Fibrillar Hydrogels Research Articles

Fibrillar Hydrogel Inducing Cell Mechanotransduction for Tissue Engineering.

One of the key strategies for tissue engineering is to design multifunctional bioinks that balance printability with cytocompatibility. Here, we describe fibrillar hydrogels produced by Schiff base formation between B-type gelatin and oxidized sodium alginate, followed by the incorporation of type I collagen, yielding a new gel (MyoColl). The resulting hydrogel exhibits a temperature- and mass-ratio-dependent sol-gel transition, showing variability of hydrogel properties depending on the component ratio. MyoColl composition provides a convenient platform for biofabrication in terms of shear thinning, yielding, Young's modulus, and shape accuracy. Metabolic activity tests and fluorescent microscopy of 2D hydrogel-based mouse C2C12 myoblast cell culture show significant cytocompatibility of the developed carriers. In addition, primary signs of cell mechanotransduction and myofilament formation of 3D printed MyoColl-based cell cultures were detected and described. Due to these promising results, the described hydrogel composition has shown itself as a convenient platform for muscle tissue engineering.

An optical system for cellular mechanostimulation in 3D hydrogels

We introduce a method utilizing single laser-generated cavitation bubbles to stimulate cellular mechanotransduction in dermal fibroblasts embedded within 3D hydrogels. We demonstrate that fibroblasts embedded in either amorphous or fibrillar hydrogels engage in Ca2+ signaling following exposure to an impulsive mechanical stimulus provided by a single 250 µm diameter laser-generated cavitation bubble. We find that the spatial extent of the cellular signaling is larger for cells embedded within a fibrous collagen hydrogel as compared to those embedded within an amorphous polyvinyl alcohol polymer (SLO-PVA) hydrogel. Additionally, for fibroblasts embedded in collagen, we find an increased range of cellular mechanosensitivity for cells that are polarized relative to the radial axis as compared to the circumferential axis. By contrast, fibroblasts embedded within SLO-PVA did not display orientation-dependent mechanosensitivity. Fibroblasts embedded in hydrogels and cultured in calcium-free media did not show cavitation-induced mechanotransduction; implicating calcium signaling based on transmembrane Ca2+ transport. This study demonstrates the utility of single laser-generated cavitation bubbles to provide local non-invasive impulsive mechanical stimuli within 3D hydrogel tissue models with concurrent imaging using optical microscopy. Statement of significanceCurrently, there are limited methods for the non-invasive real-time assessment of cellular sensitivity to mechanical stimuli within 3D tissue scaffolds. We describe an original approach that utilizes a pulsed laser microbeam within a standard laser scanning microscope system to generate single cavitation bubbles to provide impulsive mechanostimulation to cells within 3D fibrillar and amorphous hydrogels. Using this technique, we measure the cellular mechanosensitivity of primary human dermal fibroblasts embedded in amorphous and fibrillar hydrogels, thereby providing a useful method to examine cellular mechanotransduction in 3D biomaterials. Moreover, the implementation of our method within a standard optical microscope makes it suitable for broad adoption by cellular mechanotransduction researchers and opens the possibility of high-throughput evaluation of biomaterials with respect to cellular mechanosignaling.

Abstract A054: Collagen fiber architecture modulates PDAC cell and organoid phenotype

