From inflammation to cytotoxicity: exposure pattern-dependent responses to alumina nanoparticles and dissolved hydrochloric acid in a human alveolar-capillary barrier model at the air-liquid interface.
Combustion of composite solid propellants generates complex aerosols containing metallic nanoparticles (NPs) and acidic gases, raising concerns about inhalation toxicity. Here, we investigated the effects of alumina (Al₂O₃) NPs, alone or combined with hydrochloric acid (HCl), in a human alveolar-capillary barrier (ACB) model exposed at the air-liquid interface. The model consisted of hAELVi alveolar epithelial cells co-cultured with HPMEC-ST1.6R pulmonary microvascular endothelial cells and exposed using a Vitrocell Cloud system under acute or repeated scenarios. Deposited Al₂O₃ NPs doses corresponding to low-to-high alveolar exposure scenarios were 0.007, 1.35 ± 0.1 and 2.3 ± 0.2µg/cm², with or without HCl at 1.37mM. Barrier integrity, viability, inflammatory and remodeling mediators, oxidative stress and DNA damage were assessed 24h after the final exposure. The co-culture formed a tight functional barrier with high transepithelial electrical resistance, low permeability and organized junctional protein expression. Acute exposure to the highest Al₂O₃ NPs dose induced a compartment-specific inflammation-related response, based on the biomarkers investigated, with increased IL-1α, IL-8, IL-1β, CCL2, DKK1 and angiogenin, without cytotoxicity or barrier disruption. Repeated exposure shifted the response toward moderate cytotoxicity at the highest dose, with viability decreasing to 77% for Al₂O₃ NPs alone and 66% for Al₂O₃ NPs with HCl, while permeability remained unchanged. No significant oxidative stress or γ-H2AX-associated DNA damage was detected. HCl co-exposure did not enhance Al₂O₃ NPs toxicity. However, because HCl was administered in dissolved form prior to nebulization, the contribution of potential gas-particle interactions occurring in combustion plumes could not be evaluated and the results should not be interpreted as evidence for or against synergistic effects arising from gas-particle interactions occurring in combustion plumes. These findings show that exposure pattern critically shapes alveolar-capillary responses, supporting human air-liquid interface co-culture models for mechanistic inhalation toxicology.