Delivery of Immunomodulatory Molecules for Muscle and Nerve Regeneration

• Lindsay Gallagher, Biomedical Engineering, University of Delaware

• Brian Kwee, Biomedical Engineering, University of Delaware

An area of immunoengineering is creating biomaterials that provide controlled delivery of immunomodulatory cytokines to areas of injury. Drugs can be injected via bolus solution, but the drugs have a short presence in vivo due to their relatively short half-life. If the drugs can be protected from degradation with a biomaterial, they will provide prolonged support to the injury. One way that this can be done is via an alginate hydrogel. This hydrogel can be a useful tool in muscle and nerve regeneration. These hydrogels can target regulatory T-cells, immune cells that can cause regeneration by reducing inflammation.  Interleukin-33 (IL-33) is a protein cytokine that recruits regulatory T-cells. Amphiregulin is a protein that causes the activation of those regulatory T-cells. Both of these cytokines can be incorporated into an alginate hydrogel. The hydrogel can then release these cytokines over time at an injury site. One focus area in our lab is optimizing this release. This can be done by manipulating electrostatic interactions between the hydrogel and the drug. We tested these drugs’ release in vitro by taking media samples and measuring the concentration using an enzyme-linked immunosorbent assay (ELISA) kit. We were able to find that our hydrogel was able to deliver a sustained release of amphiregulin over thirty days. For IL-33 we used the addition of laponite (a nanoparticle with a distribution of charges) to provide a sustained release over 30 days. We also collaborated with graduate students in the lab to test these hydrogels in vivo in BALB/c mice with ischemic injury. The injury is induced by using the hindlimb ischemia model. This represents a peripheral artery disease and causes muscle and nerve degeneration. After the surgery, a hydrogel loaded with either Amphiregulin or IL-33 is injected into the calf and thigh of the mice one day after surgery. Then the efficacy of the gel in promoting muscle regeneration is tested throughout the following two weeks. Necrosis is observed in the mouse’s leg and blood perfusion imaging is performed. At the end of the study, muscle function testing and histology of the calf and tibialis anterior muscle are done. We have quantified the extent of regeneration in the histology slides of the muscle. Through histology, we found that amphiregulin decreased the amount of fatty degeneration that occurred in the calf of mice.