Abstract Introduction Collagen plays a significant role in the development of pancreatic ductal adenocarcinoma (PDAC). Disease progression is mediated by a dense collagenous stroma, which acts as a barrier to drug delivery and immune cell access. Current in vitro models do not well represent the collagenous stroma seen in PDAC. The aim of this study is to develop a fibrillar collagen-based hydrogel system which mimics the tumour microenvironment seen in PDAC and assess cell-mediated collagen remodeling. Methods Metastatic SUIT2-007 pancreatic cancer cells, non-metastatic BxPC3 pancreatic cancer cells, and human pancreatic stellate cells (hPSCs) were encapsulated in bovine type I collagen gels of various concentrations. KPC mouse derived tumour organoids were also encapsulated in bovine type I collagen gels of 2 mg/ml and 6 mg/ml concentrations, basement membrane extract (BME), and a combination of BME and collagen gel. Cell viability and proliferation was assessed using Live/Dead imaging and CellTitre-Glo 3D respectively. Cell mediated collagen remodeling was assessed via monitoring of gel contraction on the mm scale, and confocal reflectance microscopy on the cellular scale. Collagen contraction around cells was assessed by measuring the pixel intensity of collagen fibrils within the vicinity of the cell. Cell phenotype was assessed via immunofluorescent staining and qPCR. Results Collagen concentration alters the hydrogel structure and stiffness, with 3 mg/ml collagen resulting in straighter, shorter and thicker fibrils. Metastatic SUIT-007 cells contracted collagen gels more rapidly than non-metastatic BxPC3 cells, with both achieving similar contraction by day 7. SUIT2-007 cells remodel and align soft 1 mg/ml gels while 3 mg/ml gels proved to be sufficiently stiff to resist remodeling. Cellular collagen association was increased in more elongated SUIT-007 cells than in BxPC3 cells. Organoid proliferation varies with hydrogel collagen concentration. Conclusion Metastatic SUIT2-007 cells were able to remodel collagen more rapidly and to a greater extent than non-metastatic BxPC3 cells. Collagen supports an invasive organoid phenotype. Collagen gels combined with confocal reflectance microscopy provide a platform for the quantitative assessment of cell-matrix interactions in PDAC. Acknowledgements This project was funded by a Medical Traineeship from UCD School of Medicine, and summer student scholarships from Irish Cancer Society and Breakthrough Cancer Research. We acknowledge the Conway Imaging Core Facility for assistance with microscopy. Citation Format: Ciara L Doyle, Shanu Xavier, Jarren Y.S Lu, Leah Fallon, Anwesha Sarkar, Jan J Baguyo, Maela Gars, Stephen D Thorpe. Collagen fiber architecture modulates PDAC cell and organoid phenotype [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Advances in Pancreatic Cancer Research; 2024 Sep 15-18; Boston, MA. Philadelphia (PA): AACR; Cancer Res 2024;84(17 Suppl_2):Abstract nr A054.

Structure-properties relationships of defined CNF single-networks crosslinked by telechelic PEGs

The high structural anisotropy and colloidal stability of cellulose nanofibrils' enable the creation of self-standing fibrillar hydrogel networks at very low solid contents. Adding methacrylate moieties on the surface of TEMPO oxidized CNFs allows the formation of more robust covalently crosslinked networks by free radical polymerization of acrylic monomers, exploiting the mechanical properties of these networks more efficiently. This technique yields strong and elastic networks but with an undefined network structure. In this work, we use acrylate-capped telechelic polymers derived from the step-growth polymerization of PEG diacrylate and dithiothreitol to crosslink methacrylated TEMPO-oxidized cellulose nanofibrils (MATO CNF). This combination resulted in flexible and strong hydrogels, as observed through rheological studies, compression and tensile loading. The structure and mechanical properties of these hydrogel networks were found to depend on the dimensions of the CNFs and polymer crosslinkers. The structure of the networks and the role of individual components were evaluated with SAXS (Small-Angle X-ray Scattering) and photo-rheology. A thorough understanding of hybrid CNF/polymer networks and how to best exploit the capacity of these networks enable further advancement of cellulose-based materials for applications in packaging, soft robotics, and biomedical engineering.

Hierarchical Design of Tissue-Mimetic Fibrillar Hydrogel Scaffolds.

Most tissues of the human body present hierarchical fibrillar extracellular matrices (ECMs) that have a strong influence over their physicochemical properties and biological behavior. Of great interest is the introduction of this fibrillar structure to hydrogels, particularly due to the water-rich composition, cytocompatibility, and tunable properties of this class of biomaterials. Here, the main bottom-up fabrication strategies for the design and production of hierarchical biomimetic fibrillar hydrogels and their most representative applications in the fields of tissue engineering and regenerative medicine are reviewed. For example, the controlled assembly/arrangement of peptides, polymeric micelles, cellulose nanoparticles (NPs), and magnetically responsive nanostructures, among others, into fibrillar hydrogels is discussed, as well as their potential use as fibrillar-like hydrogels (e.g., those from cellulose NPs) with key biofunctionalities such as electrical conductivity or remote stimulation. Finally, the major remaining barriers to the clinical translation of fibrillar hydrogels and potential future directions of research in this field are discussed.

