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.

Multifunctional 3D-Printed Wound Dressings.

Personalized wound dressings provide enhanced healing for different wound types; however multicomponent wound dressings with discretely controllable delivery of different biologically active agents are yet to be developed. Here we report 3D-printed multicomponent biocomposite hydrogel wound dressings that have been selectively loaded with small molecules, metal nanoparticles, and proteins for independently controlled release at the wound site. Hydrogel wound dressings carrying antibacterial silver nanoparticles and vascular endothelial growth factor with predetermined release profiles were utilized to study the physiological response of the wound in a mouse model. Compared to controls, the application of dressings resulted in improvement in granulation tissue formation and differential levels of vascular density, dependent on the release profile of the growth factor. Our study demonstrates the versatility of the 3D-printed hydrogel dressings that can yield varied physiological responses in vivo and can further be adapted for personalized treatment of various wound types.

Nanofibrillar Hydrogel Recapitulates Changes Occurring in the Fibrotic Extracellular Matrix.

Fibrosis is a pathological condition that leads to excessive deposition of collagen and increased tissue stiffness. Understanding the mechanobiology of fibrotic tissue necessitates the development of effective in vitro models that recapitulate its properties and structure; however, hydrogels that are currently used for this purpose fail to mimic the filamentous structure and mechanical properties of the fibrotic extracellular matrix (ECM). Here, we report a nanofibrillar hydrogel composed of cellulose nanocrystals and gelatin, which addresses this challenge. By altering the composition of the hydrogel, we mimicked the changes in structure, mechanical properties, and chemistry of fibrotic ECM. Furthermore, we decoupled the variations in hydrogel structure, properties, and ligand concentration. We demonstrate that this biocompatible hydrogel supports the three-dimensional culture of cells relevant to fibrotic diseases. This versatile hydrogel can be used for in vitro studies of fibrosis of different tissues, thus enabling the development of novel treatments for fibrotic diseases.

Matrix Stiffness-Regulated Growth of Breast Tumor Spheroids and Their Response to Chemotherapy.

Interactions between tumor cells and the extracellular matrix (ECM) are an important factor contributing to therapy failure in cancer patients. Current in vitro breast cancer spheroid models examining the role of mechanical properties on spheroid response to chemotherapy are limited by the use of two-dimensional cell culture, as well as simultaneous variation in hydrogel matrix stiffness and other properties, e.g., hydrogel composition, pore size, and cell adhesion ligand density. In addition, currently used hydrogel matrices do not replicate the filamentous ECM architecture in a breast tumor microenvironment. Here, we report a collagen-alginate hydrogel with a filamentous architecture and a 20-fold variation in stiffness, achieved independently of other properties, used for the evaluation of estrogen receptor-positive breast cancer spheroid response to doxorubicin. The variation in hydrogel mechanical properties was achieved by altering the degree of cross-linking of alginate molecules. We show that soft hydrogels promote the growth of larger MCF-7 tumor spheroids with a lower fraction of proliferating cells and enhance spheroid resistance to doxorubicin. Notably, the stiffness-dependent chemotherapeutic response of the spheroids was temporally mediated: it became apparent at sufficiently long cell culture times, when the matrix stiffness has influenced the spheroid growth. These findings highlight the significance of decoupling matrix stiffness from other characteristics in studies of chemotherapeutic resistance of tumor spheroids and in development of drug screening platforms.

Nanoparticle-laden droplets of liquid crystals: Interactive morphogenesis and dynamic assembly.

Defects in liquid crystals serve as templates for nanoparticle (NP) organization; however, NP assembly in cholesteric (Ch) liquid crystals is only beginning to emerge. We show interactive morphogenesis of NP assemblies and a Ch liquid crystalline host formed by cellulose nanocrystals (CNCs), in which both the host and the guest experience marked changes in shape and structure as a function of concentration. At low NP loading, Ch-CNC droplets exhibit flat-ellipsoidal packing of Ch pseudolayers, while the NPs form a toroidal ring- or two cone-shaped assemblies at droplet poles. Increase in NP loading triggers reversible droplet transformation to gain a core-shell morphology with an isotropic core and a Ch shell, with NPs partitioning in the core and in disclinations. We show programmable assembly of droplets carrying magnetic NPs. This work offers a strategy for NP organization in Ch liquid crystals, thus broadening the spectrum of architectures of soft nanostructured materials.

