Hydrogel Mesh Size Research Articles

Calcium Cross-Linked Cellulose Nanofibrils: Hydrogel Design for Local and Controlled Nitric Oxide Release.

Nitric oxide (NO) holds promise for wound healing due to its antimicrobial properties and role in promoting vasodilation and tissue regeneration. The local delivery of NO to target cells or organs offers significant potential in numerous biomedical applications, especially when NO donors are integrated into nontoxic viscous matrices. This study presents the development of robust cellulose nanofibril (CNF) hydrogels designed to control the release of nitric oxide (NO) generated in situ from a NO-donor molecule (S-nitrosoglutathione, GSNO) obtained from the nitrosation of its precursor molecule glutathione (GSH). CNF, efficiently isolated from sugar cane bagasse, exhibited a high aspect ratio and excellent colloidal stability in water. Although depletion forces could be observed upon the addition of GSH, this effect did not significantly alter the morphology of the CNF network at low GSH concentrations (<20 mM). Ionic cross-linking with Ca2+ resulted in nontoxic and robust hydrogels (elastic moduli ranging from 300 to 3000 Pa) at low CNF solid content. The release rate of NO from GSNO decreased in CNF from 1.61 to 0.40 mmol. L-1·h-1 when the nanofibril content raised from 0.3 to 1.0 wt %. The stabilization effect monitored for 16 h was assigned to hydrogel mesh size, which was easily tailored by modifying the concentration of CNF in the initial suspension. These results highlight the potential of CNF-based hydrogels in biomedical applications requiring a precise NO delivery.

Hydrogel interfaced glass nanopore for high-resolution sizing of short DNA fragments

Solid-state nanopores, known for their label-free detection and operational simplicity, face challenges in accurately sizing the short nucleic acids due to fast translocation and a lack of enzyme-based control mechanisms as compared to their biological counterparts with sizing resolutions still limited to ≥100 bp. Here, we present a facile polyethylene glycol-dimethacrylate (PEG-DMA) hydrogel interfaced glass nanopore (HIGN) system by inserting glass nanopore into the hydrogel to achieve sub-100 base pair (bp) resolution in short DNA sizing analysis. We systematically investigated the effects of hydrogel mesh size, spatial configurations of glass nanopores about the hydrogel, applied bias voltage, and analyte concentration on the transport dynamics of 200 bp double-stranded DNA (dsDNA). A 7.5 w/v% PEG-DMA hydrogel induced ∼11x increase in the mean dwell times compared with bare solution nanopore system. The insertion locations and depths of the glass nanopore into the hydrogel resulted in 7.16% and 5.28% coefficients of variation (CV) for mean normalized event frequencies. This enhancement of dwell times and invariability in translocation characteristics enables precise sizing of dsDNA fragments under 400 bp using HIGN, with an achieved size resolution of 50 bp with observable mean normalized peak amplitude (ΔI/Io) of ∼0.005. Furthermore, we have demonstrated the capability of HIGN to perform multiplex detection of influenza A virus (IAV) and severe acute respiratory syndrome-coronavirus-2 (SARS-CoV-2) through reverse transcriptase-polymerase chain reaction (RT-PCR). These results demonstrated the potential of HIGN as a versatile tool in nucleic acid analysis and multiplexed label-free molecular diagnostics.

Controlled stiffness and diffusivity of poly(ethylene glycol) hydrogel formed with cellulose-nanofiber framework

Hydrogels composed of polymer networks are widely used in industry and scientific research for high water retention and unique mechanical properties. Nevertheless, in ordinary hydrogel formation, the trade-off relationship between stiffness and mesh size remains a crucial consideration for practical applications. This study describes a facile approach to controlling hydrogel stiffness and mesh size by hybridizing poly (ethylene glycol) diacrylate (PEGDA) and methacrylated TEMPO-oxidized cellulose nanofibers (T-CNFMA). After disintegrating T-CNF by ultrasonication, T-CNFMA was synthesized resulting in a degree of substitution of 2.04 mmol/g. Incorporation of T-CNFMA in the PEGDA network allowed for independent control of hydrogel stiffness and mesh size by reinforcing the whole hydrogel network as a framework. Consequently, the swelling ratio and shear modulus could be manipulated by controlling the PEGDA/T-CNFMA ratio. Structural analyses revealed that an increase in the T-CNFMA content in the presence of a low amount of PEGDA resulted in a large mesh size with a constant stiffness. The diffusivity test was also consistent with the properties of the hydrogels. This result indicates that the incorporation of T-CNF in hydrogel network is useful to control the physical property of the hydrogel, especially for varying mesh size, regardless of stiffness alteration.

