Biology subjects

Caliari, S. R.

Publications and source records attributed to Caliari, S. R..

23 records · Page 2Linked to original sources

bioRxiv2021

Guest-host supramolecular assembly of injectable hydrogel fibers for cell encapsulation

The fibrous architecture of the extracellular matrix (ECM) is recognized as an integral regulator of cell function. However, there is an unmet need to develop mechanically robust biomaterials mimicking nanofibrous tissue topography that are also injectable to enable minimally invasive delivery. In this study we have developed a fibrous hydrogel composed of supramolecularly-assembled hyaluronic acid (HA) nanofibers that exhibits mechanical integrity, shear-thinning, rapid self-healing, and cytocompatibility. HA was modified with methacrylates to permit fiber photocrosslinking following electrospinning and either guest adamantane or host {beta}-cyclodextrin groups to guide supramolecular fibrous hydrogel assembly. Analysis of fibrous hydrogel rheological properties showed that the mixed guest-host fibrous hydrogel was more mechanically robust (6.6 {+/-} 2.0 kPa, storage modulus (G')) than unmixed guest hydrogel fibers (1.0 {+/-} 0.1 kPa, G') or host hydrogel fibers (1.1 {+/-} 0.1 kPa, G') separately. The reversible nature of the guest-host supramolecular interactions also allowed for shear-thinning and self-healing behavior as demonstrated by cyclic deformation testing. Human mesenchymal stromal cells (hMSCs) encapsulated in fibrous hydrogels demonstrated satisfactory viability following injection and after seven days of culture (> 85%). Encapsulated hMSCs were more spread and elongated when cultured in viscoelastic guest-host hydrogels compared to non-fibrous elastic controls, with hMSCs also showing significantly decreased circularity in fibrous guest-host hydrogels compared to non-fibrous guest-host hydrogels. Together, these data highlight the potential of this injectable fibrous hydrogel platform for cell and tissue engineering applications requiring minimally invasive delivery.

bioengineering↗

bioRxiv2021

The combined influence of viscoelastic and adhesive cues on fibroblast spreading and focal adhesion formation

Tissue fibrosis is characterized by progressive extracellular matrix (ECM) stiffening and loss of viscoelasticity that ultimately results in reduced organ functionality. Cells bind to the ECM through integrins, where av integrin engagement in particular has been correlated with fibroblast activation into contractile myofibroblasts that drive fibrosis progression. There is a significant unmet need for in vitro hydrogel systems that deconstruct the complexity of native tissues to better understand the individual and combined effects of stiffness, viscoelasticity, and integrin engagement on fibroblast behavior. Here, we developed hyaluronic acid hydrogels with independently tunable cell-instructive properties (stiffness, viscoelasticity, ligand presentation) to address this challenge. Hydrogels with mechanics matching normal or fibrotic lung tissue were synthesized using a combination of covalent crosslinks and supramolecular interactions to tune viscoelasticity. Cell adhesion was mediated through incorporation of either RGD peptide or engineered fibronectin fragments promoting preferential integrin engagement via v{beta}3 or 5{beta}1. We showed that preferential v{beta}3 engagement enabled human lung fibroblasts to assume a myofibroblast-like phenotype on fibrosis-mimicking stiff elastic hydrogels with increased spreading, actin stress fiber organization, and focal adhesion maturation as indicated by paxillin organization. In contrast, preferential 5{beta}1 binding suppressed these metrics. Viscoelasticity, mimicking the mechanics of healthy tissue, largely curtailed fibroblast spreading and focal adhesion organization independent of adhesive ligand type, highlighting its role in preventing fibroblast activation. Together these results provide new insights into how mechanical and adhesive cues collectively guide disease-relevant cell behaviors.

bioengineering↗

bioRxiv2020

Aligned and Conductive 3D Collagen Scaffolds for Skeletal Muscle Tissue Engineering

Skeletal muscle is characterized by its three-dimensional (3D) anisotropic architecture composed of highly aligned, organized, and electrically excitable muscle fibers that enable normal locomotion. Biomaterial-based tissue engineering approaches to repair skeletal muscle injuries are limited due to difficulties in combining 3D structural alignment (to guide cell/matrix organization) and electrical conductivity (to enable electrically excitable myotube assembly and maturation). In this work we successfully produced aligned and electrically conductive 3D collagen scaffolds using a freeze-drying approach. Conductive polypyrrole (PPy) microparticles were synthesized and directly mixed into a suspension of type I collagen and chondroitin sulfate followed by directional lyophilization. Scanning electron microscopy (SEM), energy-dispersive spectroscopy (EDS), and confocal microscopy analyses showed that directional solidification resulted in scaffolds with longitudinally aligned macropores (transverse plane: 155 {+/-} 27 {micro}m, longitudinal plane: 218 {+/-} 49 {micro}m) with homogeneously-distributed PPy content. Chronopotentiometry verified that PPy incorporation resulted in a five-fold increase in conductivity when compared to non-PPy containing collagen scaffolds without detrimentally affecting C2C12 mouse myoblast metabolic activity. Furthermore, the aligned scaffold microstructure provided contact guidance cues that directed myoblast growth and organization. Incorporation of PPy also promoted enhanced myotube formation and maturation as measured by myosin heavy chain (MHC) expression and number of nuclei per myotube. Together these data suggest that aligned and conductive 3D collagen scaffolds could be useful for skeletal muscle tissue engineering.

bioengineering↗

bioRxiv2020

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 (CNS) motivates the need for better understanding of the oligodendrocytes that give rise to myelination. In this work, we introduce a 3D hyaluronic acid (HA) hydrogel system to study the effects of mechanical properties on the behavior of oligodendrocyte progenitor cells (OPCs), the cells that differentiate into myelin-producing oligodendrocytes in the CNS. We tuned the stiffness of the hydrogels to match 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 (169.8 {+/-} 42.1 Pa and 793.9 {+/-} 203.3 Pa) supported greater adenosine triphosphate (ATP) levels per cell than the highest stiffness hydrogels (2178.7 {+/-} 127.2 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.

bioengineering↗

bioRxiv2019

Spatial control of viscoelasticity in phototunable hyaluronic acid hydrogels

Viscoelasticity has emerged as a critical regulator of cell behavior. However, there is an unmet need to develop biomaterials where viscoelasticity can be spatiotemporally tuned to mimic the dynamic and heterogeneous nature of tissue microenvironments. Toward this objective, we developed a modular hyaluronic acid hydrogel system combining light-mediated covalent and supramolecular crosslinking to afford spatiotemporal control of network viscoelastic properties. Covalently crosslinked elastic hydrogels or viscoelastic hydrogels combining covalent and supramolecular interactions were fabricated to match healthy and fibrotic liver stiffness. LX-2 human hepatic stellate cells cultured on viscoelastic substrates displayed reduced spreading, less actin stress fiber organization, and lower myocardin-related transcription factor A (MRTF-A) nuclear localization compared to cells on elastic hydrogels. We further demonstrated the dynamic capabilities of our hydrogel system through photomediated secondary incorporation of either covalent or supramolecular crosslinks to modulate viscoelastic properties. We used photopatterning to create hydrogel models with well-controlled patterned regions of stiff elastic mechanics representing fibrotic tissue nodules surrounded by regions of soft viscoelastic hydrogel mimicking healthy tissue. Cells responded to the local mechanics of the patterned substrate with increased spreading in fibrosis-mimicking regions. Together, this work represents an important step forward toward the creation of hydrogel models with spatiotemporal control of both stiffness and viscoelastic cell-instructive cues.

bioengineering↗