Integrated in silico and in vivo approaches reveal naltrexone as a synergistic modulator of TLR4 and μ-opioid receptors restoring redox homeostasis via Nrf2/Keap-1/HO-1 and HIF-1α/VEGF pathways in COPD.

Chronic obstructive pulmonary disease (COPD) is a progressive, multifactorial respiratory disorder in which oxidative stress plays a central role. Oxidative stress arises from an imbalance between the excessive generation and accumulation of reactive oxygen species (ROS) in cells and reduced capacity of antioxidant defenses to eliminate them. Toll-like receptor-4 (TLR4) and μ-opioid receptor (µOR) signalling have emerged as key mediators of COPD pathogenesis by amplifying pro-inflammatory pathways. Naltrexone (NTX) was selected over naloxone based on comparative ADMET and molecular docking analyses, which revealed that NTX possesses more favourable pharmacokinetic profile and stronger binding affinities toward both µOR and TLR4, supporting its suitability as a dual-target therapeutic agent. Present study explores the therapeutic potential of ultra-low dose naltrexone (ULDN; 10 µg/kg body weight (BW), 1 µg/kg BW and 0.1 µg/kg BW) in mitigating cigarette smoke (CS) induced pulmonary inflammation and injury via modulation of key immunoinflammatory pathways. CS exposure resulted significant increased lung inflammation, oxidative damage, and dysregulated redox signalling pathways. NTX administration markedly attenuated airway inflammation and emphysematous changes and restored redox homeostasis via enhancing antioxidant enzyme activities (superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GPx), glutathione S-transferases (GST) and reducing oxidative stress markers (ROS, Nitric oxide (NO), myeloperoxidase (MPO), Total oxidant status (TOS) and Oxidative stress index (OSI). NTX attenuated apoptosis through downregulation of cleaved caspase-3, protein carbonyls and lipid peroxidation. NTX downregulated TLR4 and µOR expression and modulated antioxidant defence by regulating Nrf2/keap-1/HO-1 and HIF-1α/VEGF cascades thereby disrupting key inflammatory and hypoxia-induced pathways. Our study evaluated that the lowest dose of NTX (0.1 µg/kg BW) exhibited the most pronounced therapeutic effects compared with 1 µg/kg BW and 10 µg/kg BW. This dose exhibiting strongest anti-inflammatory and antioxidant efficacy in COPD by inhibition of TLR4 and µOR signalling, highlights its potential as a dual-target therapeutic strategy against oxidative stress and airway inflammation.