Chitosan-Artesunate nanoparticles: A dual anti-fibrotic and anti-inflammatory strategy for preventing bleb fibrosis post-glaucoma filtration surgery.

Abstract

Glaucoma filtration surgery (GFS) effectively lowers intraocular pressure in glaucoma patients, but postoperative bleb fibrosis often leads to surgical failure. Artesunate (ART) has demonstrated antifibrotic potential; however, its clinical use is limited by poor solubility and rapid degradation. This study aimed to develop chitosan-ART nanoparticles (CS@ART NPs) to improve ART's therapeutic efficacy in preventing bleb fibrosis. CS@ART NPs were synthesized using an ionic gelation method for chitosan encapsulation. Their characterization, including analyses of morphology, hydrodynamic properties, surface charge, encapsulation efficiency, drug release kinetics, stability, chemical structure, and mucoadhesive interactions, was carried out using various techniques such as TEM, DLS, zeta potential analysis, HPLC, FT-IR, ¹H-NMR, and adhesion assays. The antifibrotic effects were evaluated in a rabbit GFS model through subconjunctival injection. Histological analysis as well as immunohistochemistry for fibrosis markers α-SMA and fibronectin were detected. In vitro studies were conducted using human primary ocular fibroblasts stimulated with TGF-β1 to assess anti-inflammatory and anti-proliferative effects, measured by EdU incorporation, Western blot for signaling pathway components, and cytokine expression. CS@ART NPs exhibited a uniform size distribution (135.73 ± 0.90 nm), stable dispersion, high encapsulation efficiency (86.4%), and sustained drug release. In the GFS model, a single subconjunctival injection of CS@ART significantly reduced collagen deposition, as well as α-SMA and fibronectin expression at the surgical site. In vitro, CS@ART demonstrated superior antifibrotic effects with a significantly lower IC50 for inhibiting fibroblast proliferation compared to ART alone. Mechanically, CS@ART suppressed the Cyclin D1-CDK4/6, TGF-β1/SMAD, and PI3K/Akt signaling pathways. Additionally, CS@ART showed marked anti-inflammatory effects, reducing inflammatory cell infiltration and IL-6 expression. CS@ART NPs play a dual role both alleviate bleb fibrosis and inflammation after GFS as a promising therapeutic strategy for improving surgical outcomes in glaucoma patients.