Using organoid platform to investigate the effect of smoking on congenital lung malformations

BACKGROUND: Congenital lung malformation (CLMs) is a series of developmental lung abnormalities that may persist into adulthood. Some centres would advocate an observation approach rather than opting for surgical excision. However, the potential impact of environmental factors, particularly cigarette smoke exposure, on CLM-affected lung tissue remains poorly understood. AIM: This study aimed to evaluate the effects of cigarette smoke extract (CSE) on paired normal and diseased airway organoids (AOs) derived from pediatric lung tissue diagnosed as intralobar sequestration, using N-[N-(3,5-Difluorophenacetyl)-L-alanyl]-S-phenylglycine t-butyl ester (DAPT), a γ-secretase inhibitor, to modulate airway epithelial differentiation. METHOD: AOs were established from paired normal and diseased regions of pediatric lung tissue. Organoids were cultured under two conditions: without DAPT or with DAPT pretreatment, followed by 4 % CSE exposure. Mucus viscosity, MUC5AC protein expression, PAS staining, and RT-qPCR analysis of airway epithelial cell markers, tight junction proteins (E-cadherin, ZO-1), and proinflammatory cytokines were measured. RESULTS: Disease-derived AOs exhibited higher mucus secretion compared to normal tissue-derived AOs. CSE exposure enhanced mucus viscosity and MUC5AC expression in both groups, with more pronounced effects in diseased organoids. CSE-induced upregulation of mucins and pro-inflammatory cytokines was particularly evident in disease-derived organoids without DAPT treatment. DAPT pretreatment attenuated these CSE-induced changes, while tight junction protein expression remained stable across conditions. CONCLUSION: Our findings demonstrate that diseased airway organoids show enhanced susceptibility to cigarette-smoke -induced mucus hypersecretion and inflammatory responses, suggesting that patients with congenital airway malformations may be particularly vulnerable to the effects of cigarette smoke exposure.