Covalently Adaptable Elastin-like Protein – Hyaluronic Acid (ELP – HA) Hydrogels with Secondary Thermo-Responsive Crosslinking for Mesenchymal Stem Cell Delivery

Shear-thinning, self-healing hydrogels are promising vehicles for therapeutic cargo delivery due to their ability to be injected using minimally invasive surgical procedures. We present an injectable hydrogel using a novel combination of dynamic covalent crosslinking with thermo-responsive engineered proteins. Ex situ at room temperature, rapid gelation occurs through the formation of dynamic covalent hydrazone bonds by simply mixing two components: hydrazine-modified elastin-like protein (ELP-HYD) and aldehyde-modified hyaluronic acid (HA-ALD). This shear-thinning network provides significant mechanical protection to encapsulated human mesenchymal stem cells (MSCs) at multiple injection rates during syringe needle injection and rapidly recovers after ejection to retain the cells homogeneously within a 3D environment. In situ at physiological temperature, ELP undergoes a thermal phase transition within the first network, confirmed by the observation of ELP-rich thermal aggregates using Coherent anti-Stokes Raman scattering (CARS) microscopy. The formation of the secondary network further reinforces and stiffens the network. During in vitro culture post ejection, MSCs are observed to maintain their ability to differentiate into multiple lineages, including chondrogenic, adipogenic, and osteogenic cell types. Together, these data demonstrate the promising potential of these ELP-HA hydrogels for injectable stem cell transplantation and tissue regeneration applications.