Functional silk sericin-calcium loaded hydrogels: advancing towards human skin equivalents

Silk sericin (SS), is a protein traditionally discarded during industrial silk processing, contaminating waste waters, with negative economic and ecological impact to the environment. In recent years there has been a growing interest in the recovery and utilization of SS due to its interesting biological properties. SS-based biomaterial platforms, such as hydrogels, are capable of cell incorporation and maintenance over time, acting as a nutritive natural-based environment for cell proliferation1. This opens new avenues to develop more reliable and reproducible in vitro models for a better understanding of human skin conditions while minimizing animal studies. Our team has previously developed an enzymatic crosslinked SS hydrogel using horseradish peroxidase (HRP), to be applied in situ for wound healing. This hydrogel promoted cell viability and complete skin regeneration after 21 days when applied in a diabetic wound model2. These promising results have motivated the use of this formulation as a platform for cell encapsulation, in an approach to develop a natural-based human skin equivalent (HSE). The incorporation of nanoparticles (NPs) within hydrogels is reported to further enhance the biological behavior of encapsulated cells3. In this context, calcium plays an important role in maintaining skin homeostasis and modulating cell proliferation and differentiation4. In a recent study, we explored hydroxyapatite (HAp) and HAp/SS NPs as materials to enhance the adhesion and proliferation of human dermal fibroblasts (HDFs), validating the use of this particulate system to support cell growth. The NPs were produced using a continuous manufacturing process in a new modular oscillatory flow plate reactor (MOFPR). The reaction system enables the production of tailored and homogeneous NPs. In the present work, HDFs and HaCaT were incorporated within a SS/HRP hydrogel to construct a HSE. The system was further optimized with the addition of NPs to the system: a screening was conducted using different HAp and HAp/SS NP concentrations. Our results show that the HAp/SS particles at a low concentration, were associated with the best biological performance (0.05 mg/mL). The co-culture SS system was assembled with a stable silk-fibroin (SF) porous scaffold embedded with human adipose tissue with the addition of neural cells (hiNSCs), as reported by Vidal et al.5 to develop a full-thickness HSE (Figure 1). The sustained viability of the cells in the model over 21 days suggests the formation of a stable and reproducible model representing well some of the characteristics and functionality of native skin (Figure 2).