Investigating the structure and properties of polyurethane hydrogels with varying soft and hard segments

AbstractIn this study, polyurethane (PU) hydrogels were synthesized via mercapto curing reaction to elucidate the effect of molecular interactions between isocyanate and soft segments on the properties of hydrogels. Further, the mesh size, mechanical properties, hydrophilicity, and biological properties of the PU hydrogels were determined. In the isocyanate series, the structural regularity and rigidity of 4,4′‐dicyclohexylmethane diisocyanate (HMDI) favored the formation of hydrogel materials with small mesh size, high modulus, and low water absorption. In contrast, l‐lysine diisocyanate (LDI) favored the materials with large mesh size, low modulus, and good hydrophilicity. In the soft‐segment series, the strong hydrogen bonds of polycarbonate diol (PCDL) favored the formation of materials with small mesh size, dense cross‐link points, and high modulus, whereas weak hydrogen bonds of polytetrahydrofuran ether glycol (PTMG) favored the hydrogel materials with small mesh size, few crosslink points, and low modulus. PU hydrogels exhibit excellent cytocompatibility, anti‐cell adhesion, and anti‐inflammatory properties. Therefore, this study offers valuable insights into understanding the chain structure and macroscopic properties, thus contributing to preparing PU hydrogels with varying performances, as desired.

Extremely large Seebeck coefficient of gelatin methacryloyl (GelMA)-based thermogalvanic cells by the dual effect of ion-induced crystallization and nanochannel control

The quasi-solid state thermogalvanic cells (TGCs) have been extensively investigated due to their high Seebeck coefficient (S) and suitability for wearable power generation. However, the influence of hydrogel nanostructure on Seebeck coefficient remains underexplored. In this study, biocompatible TGCs based on gelatin methacryloyl (GelMA) are fabricated via a fast-photocrosslinking process. Moreover, guanidium chloride (GdmCl) salts are directly introduced into the TGCs to enhance the Seebeck coefficient via ion-induced crystallization in the p-type FeCN63−/FeCN64− redox system. Small angle X-ray scattering reveals controlled GelMA hydrogel mesh size adjusting the amount of GdmCl, leading to improved intermolecular interactions between the guanidium cations and GelMA polymer. The optimized p-type Seebeck coefficient reaches an unprecedented 21.6 mV K−1, which is the highest thus far reported in the context of the TGC system. Furthermore, a prototype wearable thermoelectric generator (TEG) with an output voltage of 1.6 V under a temperature difference (ΔT) of 9 °C is able to power an LED directly without requiring an additional voltage booster. In brief, the dual effect of ion-induced crystallization and nanochannel control substantially enhances the Seebeck coefficient of the TGC, thereby offering a promising strategy for enhancing the performance of low-grade thermal energy harvesting wearable TEGs.

Adenylate Kinase Fused to Spidroin as a Catalyst for Decreasing Leakage out of 3D-Bioprinted Hydrogels and for ATP Regeneration.

