Chitosan-Coated Oleuropein Nanoliposomes Enable Dose-Sparing Suppression of Serum Inflammatory Markers in a TNF-α-Induced Vascular Inflammation Model

Abstract

Vascular inflammation, marked by endothelial dysfunction and overexpression of proinflammatory cytokines such as interleukin-6 (IL-6) and monocyte chemoattractant protein-1 (MCP-1), is a key contributor to atherosclerosis. Oleuropein (Ole), a phenolic compound with proven anti-inflammatory properties, suffers from poor aqueous stability and limited oral bioavailability, constraining its clinical application. To overcome these limitations, we developed and optimized chitosan (CS)-coated liposomes for oral delivery of Ole. Using a Box-Behnken design, the optimized formulation exhibited a particle size of 168.73 ± 3.86 nm, a polydispersity index (PDI) of 0.17 ± 0.01, a moderate negative zeta potential (−22.50 ± 2.39 mV), an encapsulation efficiency of 87.17 ± 0.27%, and a loading capacity of 12.22 ± 0.25%. The CS-coated formulation retained acceptable colloidal stability under refrigerated storage, enabled pH-responsive sustained release governed by Fickian diffusion, and enhanced endothelial nanocarrier uptake in EA.hy926 cells. The formulation was well tolerated in vitro, with cell viability exceeding 90% across a 500–4000 ng/mL concentration range. In a TNF-α-induced murine model of acute vascular inflammation, daily oral administration of CS-Ole-Lip (20 mg/kg) significantly suppressed serum IL-6 by 6.90-fold (p < 0.05) and MCP-1 by 2.45-fold relative to TNF-α controls (p < 0.0001), with numerically greater suppression than that observed with free Ole and uncoated Ole-Lip at the same dose. CS-Ole-Lip at 2 mg/kg, one-tenth of the 20 mg/kg dose evaluated in this study, produced substantial serum cytokine suppression, suggesting a potential dose-sparing effect. No treatment-associated elevations in the evaluated serum markers of hepatic or renal function or overt histological abnormalities in the examined organs were observed. These findings support CS-Ole-Lip as a stable oral nanocarrier with enhanced anti-inflammatory pharmacodynamic activity. The mechanistic basis for the dose-sparing effect requires direct pharmacokinetic and molecular validation.