In vivo, in vitro, and in silico approaches in the detailed study of di-butyl phthalate (DBP), a plasticizer-induced lung fibrosis via Nrf-2/Keap-1/HO-1 pathway and its regulation

Abstract

The alveolar epithelium is a crucial barrier against external threats, yet it becomes a key player in initiating pulmonary fibrosis when compromised. Despite its importance, the intricate relationship between, DBP exposure and alveolar epithelial cell injury ensuing pro-fibrotic effects remains poorly understood. Phthalates, ubiquitous in nature, pose a significant risk to lung health upon inhalation, acting as immune triggers that cause airway inflammation and epithelial damage. We aimed to investigate the impact of intranasal administration of Di-butyl Phthalate (DBP) inhalation, and its probable effects on normal and asthmatic lungs. DBP was administered via intranasal route in normal and OVA-induced asthmatic mice. DBP exposure enhanced oxidative stress and inflammatory parameters, leading to exacerbated asthmatic response and oxidative lung damage. Enhanced accumulation of immune cells, bronchial thickening, and collagen deposition was noted in histopathological investigations of DBP-exposed lung sections. Curcumin, a plant-derived molecule, significantly mitigated DBP-exposed asthma exacerbations by suppressing NF-κB expression and enhancing NRF2 levels via the Nrf-2/Keap-1/HO-1 signaling pathway. FACS analysis revealed increased CD11b+ cells (32 %) in asthmatic mice which were reduced in the curcumin pre-treatment group (10.5 %). Enhanced epithelial to mesenchymal transition (EMT) was noted in mice lungs and A549 cells where E-cadherin expression was reduced as compared to Vimentin, and α-SMA. Apart from aggravated airway inflammation, DBP exposure damages healthy lungs also. MMP-9/TIMP-1 ratios and collagen-1 levels were restored which were enhanced after DBP exposure. Moreover, antioxidant enzyme levels such as NQO-1, HO-1, and Catalase were significantly enhanced (p < 0.01) and comparable to dexamethasone, a conventional corticosteroid. Notably, both dexamethasone and curcumin treatments effectively regulated the stimulation and accumulation of Nrf-2 in the nucleus, promoting antioxidant production and offering potential therapeutic benefits in mitigating pulmonary fibrosis. OVA and DBP alone caused DNA damage in the lung cells where increased percentage of damaged DNA movement in the tail, tail length, tail moment, and olive tail moment indicated severe damage in the DBP and OVA combined exposure strategies. Dexamethasone and Curcumin treatments reduced the extent of the DNA damage indicating anti-inflammatory and ant-oxidative potentials. Moreover, in silico studies are supportive of therapeutic potential of Curcumin and Dexamethasone in DBP-induced lung inflammation and fibrosis.