Numerous metabolic reactions and pathways use adenosine 5'-triphosphate (ATP) as an energy source and as a phosphorous or pyrophosphorous donor. Based on three-dimensional (3D)-printing, enzyme immobilization can be used to improve ATP regeneration and operability and reduce cost. However, due to the relatively large mesh size of 3D-bioprinted hydrogels soaked in a reaction solution, the lower-molecular-weight enzymes cannot avoid leaking out of the hydrogels readily. Here, a chimeric adenylate-kinase-spidroin (ADK-RC) is created, with ADK serving as the N-terminal domain. The chimera is capable of self-assembling to form micellar nanoparticles at a higher molecular scale. Although fused to spidroin (RC), ADK-RC remains relatively consistent and exhibits high activity, thermostability, pH stability, and organic solvent tolerance. Considering different surface-to-volume ratios, three shapes of enzyme hydrogels are designed, 3D bioprinted, and measured. In addition, a continuous enzymatic reaction demonstrates that ADK-RC hydrogels have higher specific activity and substrate affinity but a lower reaction rate and catalytic power compared to free enzymes in solution. With ATP regeneration, the ADK and ADK-RC hydrogels significantly increase the production of d-glucose-6-phosphate and obtain an efficient usage frequency. In conclusion, enzymes fused to spidroin might be an efficient strategy for maintaining activity and reducing leakage in 3D-bioprinted hydrogels under mild conditions.

Tunable enzymatically degradable hydrogels for controlled cargo release with dynamic mechanical properties.

Here, we designed enzymatically degradable hydrogels with tunable mesh sizes and crosslinking points to evaluate the effectiveness of network structure estimations in predicting dynamic mechanical properties and cargo retention or release. Poly(ethylene glycol) (PEG) hydrogels were prepared through a thiol-ene click reaction between four- or eight-arm PEG functionalized with vinyl sulfone and cysteine residues of collagenase-degradable peptides to create well-defined, homogenous, and robust materials with a range of mesh sizes estimated from the elasticity theory or Flory-Rehner theory. Time-dependent changes in mechanical properties associated with hydrogel degradation, i.e., dynamics of storage modulus, which is determined by the relationship between the hydrogel mesh and enzyme sizes, were characterized. The shear modulus G' decreased by enzyme addition, and the degradation rate decreased with the initial crosslinking density of the hydrogel. The degradation rate could also be controlled with the reactivity of peptide sequences against collagenase. With these findings, the retention and release of FITC-dextran were successfully controlled by tuning the mesh size and degradability of the hydrogel. This report provides useful insights for designing hydrogels as cell scaffolds or functional molecular delivery matrices with tunable dynamic mechanical properties and the resulting release of loaded drugs or proteins.

Polysaccharide gum based network hydrogels for controlled drug delivery of ceftriaxone: Synthesis, characterization and biomedical evaluations

In the present work dietary fibers gum acacia (GA) and tragacanth gum (TG) were applied in designing drug delivery (DD) carrier due to their therapeutic role and colon degradation. The grafted product was formed in the form of network hydrogels with polyacrylamide [poly(AAm)]. The polymer samples were characterized by Scanning electron micrograph (SEM), Electron dispersion X-ray analysis (EDAX), X-ray photoelectron spectroscopy (XPS), X-ray diffraction (XRD), Fourier transform infrared spectroscopy (FTIR) and Thermo-gravimetric analysis (TGA). The formation of the grafted product and its heterogenous and amorphous nature was confirmed from FTIR, SEM and XRD. The mesh size of the network hydrogels was about 32.51 nm. The antibiotic drug ceftriaxone diffusion was non-Fickian from drug encapsulated GA-TG-cl-poly(AAm) network and release was fitted in first order kinetic model. The release of drug from network was slow and controlled manner due to drug-polymer interactions porous nature of the hydrogels. The polymer-blood interaction showed haemolysis values less than five percent which inferred biocompatible nature and polymer-biomembrane interactions indicated the muco-adhesiveness of the DD system. These properties reflected the suitability of the network hydrogel for site specific drug delivery.

Hydrogel Mesh Size and Its Impact on Predictions of Mathematical Models of the Solute Diffusion Coefficient

Hydrogel Mesh Size and Its Impact on Predictions of Mathematical Models of the Solute Diffusion Coefficient

Regulation of cell attachment, spreading, and migration by hydrogel substrates with independently tunable mesh size

