Immunomodulatory extracellular matrix hydrogel induces tissue regeneration in a model of partial glossectomy.

While oropharyngeal cancer treatment regimens, including surgical resection, irradiation, and chemotherapy, are effective at removing tumors, they lead to muscle atrophy, denervation, and fibrosis, contributing to the pathogenesis of oropharyngeal dysphagia - difficulty swallowing. Current standard of care of rehabilitative tongue strengthening and swallowing exercises is ineffective. Here, we evaluate an alternative approach utilizing an acellular and injectable biomaterial to preserve muscle content and reduce fibrosis of the tongue after injury. Skeletal muscle extracellular matrix (SKM) hydrogel is fabricated from decellularized porcine skeletal muscle tissue. A partial glossectomy injury in the rat is used to induce tongue fibrosis, and SKM hydrogels along with saline controls are injected into the site of scarring two weeks after injury. Tissues are harvested at 3 and 7 days post-injection for gene expression and immunohistochemical analyses, and at 4 weeks post-injection to evaluate histomorphological properties. SKM hydrogel reduces scar formation and improves muscle regeneration at the site of injury compared to saline. SKM additionally modulates the immune response towards an anti-inflammatory phenotype. This study demonstrates the immunomodulatory and tissue-regenerative capacity of an acellular and minimally invasive ECM hydrogel in a rodent model of tongue injury.

Immunomodulatory contribution of mast cells to the regenerative biomaterial microenvironment.

Bioactive immunomodulatory biomaterials have shown promise for influencing the immune response to promote tissue repair and regeneration. Macrophages and T cells have been associated with this response; however, other immune cell types have been traditionally overlooked. In this study, we investigated the role of mast cells in the regulation of the immune response to decellularized biomaterial scaffolds using a subcutaneous implant model. In mast cell-deficient mice, there was dysregulation of the expected M1 to M2 macrophage transition typically induced by the biomaterial scaffold. Polarization progression deviated in a sex-specific manner with an early transition to an M2 profile in female mice, while the male response was unable to properly transition past a pro-inflammatory M1 state. Both were reversed with adoptive mast cell transfer. Further investigation of the later-stage immune response in male mice determined a greater sustained pro-inflammatory gene expression profile, including the IL-1 cytokine family, IL-6, alarmins, and chemokines. These results highlight mast cells as another important cell type that influences the immune response to pro-regenerative biomaterials.

Dose optimization of decellularized skeletal muscle extracellular matrix hydrogels for improving perfusion and subsequent validation in an aged hindlimb ischemia model.

Peripheral artery disease (PAD) affects more than 27 million individuals in North America and Europe, and current treatment strategies mainly aim to restore blood perfusion. However, many patients are ineligible for existing procedures, and these therapies are often ineffective. Previous studies have demonstrated success of an injectable decellularized skeletal muscle extracellular matrix (ECM) hydrogel in a young rat hindlimb ischemia model of PAD, but further pre-clinical studies are necessary prior to clinical translation. In this study, varying concentrations of a skeletal muscle ECM hydrogel were investigated for material properties and in vivo effects on restoring blood perfusion. Rheological measurements indicated an increase in viscosity and mechanical strength with the higher concentrations of the ECM hydrogels. When injecting dye-labelled ECM hydrogels into a healthy rat, differences were also observed for the spreading and degradation rate of the various concentrations. The three concentrations for the ECM hydrogel were then further examined in a young rat hindlimb ischemia model. The efficacy of the optimal ECM hydrogel concentration was then further confirmed in an aged mouse hindlimb ischemia model. These results further validate the use of decellularized skeletal muscle ECM hydrogels for improving blood perfusion in small animal models of PAD.

Manufacturing considerations for producing and assessing decellularized extracellular matrix hydrogels.

Although several decellularized extracellular matrix (ECM) sheets or patches have been commercialized for use in the clinic, only one injectable decellularized ECM hydrogel, a decellularized myocardial matrix, has reached clinical trials. Consequently, very little information is available for established manufacturing standards or assessments of these materials. Here we present detailed methodology for investigating three parameters related to manufacturing optimization for a porcine derived skeletal muscle ECM hydrogel - animal-to-animal variability, bioburden reduction, and harvesting conditions. Results from characterization assays, including residual dsDNA content and sulfated glycosaminoglycan content, did not yield noteworthy differences amongst individual animals or following the addition of a bioburden reducing agent. However, the tissue collected under different harvesting conditions contained varying amounts of fat, and the protein compositions of the decellularized products differed, which could ultimately impact subsequent efficacy in vitro or in vivo. As decellularized ECM hydrogels continue to be evaluated for various applications, the differences between laboratory-scale and manufacturing-scale material batches should be thoroughly considered to avoid costly and timely optimization during scale-up.