Optimized Method for Establishing Primary Human Mesothelial Cell Cultures Preserving Epithelial Phenotype

Mesothelial cells (MCs) are highly relevant for studying the pathogenesis of serosal diseases, fibrosis, inflammation, and tumor progression. However, the isolation and maintenance of an epithelial-like phenotype of MCs in vitro remain methodologically challenging due to their tendency to undergo mesothelial-to-mesenchymal transition (MMT). In this work, we propose a combined protocol utilizing collagen IV coating, conditioned medium, and short-term ROCK inhibitor treatment, which improves cell survival. This approach enables the establishment of primary human cultures suitable for investigating mesothelial cell functional activity and for assessing the efficacy of potential therapeutic strategies.