in impedance between doping methodologies and their individual capabilities to improve cellular infiltration and differentiation when stimulated by electropositive and negative potentials.

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References (Optional): : Chaudhari, N.K., et al., Journal of Materials Chemistry A, 2017.; Hixon, K.R., et al., Acta Biomaterialia, 2017.; Khare, D., et al., Biomaterials, 2020.