Solidified water at room temperature hosting tailored fluidic channels by using highly anisotropic cellulose nanofibrils

Highly anisotropic cellulose nanofibrils can solidify liquid water, creating self-supporting structures by incorporating a tiny number of fibrils. These fibrillar hydrogels can contain as much as 99.99 wt% water. The structure and mechanical properties of fibrillar networks have so far not been completely understood, nor how they solidify the bulk water at such low particle concentrations. In this work, the mechanical properties of cellulose fibrillar hydrogels in the dilute regime from a wt% perspective have been studied, and an elastoplastic model describing the network structure and its mechanics is presented. A significant insight from this work is that the ability of the fibrils to solidify water is very dependent on particle stiffness and the number of contact points it can form in the network structure. The comparison between the experimental results and the theoretical model shows that the fibrillar networks in the dilute regime form via a non-stochastic process since the fibrils have the time and freedom to find contact points during network formation by translational and rotational diffusion. The formed, dilute fibrillar network deforms by sliding fibril contacts upon straining the network beyond its elastic limit. Our results also show that before macroscopic failure, the fibril contacts are restored once the load is released. The exceptional properties of this solidified water are exploited to host fluidic channels, allowing directed fluid transportation in water. Finally, the microfluidic channels formed in the hydrogels are tailored by the layer-by-layer technique to be interactive against external stimuli, a characteristic envisioned to be useful in biomedical applications.

A Systems Approach to Study Collagen Type I Self-Assembly: Kinetics and Morphology.

Collagen type I, the main component of the extracellular matrix in vertebrates, is widely used in tissue engineering applications. This is on account that collagen molecules can self-assemble under certain conditions into 3D fibrillar hydrogels. Although there is an extensive body of literature studying collagen self-assembly, there is a lack of systematic understanding on how different experimental factors, such as pH and temperature, and their cumulative effects guide the self-assembly process. In this work, a comprehensive workflow to study the interactive effects of several assembly parameters on the collagen self-assembly process is implemented. This workflow consists of: 1) efficient statistical sampling based on Design of Experiments, 2) high-throughput and automated data collection and 3) automated data analysis. This approach enables to screen several parameters simultaneously and derive a set of mathematical equationsthat link parameters with the kinetics and morphological aspects of collagen self-assembly, and can be used to design collagen constructs with predefinedcharacteristics.

Fibrillar Hydrogel Based on Cellulose Nanocrystals Crosslinked via Diels-Alder Reaction: Preparation and pH-Sensitive Release of Benzocaine.

A fibrillar hydrogel was obtained by covalent crosslinking via Diels-Alder reaction of two types of cellulose nanocrystals (CNCs) with furan and maleimide groups. Gelation has been studied at various ratios of components and temperatures in the range from 20 to 60 °C. It was shown that the rheological properties of the hydrogel can be optimized by varying the concentration and ratio of components. Due to the rigid structure of the CNCs, the hydrogel could be formed at a concentration of at least 5 wt%; however, it almost does not swell either in water with pH 5 or 7 or in the HBSS buffer. The introduction of aldehyde groups into the CNCs allows for the conjugation of physiologically active molecules containing primary amino groups due to the formation of imine bonds. Here, we used benzocaine as a model drug for conjugation with CNC hydrogel. The resulting drug-conjugated hydrogel demonstrated the stability of formulation at pH 7 and a pH-sensitive release of benzocaine due to the accelerated hydrolytic cleavage of the imine bond at pH < 7. The developed drug-conjugated hydrogel is promising as wound dressings for local anesthesia.

Strategies for Anisotropic Fibrillar Hydrogels: Design, Cell Alignment, and Applications in Tissue Engineering.