Hydrogels are widely used as substrates to investigate interactions between cells and their microenvironment as they mimic many attributes of the extracellular matrix. The stiffness of hydrogels is an important property that is known to regulate cell behavior. Beside stiffness, cells also respond to structural cues such as mesh size. However, since the mesh size of hydrogel is intrinsically coupled to its stiffness, its role in regulating cell behavior has never been independently investigated. Here, we report a hydrogel system whose mesh size and stiffness can be independently controlled. Cell behavior, including spreading, migration, and formation of focal adhesions is significantly altered on hydrogels with different mesh sizes but with the same stiffness. At the transcriptional level, hydrogel mesh size affects cellular mechanotransduction by regulating nuclear translocation of yes-associated protein. These findings demonstrate that the mesh size of a hydrogel plays an important role in cell-substrate interactions. Statement of significanceHydrogels are ideal platforms with which to investigate interactions between cells and their microenvironment as they mimic many physical properties of the extracellular matrix. However, the mesh size of hydrogels is intrinsically coupled to their stiffness, making it challenging to investigate the contribution of mesh size to cell behavior. In this work, we use hydrogel-on-glass substrates with defined thicknesses whose stiffness and mesh size can be independently tuned. We use these substrates to isolate the effects of mesh size on cell behavior, including attachment, spreading, migration, focal adhesion formation and YAP localization in the nucleus. Our results show that mesh size has significant, yet often overlooked, effects, on cell behavior, and contribute to a further understanding of cell-substrate interactions.

High-Throughput FRAP Analysis of Solute Diffusion in Hydrogels.

Increasingly accurate mathematical models have been developed to relate solute and hydrogel properties to solute diffusion coefficients in hydrogels, primarily by comparing solute sizes and hydrogel mesh sizes. Here, we use a standardized, high-throughput method for fluorescence recovery after photobleaching (FRAP) experiments and analysis to characterize the diffusion coefficients of fluorescein, three sizes of FITC-dextran, and three sizes of FITC-conjugated poly(ethylene glycol) (PEG) through 18 structurally varied poly(vinyl alcohol) (PVA) hydrogel formulations. Increasing the hydrogel mesh radii increased the diffusivities of all the tested solutes within the hydrogels. While the diffusivity of FITC-dextrans in hydrogels decreased with increasing solute size, the diffusivity of FITC-PEGs increased with increasing solute size, suggesting that a generalized hydrodynamic radius-based model is not universally applicable for solute diffusion in hydrogels. The high-throughput characterization method for solute diffusion in hydrogels described here facilitates precise hydrogel design for biomedical applications.

Microscale Interfacial Polymerization on a Chip

AbstractForming hydrogels with precise geometries is challenging and mostly done using photopolymerization, which involves toxic chemicals, rinsing steps, solvents, and bulky optical equipment. Here, we introduce a new method for in situ formation of hydrogels with a well‐defined geometry in a sealed microfluidic chip by interfacial polymerization. The geometry of the hydrogel is programmed by microfluidic design using capillary pinning structures and bringing into contact solutions containing hydrogel precursors from vicinal channels. The characteristics of the hydrogel (mesh size, molecular weight cut‐off) can be readily adjusted. This method is compatible with capillary‐driven microfluidics, fast, uses small volumes of reagents and samples, and does not require specific laboratory equipment. Our approach creates opportunities for filtration, hydrogel functionalization, and hydrogel‐based assays, as exemplified by a rapid, compact competitive immunoassay that does not require a rinsing step.

Microscale Interfacial Polymerization on a Chip.

Forming hydrogels with precise geometries is challenging and mostly done using photopolymerization, which involves toxic chemicals, rinsing steps, solvents, and bulky optical equipment. Here, we introduce a new method for in situ formation of hydrogels with a well‐defined geometry in a sealed microfluidic chip by interfacial polymerization. The geometry of the hydrogel is programmed by microfluidic design using capillary pinning structures and bringing into contact solutions containing hydrogel precursors from vicinal channels. The characteristics of the hydrogel (mesh size, molecular weight cut‐off) can be readily adjusted. This method is compatible with capillary‐driven microfluidics, fast, uses small volumes of reagents and samples, and does not require specific laboratory equipment. Our approach creates opportunities for filtration, hydrogel functionalization, and hydrogel‐based assays, as exemplified by a rapid, compact competitive immunoassay that does not require a rinsing step.

Prediction of Viscoelastic Properties of Enzymatically Crosslinkable Tyramine-Modified Hyaluronic Acid Solutions Using a Dynamic Monte Carlo Kinetic Approach.

