Evaluation of the Ultrastructural Effects on Conjunctival Epithelial Cells of a New Multiple-Action Artificial Tear Containing Cross-Linked Hyaluronic Acid, Cationic Liposomes, and Trehalose with Transmission Electron Microscopy: A Pilot Study

<b>Background:</b> This study aimed to evaluate the effects of Trimix^® on the microvilli and glycocalyx of ocular conjunctival epithelial cells, assessing drug persistence on the cell surface and its interaction with the glycocalyx. Microvilli, vital indicators of cellular health, are altered in inflammatory or toxic conditions, making their restoration a key therapeutic target. <b>Method:</b> Building upon previous scanning electron microscopy work, this investigation utilized Transmission Electron Microscopy (TEM) to delve into the direct interaction between Trimix and the cell membrane, elucidating its role in cellular mechanisms. The research involved both an in vitro phase, examining the drug's molecular arrangement, and an in vivo phase, treating three subjects (healthy, moderate inflammation, severe dry eye) for 30 days. Cytological samples were taken via impression cytology for TEM analysis to observe the drug's long-term action and its influence on microvillar structures, glycocalyx, and vesicular transport. <b>Results:</b> We demonstrated that Trimix stimulated vesicular transport and promoted the formation of a rudimentary glycocalyx, significantly increasing its presence and the number of microvilli in treated patients across all inflammatory grades, even in severe dry eye. <b>Conclusions:</b> In conclusion, Trimix acts as an effective glycocalyx substitute, restoring the second mucosal system (SMS) and enabling distressed cells to resume essential exchange functions, offering a novel therapeutic approach for dry eye disease.