Pollen hydration and its induced effects on human bronchial epithelial cells: A comparative analysis of three species

Pollen-induced allergic diseases are on the rise globally with recent studies pointing towards climate change co-factors potentially influencing pollen seasons and pollen load. This study investigated the hydration-induced rupturing behaviour and allergenic potential of pollen belonging to species of ryegrass (Lolium perenne), golden wattle (Acacia pycnantha) and pine (Pinus radiata). Using an in vitro model of human bronchial epithelial cells (Calu-3), this study evaluated cellular responses upon exposure to pollen extracts through multiple experimental approaches. Initial hydration experiments revealed species-specific rupturing patterns. Ryegrass pollen demonstrated the highest rupture propensity (38% immediately upon hydration), with increased rupturing over time and enhanced effects in acidic conditions. Wattle exhibited moderate rupturing that plateaued after 10 min, while pine pollen showed minimal rupturing (≤2%). Nebulisation of hydrated pollen-water suspensions generated distinct aerosol profiles, with ryegrass producing significant numbers of sub-pollen particles in the respirable range. Cell viability of Calu-3 cells showed varied concentration-dependent responses among pollen species. Pine pollen induced the most significant reduction in cell viability (51% at 25 mg/mL pollen extract concentration), followed by ryegrass and wattle pollen. Notably, ryegrass pollen uniquely triggered substantial reactive oxygen species (ROS) production, with rates increasing up to 33.8 times greater than controls at 25 mg/mL pollen extract concentration. This elevated ROS generation correlated with increased group 5 grass allergen (Phl p 5) levels (53.8 ng/mL at 25 mg/mL pollen extract concentration), suggesting a potential mechanistic link. In contrast, cell viability of Calu-3 cells was greater than 70% in the presence of wattle pollen with only a slight increase in cellular ROS generation, suggesting other mechanisms are involved in maintaining cell viability in this species. This research establishes a foundation for understanding cellular responses to pollen extracts of different species, including the impact of physical characteristics (rupture propensity and sub-pollen particle generation) and biochemical properties (ROS induction and allergen content), which can both contribute to overall allergic potential. The species-specific responses identified will also have important implications for clinical management strategies, particularly in the context of changing climate conditions.