Anti-fibrotic Effects of Atorvastatin Delivered via Keratin-based Hydrogels

Abstract

Implantations of degradable biomaterials for drug delivery or restoration and regenerative medicine cause fibrosis and acute inflammation which may lead to chronic side effects. Additionally, the fibrous encapsulation can itself inhibit the local cells from proliferating and forming new healthy tissue. Current drug delivery models require a new method to decrease the adverse effects following implantation of biomaterials in patients and inhibit the activation signaling sent to quiescent fibroblasts. This research details the novel chemical and physical properties of reduced keratin (KTN) hydrogels extracted from residual human hair with sheared cuticle layers, obtained from barbershops and beauty salons. KTN hydrogels with and without calcium ions conjugated with atorvastatin (Ator), which is thought to affect TGF-{beta} signaling pathways directly by competitively inhibiting key signal transmitters and cellular responses, were investigated via electron microscopy, absorbance spectroscopy, and computational modeling and simulation to observe the proliferative inhibition of fibroblastic cells in comparison to the alginate hydrogel controls. Hydrogel extracts were tested for non-cytotoxic effects via L929 adipose fibroblast cell culture following ISO 10993-5 standard for safety of medical device biomaterials. Two cell models were exposed to increasing serial concentrations of Ator to evaluate the half maximal effective concentration (EC50). For quiescent fibroblasts, Ators EC50 was observed at 368 M in PBS. For mesenchymal stem cells (MSCs), Ators EC50 was observed at 209 M in PBS. The mass of drug was normalized in all groups after calculating their timed absorption and release, and KTN hydrogels conjugated with calcium ions and Ator (ATCK) completely absorbed the drug < 0.5 h while KTN hydrogels with Ator alone (ATKR) completely absorbed the drug at 1h. After 7d in PBS, ATCK released Ator at 2.2 {+/-} 0.7% while Ator in calcium alginate spheres (ATLG) was released at 7.8 {+/-} 1.1% and ATKR released 11.2 {+/-} 4.3% of drug. On-going research focuses on other active fibroblastic cell types and in vivo experimentation to observe foreign body response in subcutaneous mouse tissue.