Biomimetic gradient hydrogel with fibroblast spheroids for full-thickness skin regeneration.

Hydrogel-based scaffolds have been widely investigated for their use in tissue engineering to accelerate tissue regeneration. However, replicating the physiological microenvironments of tissues with appropriate biological cues remains challenging. Recent advances in gradient hydrogels have transformed tissue-engineering research by providing precise structures that mimic the extracellular matrix of natural tissues. Unlike conventional homogeneously structured hydrogels, gradient hydrogels provide a better bio-mimicking microenvironment for combined cell therapies in chronic wound treatment by regulating various cell behaviors, such as proliferation, migration, and differentiation. Here, we present the integration of L929 mouse fibroblast spheroids into gradient hydrogels to mimic the dermal stiffness microenvironment and we investigated their impact on full-thickness skin regeneration. A stiffness gradient was achieved by modulating the concentration of methacrylated hyaluronic acid (HA-MA) with varying degrees of methacrylation, using a dual-syringe pump system. The encapsulation of L929 spheroids with gradient hydrogel facilitated skin cell organization in a hierarchically ordered configuration, leading to full-thickness wound healing that was 1.53 times faster than the untreated group in a rat model. This study provides a method for investigating the potential role of gradient hydrogels in various tissue engineering and regeneration applications.