(C-88) Effects of Interstitial Matrix Composition on the Properties of Hydrogel Microparticle-Based Scaffolds for Use in Cell Delivery

Hydrogel microparticle-based materials are of broad interest for wound healing, drug and cell delivery, and biosensor systems. These materials are particularly attractive as scaffolds for use in cell delivery, as cells can be incorporated into the void space between particles, and the particle properties can be engineered to influence cell growth and function [1]. However, cells incorporated within these scaffolds can have difficulty remaining evenly distributed within them. Poor incorporation can lead to inadequate cell delivery for therapeutic applications such as cell-based cancer treatments, and bone and tissue regeneration, limiting the overall effectiveness of these therapies. We hypothesize that adding a soft interstitial matrix within the scaffolds will allow for a more even distribution of incorporated cells compared to scaffolds without such a matrix, potentially improving the therapeutic effect of the delivered cells. This study covers the fabrication and characterization of a hydrogel microparticle-based platform designed with a soft interstitial matrix and provides a foundation for future studies on their use in cell delivery applications.

Poly(ethylene glycol) (PEG) microparticles were prepared using a formulation comprising of 10 weight % 4-arm 20kDa PEG-norbornene, crosslinking peptide (KCGPQGIAGQCK), photoinitiator (Lithium phenyl-2,4,6-trimethylbenzoylphosphinate - LAP), and cell adhesive peptide (CGRGDS) through electrospraying technique. The thiol:norbornene ratio of the synthesized PEG microparticles was 0.75:1.

The PEG microparticles were then washed and compacted in a 1 mL syringe using a 5.0 μm glass fiber syringe filter to remove any excess fluid. Subsequently, the annealing solution, containing PEG-norbornene (4 arm 20 kDa) as the interstitial matrix, along with crosslinking peptide and photoinitiator, was added to the packed particles. The annealing solution was prepared using a crosslinking thiol:norbornene ratio of 1.71. Four different concentrations of PEG-norbornene (0%, 0.5%, 1%, and 2% volume/volume) were used to investigate its effects.

Molds for the scaffolds were made using a clean glass slide treated with Rain-X water repellent and a 1 mm thick silicon sheet with four 8 mm diameter cut-outs. The particle solution was then added to the molds and exposed to UV light for the annealing process. Finally, the annealed scaffolds were placed in a 24-well plate containing phosphate-buffered saline (PBS) and allowed to swell over several days.

The PEG microparticles themselves were imaged under a light microscope after being dyed with trypan blue to determine their size. The shear storage moduli of the scaffolds were determined using an Anton Paar rheometer, and statistical analysis of the experimental data was performed using a one-way ANOVA.

The focus of this study was on the characterization of a novel hydrogel microparticle-based scaffold containing a soft interstitial matrix (Fig. 1a,b). The size distribution of the PEG microparticles used to create the scaffolds had a mean diameter of 253.7 μm, with a median of 207.5 μm and a standard deviation of 184.6 μm. A total of 190 particles were analyzed. The incorporation of varying concentrations (0%, 0.5%, 1%, and 2% volume/volume) of PEG-NB as an interstitial matrix did not lead to statistically significant differences in the storage modulus of the resulting scaffolds (p > 0.05, p = 0.8565 one-way ANOVA). The storage moduli were in the range of 2-2.5 kPa (Fig. 1c). These findings indicate that the presence of this matrix within the scaffold did not significantly alter their overall mechanical properties.

This study provides a foundation for further investigations into the optimization of hydrogel microparticle-based scaffolds for cell delivery. Future research plans include the use of a nanoindenter to generate a topographical stiffness map of the scaffolds to further characterize the interstitial matrix, and additionally a comparison of the ability to encapsulate cells within the scaffolds containing an interstitial matrix to control scaffolds that do not. Through conducting these planned investigations, we hope to demonstrate the potential this technology has as an improved cell delivery platform and by extension the potential it has in improving the therapeutic effectiveness of current cell therapies.

[1]        S. Xin, O. M. Wyman, and D. L. Alge, "Assembly of PEG Microgels into Porous Cell-Instructive 3D Scaffolds via Thiol-Ene Click Chemistry," Advanced Healthcare Materials, vol. 7, no. 11, p. 1800160, 2018, doi: https://doi.org/10.1002/adhm.201800160.