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

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)-alpha methacrylate (PEG alpha MA) backbone. Following modification and character-ization, 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 alpha-methacrylates increased crosslinking density and hybrid-hydrogel elastic modulus to mimic a fibrotic microenvironment. The incorporation of dECM into the PEG alpha MA 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 alpha-smooth muscle actin (alpha SMA). Fibroblasts expressing alpha SMA 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.

Automated summary

Tissue fibrosis is a serious health condition, and there is a need for model systems that can better mimic the changes in the fibrotic extracellular microenvironment. In this study, human decellularized extracellular matrix (dECM) was chemically modified and incorporated into a tunable hybrid-hydrogel system to recapitulate the mechanical properties of healthy tissue types and mimic a fibrotic microenvironment.