Nanocolloidal Hydrogel for Heavy Metal Scavenging.

We report a nanocolloidal hydrogel that combines the advantages of molecular hydrogels and nanoparticle-based scavengers of heavy metal ions. The hydrogel was formed by the chemical cross-linking of cellulose nanocrystals and graphene quantum dots. Over a range of hydrogel compositions, its structure was changed from lamellar to nanofibrillar, thus enabling the control of hydrogel permeability. Using a microfluidic approach, we generated nanocolloidal microgels and explored their scavenging capacity for Hg2+, Cu2+, Ni2+, and Ag+ ions. Due to the large surface area and abundance of ion-coordinating sites on the surface of nanoparticle building blocks, the microgels exhibited a high ion-sequestration capacity. The microgels were recyclable and were used in several ion scavenging cycles. These features, in addition to the sustainable nature of the nanoparticles, make this nanocolloidal hydrogel a promising ion-scavenging material.

Patterning of Structurally Anisotropic Composite Hydrogel Sheets.

Compositional and structural patterns play a crucial role in the function of many biological tissues. In the present work, for nanofibrillar hydrogels formed by chemically cross-linked cellulose nanocrystals (CNC) and gelatin, we report a microextrusion-based 3D printing method to generate structurally anisotropic hydrogel sheets with CNCs aligned in the direction of extrusion. We prepared hydrogels with a uniform composition, as well as hydrogels with two different types of compositional gradients. In the first type of gradient hydrogel, the composition of the sheet varied parallel to the direction of CNC alignment. In the second hydrogel type, the composition of the sheet changed orthogonally to the direction of CNC alignment. The hydrogels exhibited gradients in structure, mechanical properties, and permeability, all governed by the compositional patterns, as well as cytocompatibility. These hydrogels have promising applications for both fundamental research and for tissue engineering and regenerative medicine.

Injectable Shear-Thinning Fluorescent Hydrogel Formed by Cellulose Nanocrystals and Graphene Quantum Dots.

In the search for new building blocks of nanofibrillar hydrogels, cellulose nanocrystals (CNCs) have attracted great interest because of their sustainability, biocompatibility, ease of surface functionalization, and mechanical strength. Making these hydrogels fluorescent extends the range of their applications in tissue engineering, bioimaging, and biosensing. We report the preparation and properties of a multifunctional hydrogel formed by CNCs and graphene quantum dots (GQDs). We show that although CNCs and GQDs are both negatively charged, hydrogen bonding and hydrophobic interactions overcome the electrostatic repulsion between these nanoparticles and yield a physically cross-linked hydrogel with tunable mechanical properties. Owing to their shear-thinning behavior, the CNC-GQD hydrogels were used as an injectable material in 3D printing. The hydrogels were fluorescent and had an anisotropic nanofibrillar structure. The combination of these advantageous properties makes this hybrid hydrogel a promising material and fosters the development of new manufacturing methods such as 3D printing.

Supramolecular Nanofibrillar Thermoreversible Hydrogel for Growth and Release of Cancer Spheroids.

Growth of three-dimensional cancer spheroids (CSs) in man-made hydrogels mimicking natural extracellular matrix is an important and challenging task. Herein, we report on a supramolecular temperature-responsive hydrogel designed for the growth and subsequent release of CSs. A filamentous hydrogel was formed at 37 °C from an aqueous suspension of cellulose nanocrystals surface-functionalized with temperature-responsive polymer molecules. The encapsulation of cells in the hydrogel enabled effective growth of CSs with dimensions determined by the concentration of cellulose nanocrystals in the hydrogel. On demand release of CSs without loss of cell viability and spheroid integrity was achieved upon hydrogel cooling. The tumorigenic properties of the released CSs were examined by encapsulating and re-growing them in fibrin hydrogel. The results in this work can be used in fundamental cancer research and in cancer drug screening.