Bio-INvENT Lab, Department of Chemical Engineering, Siddaganga Institute of Technology, Tumakuru, India.
School of Optometry, Indiana University, Bloomington, Indiana, USA.
J Ocul Pharmacol Ther. 2022 Oct;38(8):549-560. doi: 10.1089/jop.2022.0044.
Local hotspots of elevated tear hyperosmolarity (exceeding 900 mOsM) are predicted in dry eye disease (DED) but have not been measured. This study aims to develop, characterize, and evaluate the suitability of fluorescent nanoliposomes for noninvasive sensing of the local osmolarity of the tear film. Fluorescent nanoliposomes, loaded with calcein (susceptible to self-quenching; sensor dye) and sulforhodamine 101 (SR101; reference dye), were produced by the thin-film hydration method. Dynamic light scattering measurements and Cryo-TEM showed that liposomes were negatively charged (-23.7 ± 1.5 mV), spherical (diameter = 117.9 ± 6.4 nm), and uniform in size (polydispersity index = 0.15 ± 0.02). These properties were unaffected by cold storage (4°C; 14 days), but dye leakage was significant after 3 days. Swelling and shrinkage of the liposomes by exposure to hypoosmotic and hyperosmotic media led to rapid dequenching and quenching of calcein fluorescence (F), with no effect on SR101 fluorescence (F). The ratio F/F decreased with increasing osmolarity and , obeying the Boyle van't Hoff relationship. When liposomes were dispersed in a gelatin film with dynamic radial sucrose gradients, local F/F decreased with increasing hyperosmolarity as predicted. When instilled on the hydrophilic surface of contact lenses or corneas, nanoliposomes dispersed evenly as thin films. Importantly, the measured F/F declined continuously with evaporation and consequent increase in their osmolarities. The study provides proof of principle for noninvasive measurement of local tear film osmolarity based on osmosensitive fluorescent nanoliposomes. The strategy can potentially advance our understanding of the pathophysiology of DED.
局部泪液高渗透压热点（超过 900 mOsM）在干眼病（DED）中被预测到，但尚未测量。本研究旨在开发、表征并评估荧光纳米脂质体用于非侵入性测量泪膜局部渗透压的适用性。通过薄膜水化法制备负载钙黄绿素（易自猝灭；传感器染料）和磺基罗丹明 101（SR101；参考染料）的荧光纳米脂质体。动态光散射测量和 Cryo-TEM 显示脂质体带负电荷（-23.7 ± 1.5 mV）、呈球形（直径= 117.9 ± 6.4nm）且粒径均匀（多分散指数= 0.15 ± 0.02）。这些特性不受冷藏（4°C；14 天）的影响，但染料泄漏在 3 天后显著增加。脂质体暴露于低渗和高渗介质会导致肿胀和收缩，从而快速猝灭和荧光再激发钙黄绿素荧光（F），而对 SR101 荧光（F）没有影响。F/F 比值随渗透压的增加而降低，符合 Boyle van't Hoff 关系。当脂质体在含有动态径向蔗糖梯度的明胶膜中分散时，局部 F/F 随渗透压的增加而降低，这与预测结果一致。当滴注在亲水隐形眼镜或角膜表面时，纳米脂质体均匀分散成薄膜。重要的是，随着蒸发和渗透压的增加，测量的 F/F 持续下降。该研究为基于对渗透压敏感的荧光纳米脂质体的非侵入性测量局部泪膜渗透压提供了原理验证。该策略有可能促进我们对 DED 病理生理学的理解。
