Boswellic Acid-Nanoparticles (BA-NPs) Confer Synergistic Hepatoprotection against Paracetamol-Induced Toxicity via Attenuation of Oxidative Stress, Inflammation, and Apoptosis

Abstract

Introduction: Paracetamol is the most common cause of drug- induced liver injury, which is associated with oxidative stress, cytokine release, and apoptosis. Boswellic acid (BA), with anti- inflammatory and antioxidant effects, can ameliorate liver injury but has poor solubility and bioavailability. The objective of this study was to design boswellic acid nanoparticles (BA-NPs) and test their protective effect against paracetamol-induced liver injury in mice. Methods: BA-NPs were prepared by a surfactant-assisted emulsion method and characterized by UV-Vis spectroscopy, FT-IR analysis, dynamic light scattering (DLS), zeta potential measurements, and transmission electron microscopy (TEM). Acute oral toxicity (LD₂⁽) was assessed in healthy mice. Liver- intoxicated mice were treated with paracetamol (1 g/kg, po) and pretreated for 14 days with BA- NPs (75 mg/kg, po) or silymarin (50 mg/kg). Measurements included serum liver enzymes (ALT, AST, and ALP), lipid profile markers (TC, TG, and HDL), oxidative stress indicators (GSH, CAT, GPx, SOD, and MDA), inflammatory cytokines and their markers (IL- 6 and TNF–α), the apoptotic marker p 53, caspase- 8 and STAT 3 mRNA expression measured by qPCR, and molecular docking simulations to predict boswellic acid's binding affinity to target proteins. Results: The formulated BA- NPs were spherical and monodisperse (54. 54.54 ± 2. 76 nm) with colloidal stability (zeta potential:- 18. 84 mV; PDI: 0. 26). The LD50 was estimated to be 1500 mg/kg, indicating relatively low acute toxicity. BA-NP pretreatment alleviated paracetamolinduced increases in ALT, AST, and ALP; normalized dyslipidemia; restored antioxidant activity; and decreased MDA levels. We observed that the levels of pro- inflammatory cytokines (IL- 6, TNF- α) and p 53 were strongly downregulated. The expression of both caspase-8 and STAT3 was also downregulated by BA-NPs. Molecular docking data showed that boswellic acid bound significantly to caspase-8 (ΔG = -7.97 kcal/mol) and STAT3 (ΔG = -7.95 kcal/mol), supporting its mechanism of hepatoprotection. Discussion: Our results indicate that BA- NPs prepared by Span 60/Tween 80 emulsification provide potent hepatoprotection against paracetamol- induced liver injury through synergistic, multi- mechanistic action, significantly protecting against hepatic injury by concurrently suppressing oxidative stress, inflammatory signaling pathways, and apoptosis, three interconnected molecular events critical to the pathogenesis of paracetamol hepatotoxicity. Conclusion: BA-NPs prepared via emulsification of Span 60/Tween 80 offer potent hepatoprotection against paracetamol-induced liver injury by primarily modulating oxidative stress, inflammation, and apoptosis. This approach not only highlights a potentially novel therapeutic for drug- induced hepatotoxicity but also overcomes the pharmacokinetic barriers of the native boswellic acid agent and provides an improved alternative to currently available drugs.