Efficient cellular alignment in biomaterials presents a considerable challenge, demanding the refinement of appropriate material morphologies, while ensuring effective cell-surface interactions. To address this, biomaterials are continuously researched with diverse coatings, hydrogels, and polymeric surfaces. In this context, we investigate the influence of physicochemical parameters on the architecture of fibrillar hydrogels that significantly orient the topography of flexible hydrogel substrates, thereby fostering cellular adhesion and spatial organization. Our Review comprehensively assesses various techniques for aligning polymer fibrils within hydrogels, specifically interventions applied during and after the cross-linking process. These methodologies include mechanical strains, precise temperature modulation, controlled fluidic dynamics, and chemical modulators, as well as the use of magnetic and electric fields. We highlight the intrinsic appeal of these methodologies in fabricating cell-aligning interfaces and discuss their potential implications within the fields of biomaterials and tissue engineering, particularly concerning the pursuit of optimal cellular alignment.

Amyloid Fibril and Clay Nanosheet Dual-Nanoengineered DNA Dynamic Hydrogel for Vascularized Bone Regeneration.

Dynamic hydrogels have attracted enormous interest for bone tissue engineering as they demonstrate reversible mechanics to better mimic biophysical cues of natural extracellular matrix (ECM) compared to traditional static hydrogels. However, the facile development of therapeutic dynamic hydrogels that simultaneously recapitulate the filamentous architecture of the ECM of living tissues and induce both osteogenesis and angiogenesis to augment vascularized bone regeneration remains challenging. Herein, we report a dual nanoengineered DNA dynamic hydrogel developed through the supramolecular coassembly of amyloid fibrils and clay nanosheets with DNA strands. The nanoengineered ECM-like fibrillar hydrogel network is facilely formed without a complicated and tedious molecular synthesis. Amyloid fibrils together with clay nanosheets synergistically enhance the mechanical strength and stability of the dynamic hydrogel and, more remarkably, endow the matrix with an array of tunable features, including shear-thinning, injectability, self-healing, self-supporting, and 3D printable properties. The QK peptide is further chemically grafted onto amyloid fibrils, and its sustainable release from the hydrogel matrix stimulates the tube formation and migration with human umbilical vein endothelial cells. Meanwhile, the nanoengineered hydrogel matrix promotes osteogenic differentiation of bone marrow mesenchymal stem cells due to the sustainable release of Si4+ and Mg2+ derived from clay nanosheets. Furthermore, the manipulation of enhanced vascularized bone regeneration by the dynamic hydrogel is revealed in a rat cranial bone defect model. This dual nanoengineered strategy envisions great promise in developing therapeutic dynamic hydrogels for improved and customizable bone regeneration.

Biomimetic Nanofibrillar Hydrogel with Cell-Adaptable Network for Enhancing Cellular Mechanotransduction, Metabolic Energetics, and Bone Regeneration.

The natural extracellular matrix, with its heterogeneous structure, provides a stable and dynamic biophysical framework and biochemical signals to guide cellular behaviors. It is challenging but highly desirable to develop a synthetic matrix that emulates the heterogeneous fibrous structure with macroscopic stability and microscopical dynamics and contains inductive biochemical signals. Herein, we introduce a peptide fiber-reinforced hydrogel in which the stiff ß-sheet fiber functions as a multivalent cross-linker to enhance the hydrogel's macroscopic stability. The dynamic imine cross-link between the peptide fiber and polymer network endows the hydrogel with a microscopically dynamic network. The obtained fibrillar nanocomposite hydrogel, with its cell-adaptable dynamic network, enhances cell-matrix and cell-cell interactions and therefore significantly promotes the mechanotransduction, metabolic energetics, and osteogenesis of encapsulated stem cells. Furthermore, the hydrogel can codeliver a fiber-attached inductive drug to further enhance osteogenesis and bone regeneration. We believe that our work provides valuable guidance for the design of cell-adaptive and bioactive biomaterials for therapeutic applications.

Molecular Tuning of a Benzene-1,3,5-Tricarboxamide Supramolecular Fibrous Hydrogel Enables Control over Viscoelasticity and Creates Tunable ECM-Mimetic Hydrogels and Bioinks.

