Evaluation of Trans-epithelial Penetration and Microplastic-induced Tissue Damage in a 3d Model of Human Respiratory Mucosa

Microplastics (MPs), ubiquitous pollutants, pose an emerging risk to human health.1 The accumulation of MPs, driven by the overuse of plastics, has established ingestion (through the food chain) and inhalation (through ambient air) as the primary routes of human exposure.2 Previous studies have confirmed that approximately 6,000 particles are inhaled in a polluted area over 18 cubic metres in 24 hours, which promotes their accumulation in the respiratory tract. Despite studies on the systemic effects of ingested MPs, their pulmonary impact, mechanisms of interaction with the epithelial barrier and potential tissue damage remain poorly investigated. In this study, we used a 3D air-liquid interface (ALI) culture model of human respiratory mucosa to evaluate:3 the ability of MPs to cross the epithelium and persist at the tissue level; the induction of cellular stress and epithelial damage. Samples of polystyrene MPs stained dark blue and dark red, 10 μm and 3 μm in diameter, respectively (SIGMA-ALDRICH), were diluted in PBS, counted and placed apically on the confluent and differentiated respiratory mucosa. The cultures were treated simulating acute (48 hours) and prolonged (1 week) exposures. At the end of the exposure to MPs treatments, the cultures were fixed in formalin and embedded in paraffin for subsequent analysis. The immunomorphological analyses aimed to assess the differential/potential penetration of MPs based on size, as well as any morphological changes that occurred to the epithelial layer of the mucosa. Epithelial stress markers were evaluated to determine any changes resulting from different exposure times.