The present study deals with the mathematical modeling of crosslinking kinetics of polymer–phenol conjugates mediated by the Horseradish Peroxidase (HRP)-hydrogen peroxide (H2O2) initiation system. More specifically, a dynamic Monte Carlo (MC) kinetic model is developed to quantify the effects of crosslinking conditions (i.e., polymer concentration, degree of phenol substitution and HRP and H2O2 concentrations) on the gelation onset time; evolution of molecular weight distribution and number and weight average molecular weights of the crosslinkable polymer chains and gel fraction. It is shown that the MC kinetic model can faithfully describe the crosslinking kinetics of a finite sample of crosslinkable polymer chains with time, providing detailed molecular information for the crosslinkable system before and after the gelation point. The MC model is validated using experimental measurements on the crosslinking of a tyramine modified Hyaluronic Acid (HA-Tyr) polymer solution reported in the literature. Based on the rubber elasticity theory and the MC results, the dynamic evolution of hydrogel viscoelastic and molecular properties (i.e., number average molecular weight between crosslinks, Mc, and hydrogel mesh size, ξ) are calculated.

Organisation of clay nanoplatelets in a polyelectrolyte-based hydrogel

We investigate the organisation of clay nanoplatelets within a hydrogel based on modified ionenes, cationic polyelectrolytes forming physically crosslinked hydrogels induced by hydrogen bonding and π-π stacking. Combination of small angle X-ray and neutron scattering (SAXS, SANS) reveals the structure of the polyelectrolyte network as well as the organisation of the clay additives. The clay-free hydrogel network features a characteristic mesh-size between 20 and 30 nm, depending on the polyelectrolyte concentration. Clay nanoplatelets inside the hydrogel organise in a regular face-to-face stacking manner, with a large repeat distance, following rather closely the hydrogel mesh-size. The presence of the nanoplatelets does not modify the hydrogel mesh size. Further, the clay-compensating counterions (Na+, Ca2+ or La3+) and the clay type (montmorillonite, beidellite) both have a significant influence on nanoplatelet organisation. The degree of nanoplatelet ordering in the hydrogel is very sensitive to the negative charge location on the clay platelet (different for each clay type). Increased nanoplatelet ordering leads to an improvement of the elastic properties of the hydrogel. On the contrary, the presence of dense clay aggregates (tactoids), induced by multi-valent clay counterions, destroys the hydrogel network as seen by the reduction of the elastic modulus of the hydrogel.

Selectively Cross-Linked Tetra-PEG Hydrogels Provide Control over Mechanical Strength with Minimal Impact on Diffusivity.

Synthetic hydrogels formed from poly(ethylene glycol) (PEG) are widely used to study how cells interact with their extracellular matrix. These in vivo-like 3D environments provide a basis for tissue engineering and cell therapies but also for research into fundamental biological questions and disease modeling. The physical properties of PEG hydrogels can be modulated to provide mechanical cues to encapsulated cells; however, the impact of changing hydrogel stiffness on the diffusivity of solutes to and from encapsulated cells has received only limited attention. This is particularly true in selectively cross-linked “tetra-PEG” hydrogels, whose design limits network inhomogeneities. Here, we used a combination of theoretical calculations, predictive modeling, and experimental measurements of hydrogel swelling, rheological behavior, and diffusion kinetics to characterize tetra-PEG hydrogels’ permissiveness to the diffusion of molecules of biologically relevant size as we changed polymer concentration, and thus hydrogel mechanical strength. Our models predict that hydrogel mesh size has little effect on the diffusivity of model molecules and instead predicts that diffusion rates are more highly dependent on solute size. Indeed, our model predicts that changes in hydrogel mesh size only begin to have a non-negligible impact on the concentration of a solute that diffuses out of hydrogels for the smallest mesh sizes and largest diffusing solutes. Experimental measurements characterizing the diffusion of fluorescein isothiocyanate (FITC)-labeled dextran molecules of known size aligned well with modeling predictions and suggest that doubling the polymer concentration from 2.5% (w/v) to 5% produces stiffer gels with faster gelling kinetics without affecting the diffusivity of solutes of biologically relevant size but that 10% hydrogels can slow their diffusion. Our findings provide confidence that the stiffness of tetra-PEG hydrogels can be modulated over a physiological range without significantly impacting the transport rates of solutes to and from encapsulated cells.