Traditional synthetic covalent hydrogels lack the native tissue dynamics and hierarchical fibrous structure found in the extracellular matrix (ECM). These dynamics and fibrous nanostructures are imperative in obtaining the correct cell/material interactions. Consequently, the challenge to engineer functional dynamics in a fibrous hydrogel and recapitulate native ECM properties remains a bottle-neck to biomimetic hydrogel environments. Here,the molecular tuning of a supramolecular benzene-1,3,5-tricarboxamide (BTA) hydrogelator via simple modulation of hydrophobic substituents is reported. This tuning results in fibrous hydrogels with accessible viscoelasticity over 5 orders of magnitude, while maintaining a constant equilibrium storage modulus. BTA hydrogelators are created with systematic variations in the number ofhydrophobic carbon atoms, and this is observed to control the viscoelasticity and stress-relaxation timescales in a logarithmic fashion. Some of theseBTA hydrogels are shear-thinning, self-healing, extrudable, and injectable, and can be 3D printed into multiple layers. These hydrogels show high cell viability for chondrocytes and human mesenchymal stem cells, establishing their use in tissue engineering applications. This simple molecular tuning by changing hydrophobicity (with just a few carbon atoms) provides precise control over the viscoelasticity and 3D printability in fibrillar hydrogels and can be ported onto other 1D self-assembling structures. The molecular control and design of hydrogel network dynamics can push the field of supramolecular chemistry towardthe design of new ECM-mimicking hydrogelators for numerous cell-culture and tissue-engineering applications and give access towardhighly biomimetic bioinks for bioprinting.

Ultrafast, High-Strain, and Strong Uniaxial Hydrogel Actuators from Recyclable Nanofibril Networks.

Synthetic polymeric hydrogels mimic biological tissues and are suitable for future lifelike machines. However, in polymer hydrogel actuators, actuation is isotropic, so they must be crosslinked or placed in a turgor membrane to achieve high actuation pressures, severely impeding their performance. Here, we show that organizing cellulose nanofibrils in anisotropic fibrillar networks leads to hydrogel sheets with a mechanical in-plane reinforcement that generates a uniaxial, out-of-plane strain with performance far surpassing polymer hydrogels. These fibrillar hydrogel actuators expand uniaxially by 250 times with an initial rate of 100-150% s-1 , compared to <10 times and <1% s-1 in directional strain rate for isotropic hydrogels, respectively. The blocking pressure was up to 0.9MPa, similar to turgor actuators, while the time to reach 90% of the max pressure was 1-2 minutes, compared to 10 minutes to hours for crosslinked polymers or turgor actuators. We showcase uniaxial actuators that lift objects 120 000 times their weight and soft grippers that grasp objects. In addition, the hydrogels could be recycled without a loss in performance. The uniaxial swelling allowed adding channels through the gel for local solvent delivery, further increasing the actuation rate and cyclability. Thus, fibrillar networks can overcome the major drawbacks of hydrogel actuators and is a significant advancement toward hydrogel-based lifelike machines. This article is protected by copyright. All rights reserved.

Governing the aggregation of type I collagen mediated through β-cyclodextrin

The effect of carbohydrates on collagen self-assembly behavior has been widely investigated because of their regulation on collagen fibrogenesis in vivo. In this paper, β-cyclodextrin (β-CD) was selected as an external disturbance to explore its intrinsic regulating mechanism on collagen self-assembly. The results of fibrogenesis kinetics indicated that β-CD had a bilateral regulation on collagen self-aggregation process, which was closely related to the content of β-CD: collagen protofibrils with low β-CD content were less aggregated compared to collagen protofibrils with high β-CD content. However, typical periodic stripes of ~67 nm on collagen fibrils were observed from transmission electron microscope (TEM), indicating that β-CD did not disturb the lateral arrangement of collagen molecules to form a 1/4 staggered structure. Correspondingly, the degree of aggregation of collagen self-assembled fibrils was closely correlated with the addition of β-CD content, as confirmed by field emission scanning electron microscopy (FESEM) and atomic force microscope (AFM). In addition, collagen/β-CD fibrillar hydrogel had good thermal stability and cytocompatibility. These results provide a better understanding of how to construct a structurally reliable collagen/β-CD fibrillar hydrogel as a biomedical material in a β-CD-regulated environment.

Ca2+ and Ag+ orient low-molecular weight amphiphile self-assembly into "nano-fishnet" fibrillar hydrogels with unusual β-sheet-like raft domains.

