Enríquez de Salamanca Amalia, Diebold Yolanda, Calonge Margarita, García-Vazquez Carmen, Callejo Sagrario, Vila Ana, Alonso Maria José
Instituto de Oftalmobiologia Aplicada (IOBA), University of Valladolid, Spain.
Invest Ophthalmol Vis Sci. 2006 Apr;47(4):1416-25. doi: 10.1167/iovs.05-0495.
To study the in vitro and in vivo interaction of chitosan nanoparticles (CSNPs), a new particulate drug carrier, with epithelial cells on the ocular surface.
CSNPs labeled with fluorescein isothiocyanate-bovine serum albumin were produced by ionotropic gelation. Human conjunctival epithelial cells (IOBA-NHC) were exposed for 15, 30, 60, and 120 minutes to three different CSNP concentrations. Immediately after treatment and after a 24-hour recovery period in culture medium, cell survival, and viability were measured. The association of CSNPs with IOBA-NHC cells was investigated by confocal microscopy. The influence of temperature and the effect of metabolic inhibition were studied by fluorometry. The in vivo uptake and acute tolerance of the ocular surface to CSNPs were evaluated in rabbits.
Cell survival and viability of CSNP-exposed cells were equivalent to that of the control. Uptake of CSNPs was continuous for the 2-hour duration of these experiments and was temperature dependent. Metabolic inhibition by sodium azide had no effect on CSNP uptake. The rabbit ocular surface showed no signs of inflammation or alteration after CSNP exposure compared with the control. Fluorescence microscopy of rabbit eyeball and lid sections confirmed in vivo uptake by conjunctival and corneal epithelia.
CSNPs were internalized by IOBA-NHC cells by an active transport mechanism that did not compromise cell viability. Moreover, these nanoparticles were well tolerated by the ocular surface tissues. These facts add further support for the potential use of these colloidal systems to delivery drugs to the ocular surface.
研究新型颗粒药物载体壳聚糖纳米粒（CSNPs）与眼表上皮细胞的体外和体内相互作用。
通过离子凝胶法制备异硫氰酸荧光素-牛血清白蛋白标记的CSNPs。将人结膜上皮细胞（IOBA-NHC）暴露于三种不同浓度的CSNPs中15、30、60和120分钟。处理后立即及在培养基中恢复24小时后，测量细胞存活率和活力。通过共聚焦显微镜研究CSNPs与IOBA-NHC细胞的结合情况。通过荧光测定法研究温度的影响和代谢抑制的作用。在兔体内评估眼表对CSNPs的摄取和急性耐受性。
暴露于CSNPs的细胞的存活率和活力与对照组相当。在这些实验的2小时期间，CSNPs的摄取是持续的，并且依赖于温度。叠氮化钠的代谢抑制对CSNP摄取没有影响。与对照组相比，兔眼表在暴露于CSNPs后没有炎症或改变的迹象。兔眼球和眼睑切片的荧光显微镜检查证实结膜和角膜上皮在体内摄取了CSNPs。
CSNPs通过一种不损害细胞活力的主动转运机制被IOBA-NHC细胞内化。此外，这些纳米粒在眼表组织中耐受性良好。这些事实进一步支持了这些胶体系统在向眼表递送药物方面的潜在用途。