Synthesis and characterization of site selective photo-crosslinkable glycidyl methacrylate functionalized gelatin-based 3D hydrogel scaffold for liver tissue engineering

The presented work outlined the development of a new biocompatible hydrogel material that has potential applications in soft tissue engineering. As a proof of concept, human hepatocytes were used to demonstrate the suitability of this material in providing conducive environment for cellular growth and functional development. Herein, a detailed synthesis of novel gelatin derivatives - photo-crosslinkable glycidyl methacrylate (GMA) functionalized gelatins (Gelatin-GMA), and preparation of three-dimensional (3D) hydrogel scaffolds for the encapsulated Huh-7.5 cells is reported. The Gelatin-GMA biopolymers were synthesized at two different pH values of 3.5 (acidic) and 10.5 (basic) where two different photo-crosslinkable polymers were formed utilizing –COOH & –OH groups in acidic pH, and –NH2 & –OH groups in basic pH. The hydrogels were prepared using an initiator (Irgacure I2959) in the presence of UV light. The Gelatin-GMA biopolymers were characterized using spectroscopic studies which confirmed the successful preparation of the polymer derivatives. Rheological measurement was carried out to characterize the mechanical properties and derive the mesh sizes of the 3D hydrogels. Subsequently, detailed in vitro hepatocyte compatibility and functionality studies were performed in the 3D cell seeded hydrogel platform. The 3D hydrogel design with larger mesh sizes utilizes the advantage of the excellent diffusion properties of porous platform, and enhanced cell-growth was observed, which in turn elicited favorable Huh-7.5 response. The hydrogels led to improved cellular functions such as differentiation, viability and proliferation. Overall, it showed that the Gelatin-GMA based hydrogels presented better results compared to control sample (GelMA) because of the higher mesh sizes in Gelatin-GMA based hydrogels. Additionally, the functional group studies of the two Gelatin-GMA samples revealed that the cell functionalities are almost unaffected even after the tripeptide - Arg-Gly-Asp (RGD) in Gelatin-GMA synthesized at pH 3.5 is no longer completely available.

Hydrogels for 3D Neural Tissue Models: Understanding Cell-Material Interactions at a Molecular Level.

The development of 3D neural tissue analogs is of great interest to a range of biomedical engineering applications including tissue engineering of neural interfaces, treatment of neurodegenerative diseases and in vitro assessment of cell-material interactions. Despite continued efforts to develop synthetic or biosynthetic hydrogels which promote the development of complex neural networks in 3D, successful long-term 3D approaches have been restricted to the use of biologically derived constructs. In this study a poly (vinyl alcohol) biosynthetic hydrogel functionalized with gelatin and sericin (PVA-SG), was used to understand the interplay between cell-cell communication and cell-material interaction. This was used to probe critical short-term interactions that determine the success or failure of neural network growth and ultimately the development of a useful model. Complex primary ventral mesencephalic (VM) neural cells were encapsulated in PVA-SG hydrogels and critical molecular cues that demonstrate mechanosensory interaction were examined. Neuronal presence was constant over the 10 day culture, but the astrocyte population decreased in number. The lack of astrocytic support led to a reduction in neural process outgrowth from 24.0 ± 1.3 μm on Day 7 to 7.0 ± 0.1 μm on Day 10. Subsequently, purified astrocytes were studied in isolation to understand the reasons behind PVA-SG hydrogel inability to support neural network development. It was proposed that the spatially restrictive nature (or tight mesh size) of PVA-SG hydrogels limited the astrocytic actin polymerization together with a cytoplasmic-nuclear translocation of YAP over time, causing an alteration in their cell cycle. This was confirmed by the evaluation of p27/Kip1 gene that was found to be upregulated by a twofold increase in expression at both Days 7 and 10 compared to Day 3, indicating the quiescent stage of the astrocytes in PVA-SG hydrogel. Cell migration was further studied by the quantification of MMP-2 production that was negligible compared to 2D controls, ranging from 2.7 ± 2.3% on Day 3 to 5.3 ± 2.9% on Day 10. This study demonstrates the importance of understanding astrocyte-material interactions at the molecular level, with the need to address spatial constraints in the 3D hydrogel environment. These findings will inform the design of future hydrogel constructs with greater capacity for remodeling by the cell population to create space for cell migration and neural process extension.