Low-molecular weight gelators (LMWGs) are small molecules (Mw < ∼1 kDa), which form self-assembled fibrillar network (SAFiN) hydrogels in water when triggered by an external stimulus. A great majority of SAFiN gels involve an entangled network of self-assembled fibers, in analogy to a polymer in a good solvent. In some rare cases, a combination of attractive van der Waals and repulsive electrostatic forces drives the formation of bundles with a suprafibrillar hexagonal order. In this work, an unexpected micelle-to-fiber transition is triggered by Ca2+ or Ag+ ions added to a micellar solution of a novel glycolipid surfactant, whereas salt-induced fibrillation is not common for surfactants. The resulting SAFiN, which forms a hydrogel above 0.5 wt%, has a "nano-fishnet" structure, characterized by a fibrous network of both entangled fibers and β-sheet-like rafts, generally observed for silk fibroin, actin hydrogels or mineral imogolite nanotubes, but not known for SAFiNs. The β-sheet-like raft domains are characterized by a combination of cryo-TEM and SAXS and seem to contribute to the stability of glycolipid gels. Furthermore, glycolipid is obtained by fermentation from natural resources (glucose, rapeseed oil), thus showing that naturally engineered compounds can have unprecedented properties, when compared to the wide range of chemically derived amphiphiles.

Elastoplastic behavior of anisotropic, physically crosslinked hydrogel networks comprising stiff, charged fibrils in an electrolyte.

Fibrillar hydrogels are remarkably stiff, low-density networks that can hold vast amounts of water. These hydrogels can easily be made anisotropic by orienting the fibrils using different methods. Unlike the detailed and established descriptions of polymer gels, there is no coherent theoretical framework describing the elastoplastic behavior of fibrillar gels, especially concerning anisotropy. In this work, the swelling pressures of anisotropic fibrillar hydrogels made from cellulose nanofibrils were measured in the direction perpendicular to the fibril alignment. This experimental data was used to develop a model comprising three mechanical elements representing the network and the osmotic pressure due to non-ionic and ionic surface groups on the fibrils. At low solidity, the stiffness of the hydrogels was dominated by the ionic swelling pressure governed by the osmotic ingress of water. Fibrils with different functionality show the influence of aspect ratio, chemical functionality, and the remaining amount of hemicelluloses. This general model describes physically crosslinked hydrogels comprising fibrils with high flexural rigidity - that is, with a persistence length larger than the mesh size. The experimental technique is a framework to study and understand the importance of fibrillar networks for the evolution of multicellular organisms, like plants, and the influence of different components in plant cell walls.

The effect of ethanol on fibrillar hydrogels formed by glycyrrhizic acid monoammonium salt

MotivationThe monoammonium salt of glycyrrhizic acid (AGA) is known to form fibrillar hydrogels and few studies regarding self-assembly of AGA have been published. Yet, the understanding of the fibrillar microstructures and the gelation remains vague. Thus, we attempt to achieve a deeper understanding of the microstructures and the gelation process of binary solutions of AGA in water. Further, we examine the effect of ethanol on the microstructures to pave the way for potential enhancement of drug loading in AGA hydrogels. ExperimentsA partial room temperature phase map of the ternary system AGA/ethanol/water was recorded. Small-angle X-ray and neutron scattering experiments were performed over wide ranges of compositions in both binary AGA/water and ternary AGA/ethanol/water mixtures to get access to the micro-structuring. FindingsBinary aqueous solutions of AGA form birefringent gels consisting of a network of long helical fibrils. ‘Infinitely’ long negatively charged fibrils are in equilibrium with shorter fibrils (≈25 nm), both of which have a diameter of about 3 nm and are made of around 30 stacks of AGA per helical period (≈9nm), with each stack consisting of two AGA molecules. The interaxial distance (order of magnitude ≈20 nm) varies with an almost two-dimensional swelling law. Addition of ethanol reduces electrostatic repulsion and favors the formation of fibrillar end caps, reducing the average length of shorter fibrils, as well as the formation of small, swollen aggregates. While the gel network built by the long fibrils is resilient to a significant amount of ethanol, all fibrils are finally dissolved into small aggregates above a certain threshold concentration of ethanol (≈30 wt%).

Printing Structurally Anisotropic Biocompatible Fibrillar Hydrogel for Guided Cell Alignment.

