Silica-challenged macrophage secretome drives distinct alveolar and bronchial epithelial-fibroblast crosstalk: a multicellular preclinical modelling approach

Abstract Background Silicosis is an irreversible occupational lung disease that involves complex cellular mechanisms including inflammation, epithelial-mesenchymal transition (EMT), matrix remodelling, and fibrosis. Most in vitro studies have used conventional 2D models, which fail to orchestrate tissue architecture. While animal models have provided valuable information on silicosis pathogenesis, differences in anatomy and respiratory characteristics limit their use. With rapid advancements in 3D culture technologies, it is important to develop more physiologically relevant models to improve our understanding of silicosis. Methods THP-1-derived macrophages were exposed to silica nanoparticles, and their viability, cytotoxicity, and inflammatory responses were assessed by MTT assay ,LDH assay, and real-time PCR. This silica secretome was applied to alveolar and bronchial epithelial cells and fibroblasts established in 2D, 3D spheroids, and transwell models either in mono- culture or co-cultures. Cellular viability, inflammatory response, EMT, and extracellular matrix remodelling (ECM) were evaluated by CellTiter-Glo, real-time PCR, and western blotting. Results Silica nanoparticles induced cytotoxicity and inflammation in macrophages. Silica secretome altered cell viability and inflammatory response in 2D models of epithelial cells. Alveolar epithelial cells in 3D models showed a partial EMT transition (hybrid), while alveolar epithelial-fibroblast 3D co-culture demonstrated pronounced effects on viability, inflammation, and fibrotic remodeling compared to bronchial epithelial cells. Transwell co-cultures further revealed epithelial cell type-dependent differences in fibroblast activation. Conclusion These findings highlight the significance of multicellular 3D models in better reflecting key silicosis mechanisms and support their potential use as a physiologically relevant platform for mechanistic studies and therapeutic screening.