3D Hyaluronic Acid Hydrogels for Modeling Oligodendrocyte Progenitor Cell Behavior as a Function of Matrix Stiffness.

The lack of regenerative solutions for demyelination within the central nervous system motivates the development of strategies to expand and drive the bioactivity of the cells, including oligodendrocyte progenitor cells (OPCs), that ultimately give rise to myelination. In this work, we introduce a 3D hyaluronic acid (HA) hydrogel system to study the effects of microenvironmental mechanical properties on the behavior of OPCs. We tuned the stiffness of the hydrogels to match the brain tissue (storage modulus 200-2000 Pa) and studied the effects of stiffness on metabolic activity, proliferation, and cell morphology of OPCs over a 7 day period. Although hydrogel mesh size decreased with increasing stiffness, all hydrogel groups facilitated OPC proliferation and mitochondrial metabolic activity to similar degrees. However, OPCs in the two lower stiffness hydrogel groups (170 ± 42 and 794 ± 203 Pa) supported greater adenosine triphosphate levels per cell than the highest stiffness hydrogels (2179 ± 127 Pa). Lower stiffness hydrogels also supported higher levels of cell viability and larger cell spheroid formation compared to the highest stiffness hydrogels. Together, these data suggest that 3D HA hydrogels are a useful platform for studying OPC behavior and that OPC growth/metabolic health may be favored in lower stiffness microenvironments mimicking brain tissue mechanics.

Hydrogel Properties May Influence Mesenchymal Stem Cell Lineage Progression Through Modulating GAPDH Activity

Cell traction forces, biochemical signals, and cell metabolism are each known to regulate mesenchymal stem cell (MSC) differentiation. Biomaterials have therefore been designed to manipulate cell traction force (via their viscoelasticity and adhesion ligands) to guide MSC fate decisions. Similarly, the type and density of biochemical signals presented by a material have been tailored to influence MSC differentiation. However, the potential impact of biomaterial properties on regulating MSC differentiation through modulating cell metabolism is relatively unstudied. Here, we present data indicating that hydrogel elastic modulus and mesh size regulate MSC differentiation in part through modulating the activity of the metabolic enzyme glyceraldehyde-3-phosphate-dehydrogenase (GAPDH). Toward this end, we first confirm that the differentiation profile of MSCs cultured in 2D on highly elastic, covalently crosslinked poly(ethylene glycol) diacrylate (PEGDA) hydrogels differs substantially from that of MSCs encapsulated within the same hydrogel formulations, indicating a dependence in 3D on a variable(s) beyond elastic modulus. Further results indicate that the GAPDH activity of MSCs in 3D hydrogels is a function of elastic modulus and mesh size, suggesting that GAPDH activity may be one of these variables. Studies in 2D supported a positive correlation between hydrogel elastic modulus and GAPDH activity. Additionally, inhibition of GAPDH activity on 2D surfaces induced alterations in the profiles of key differentiation markers, indicating that GAPDH activity can impact lineage progression. Cumulatively, these findings suggest that the potential impact of hydrogel properties on cell metabolism should be considered when evaluating biomaterial-driven MSC differentiation. Cell traction forces, biochemical signals, and cell metabolism are each known to regulate mesenchymal stem cell (MSC) differentiation. Biomaterials have therefore been designed to manipulate cell traction force (via their viscoelasticity and adhesion ligands) to guide MSC fate decisions. Similarly, the type and density of biochemical signals presented by a material have been tailored to influence MSC differentiation. However, the impact of biomaterial properties on regulating MSC differentiation through modulating cell metabolism is relatively unstudied. Here, we present data indicating that hydrogel elastic modulus and mesh size regulate MSC differentiation in part through modulating the activity of the metabolic enzyme glyceraldehyde-3-phosphate-dehydrogenase (GAPDH).