Many fibrous biological tissues exhibit structural anisotropy due to the alignment of fibers in the extracellular matrix. To study the impact of such anisotropy on cell proliferation, orientation, and mobility, it is important to recapitulate and achieve control over the structure of man-made hydrogel scaffolds for cell culture. Here, we report a chemically crosslinked fibrous hydrogel due to the reaction between aldehyde-modified cellulose nanofibers and gelatin. We explored two ways to induce structural anisotropy in this gel by extruding the hydrogel precursor through two different printheads. The cellulose nanofibers in the hydrogel ink underwent shear-induced alignment during extrusion and retained it in the chemically crosslinked hydrogel. The degree of anisotropy was controlled by the ink composition and extrusion flow rate. The structural anisotropy of the hydrogel extruded through a nozzle affected the orientation of human dermal fibroblasts that were either seeded on the hydrogel surface or encapsulated in the extruded hydrogel. The reported straightforward approach to constructing fibrillar hydrogel scaffolds with structural anisotropy can be used in studies of the biological impact of tissue anisotropy.

Exploiting Minimalistic Backbone Engineered γ-Phenylalanine for the Formation of Supramolecular Co-Polymer.

Ordered supramolecular hydrogels assembled by modified aromatic amino acids often exhibit low mechanical rigidity. Aiming to stabilize the hydrogel and understand the impact of conformational freedom and hydrophobicity on the self-assembly process, two building blocks based on 9-fluorenyl-methoxycarbonyl-phenylalanine (Fmoc-Phe) gelator which contain two extra methylene units in the backbone, generating Fmoc-γPhe and Fmoc-(3-hydroxy)-γPhe are designed. Fmoc-γPhe spontaneously assembled in aqueous media forming a hydrogel with exceptional mechanical and thermal stability. Moreover, Fmoc-(3-hydroxy)-γPhe, with an extra backbone hydroxyl group decreasing its hydrophobicity while maintaining some molecular flexibility, self-assembled into a transient fibrillar hydrogel, that later formed microcrystalline aggregates through a phase transition. Molecular dynamics simulations and single crystal X-ray analyses reveal the mechanism underlying the two residues' distinct self-assembly behaviors. Finally, Fmoc-γPhe and Fmoc-(3-OH)-γPhe co-assembly to form a supramolecular hydrogel with notable mechanical properties are demonstrated. It has been believed that the understanding of the structure-assembly relationship will enable the design of new functional amino acid-based hydrogels.

Stress-controlled shear flow alignment of collagen type I hydrogel systems

Disease research and drug screening platforms require in vitro model systems with cellular cues resembling those of natural tissues. Fibrillar alignment, occurring naturally in extracellular matrices, is one of the crucial attributes in tissue development. Obtaining fiber alignment in 3D, in vitro remains an important challenge due to non-linear material characteristics. Here, we report a cell-compatible, shear stress-based method allowing to obtain 3D homogeneously aligned fibrillar collagen hydrogels. Controlling the shear-stress during gelation results in low strain rates, with negligible effects on the viability of embedded SH-SY5Y cells. Our approach offers reproducibility and tunability through a paradigm shift: The shear-stress initiation moment, being the critical optimization parameter in the process, is related to the modulus of the developing gel, whereas state of the art methods often rely on a predefined time to initiate the alignment procedure. After curing, the induced 3D alignment is maintained after the release of stress, with a linear relation between the total acquired strain and the fiber alignment. This method is generally applicable to 3D fibrillar materials and stress/pressure-controlled setups, making it a valuable addition to the fast-growing field of tissue engineering. Statement of significanceControlling fiber alignment in vitro 3D hydrogels is crucial for developing physiologically relevant model systems. However, it remains challenging due to the non-linear material characteristics of fibrillar hydrogels, limiting the scalability and repeatability.Our approach tackles these challenges by utilizing a stress-controlled rheometer allowing us to monitor structural changes in situ and determine the optimal moment for applying a shear-stress inducing alignment. By careful parameter control, we infer the relationship between time, induced strain, alignment and biocompatibility.This tunable and reproducible method is both scalable and generally applicable to any fibrillar hydrogel, therefore, we believe it is useful for research investigating the link between matrix anisotropy and cell behavior in 3D systems, organ-on-chip technologies and drug research.