Chemical Modification of Human Decellularized Extracellular Matrix for Incorporation into Phototunable Hybrid-Hydrogel Models of Tissue Fibrosis

Abstract

Tissue fibrosis remains a serious health condition with high morbidity and mortality rates. There is a critical need to engineer model systems that better recapitulate the spatial and temporal changes in the fibrotic extracellular microenvironment and enable study of the cellular and molecular alterations that occur during pathogenesis. Here, we present a process for chemically modifying human decellularized extracellular matrix (dECM) and incorporating it into a dynamically tunable hybrid-hydrogel system containing a poly(ethylene glycol)-α methacrylate (PEGαMA) backbone. Following modification and characterization, an off-stoichiometry thiol-ene Michael addition reaction resulted in hybrid-hydrogels with mechanical properties that could be tuned to recapitulate many healthy tissue types. Next, photoinitiated, free-radical homopolymerization of excess α-methacrylates increased crosslinking density and hybrid-hydrogel elastic modulus to mimic a fibrotic microenvironment. The incorporation of dECM into the PEGαMA hydrogel decreased the elastic modulus and, relative to fully synthetic hydrogels, increased the swelling ratio, the average molecular weight between crosslinks, and the mesh size of hybrid-hydrogel networks. These changes were proportional to the amount of dECM incorporated into the network. Dynamic stiffening increased the elastic modulus and decreased the swelling ratio, average molecular weight between crosslinks, and the mesh size of hybrid-hydrogels, as expected. Stiffening also activated human fibroblasts, as measured by increases in average cellular aspect ratio (1.59 ± 0.02 to 2.98 ± 0.20) and expression of α-smooth muscle actin (αSMA). Fibroblasts expressing αSMA increased from 25.8 to 49.1% upon dynamic stiffening, demonstrating that hybrid-hydrogels containing human dECM support investigation of dynamic mechanosensing. These results improve our understanding of the biomolecular networks formed within hybrid-hydrogels: this fully human phototunable hybrid-hydrogel system will enable researchers to control and decouple the biochemical changes that occur during fibrotic pathogenesis from the resulting increases in stiffness to study the dynamic cell–matrix interactions that perpetuate fibrotic diseases.

Physically and Chemically Cross-Linked Poly(vinyl alcohol)/Humic Acid Hydrogels for Agricultural Applications

“…The mesh size of the GA12 sample (5.06 nm) was higher than that calculated for the GA9 network (3.98 nm); however, without adding HA, the mesh size of the PVA network was the highest (5.67 nm). The higher values of mesh sizes in the GA0 and GA12 samples compared to that of GA9 hydrogels may favor the loading of high molecular weight compounds within their network structures …”

“…The higher values of mesh sizes in the GA0 and GA12 samples compared to that of GA9 hydrogels may favor the loading of high molecular weight compounds within their network structures. 47 The theoretical shear modulus of GA hydrogels was determined to compare the relative behavior of samples with the experimental results obtained in compression mode. The theoretical shear modulus increased for the samples in the order: GA12, GA0, and GA9.…”

“…The differences between theorical estimations and experimental results may be attributed to a nonideal cross-linking behavior. 47 Swelling Measurements. The swelling profiles were obtained for each hydrogel at room temperature in deionized water and a soil extract.…”

Thiolation-Based Protein-Protein Hydrogels for Improved Wound Healing

“…To overcome this obstacle, we employed Traut’s reagent to thiolate lysine residues in BSA. Traut’s reagent is well-known for its capacity to add thiol groups to the primary amines (-NH2) of lysine, thereby preserving charge properties akin to the original amino group[14, 17, 28, 29]. The thiolated BSA, functioning as a globular protein scaffold, was crosslinked with a genetically engineered collagen-like protein (CLP).…”

Spatial and Temporal Control of 3D Hydrogel Viscoelasticity through Phototuning

“…Tissue mechanical properties vary throughout time during many biological processes, such as development and aging, and diseases such as organ fibrosis and solid tumor progression. − Similarly, tissues are spatially heterogeneous in terms of mechanical properties. , Dynamically tunable hydrogels have been developed to mimic the spatial and temporal changes that occur in tissues. − These approaches primarily rely on methods to alter hydrogel cross-link density in a temporal or spatial gradient or with a user-directed stimulus like light, heat, or ultrasound. Cross-link density is directly related to the hydrogel network elasticity; thus, the stiffness of these gels can be altered spatiotemporally.…”