Lee Seung-Mok, Lee Da-In, Park Sung-Hee, Yu Hak-Sun, Lee Ji-Eun, Lee Jong-Soo
*Department of Geological Sciences, Pusan National University College of Natural Sciences, Pusan, Korea; †Department of Parasitology, Pusan National University College of Medicine, Yangsan, Korea; ‡Department of Ophthalmology, Pusan National University College of Medicine, Yangsan Pusan National University Hospital, Yangsan, Korea; and §Department of Ophthalmology, Pusan National University College of Medicine, Medical Research Institute, Pusan National University Hospital, Pusan, Korea.
Cornea. 2017 Dec;36(12):1538-1543. doi: 10.1097/ICO.0000000000001340.
The aim of the study was to evaluate the effect of multipurpose contact lens (CL) solution (MPS) combined with autophagy inhibitors on the adhesion properties of Acanthamoeba castellanii (AC) trophozoites to silicone hydrogel CLs.
AC trophozoites were inoculated onto discs trimmed from a silicone hydrogel CL with plasma surface treatment. After 18 hours, the number of adherent AC trophozoites on the CL treated with MPS was counted under phase-contrast microscopy. We analyzed the efficacy of MPS combined with autophagy inhibitors, 3-methyladenine (0.1, 1, and 5 mM) and chloroquine (10, 100 μM, and 1 mM), on Acanthamoeba adhesion by electron microscopy.
Adhesion of AC trophozoites to the CL treated with MPS (average number of trophozoites adhered to the CL: 61.2 ± 8.1) was significantly lower compared with that of the CL treated without MPS (83.8 ± 10.2) (P = 0.027). In MPS application, the number of adhered AC trophozoites treated with 3-methyladenine [8.2 ± 2.5 for 5 mM (P = 0.008)] or chloroquine [19.4 ± 7.6 for 100 μM (P = 0.038) and 5.3 ± 1.9 for 1 mM (P = 0.001)] was significantly reduced compared with the sample without autophagy inhibitors. However, the number of adherent AC trophozoites was not significantly reduced in less than 5 mM in 3-methyladenine [50.4 ± 5.1 for 0.1 mM (P = 0.084) and 43.1 ± 5.0 for 1 mM (P = 0.079)] and 100 μM in chloroquine [40.6 ± 13.5 for 10 μM (P = 0.075)]. 3-methyladenine induced blebby structures or disrupted the membranes of AC trophozoites. AC trophozoites treated with chloroquine showed undigested organelles in the cytoplasm of Acanthamoeba cells.
MPS combined with 3-methyladenine or chloroquine reduced the adhesion rate of AC trophozoites rather than MPS containing only polyhexamethylene biguanide. Appropriate concentrations of autophagy inhibitors, 3-methyladenine and chloroquine, added to commercial MPS should be considered to decrease the clinical rate of Acanthamoeba keratitis.
本研究旨在评估多功能隐形眼镜护理液（MPS）联合自噬抑制剂对卡氏棘阿米巴（AC）滋养体与硅水凝胶隐形眼镜黏附特性的影响。
将经等离子体表面处理的硅水凝胶隐形眼镜修剪成圆盘状，接种AC滋养体。18小时后，在相差显微镜下计数经MPS处理的隐形眼镜上黏附的AC滋养体数量。我们通过电子显微镜分析了MPS联合自噬抑制剂3 - 甲基腺嘌呤（0.1、1和5 mM）和氯喹（10、100 μM和1 mM）对棘阿米巴黏附的效果。
与未经MPS处理的隐形眼镜（83.8 ± 10.2）相比，AC滋养体对经MPS处理的隐形眼镜的黏附（黏附于隐形眼镜的滋养体平均数量：61.2 ± 8.1）显著降低（P = 0.027）。在使用MPS时，与未添加自噬抑制剂的样本相比，用3 - 甲基腺嘌呤处理的黏附AC滋养体数量[5 mM时为8.2 ± 2.5（P = 0.008）]或氯喹处理的[100 μM时为19.4 ± 7.6（P = 0.038），1 mM时为5.3 ± 1.9（P = 0.0..1）]显著减少。然而，3 - 甲基腺嘌呤浓度低于5 mM[0.1 mM时为50.4 ± 5.1（P = 0.084），1 mM时为43.1 ± 5.0（P = 0.079）]和氯喹浓度低于100 μM[10 μM时为40.6 ± 13.5（P = 0.075）]时，黏附的AC滋养体数量没有显著减少。3 - 甲基腺嘌呤诱导形成泡状结构或破坏AC滋养体的膜。用氯喹处理的AC滋养体在棘阿米巴细胞的细胞质中显示有未消化的细胞器。
MPS联合3 - 甲基腺嘌呤或氯喹可降低AC滋养体的黏附率，而不仅仅是含聚六亚甲基双胍的MPS。应考虑在商业MPS中添加适当浓度的自噬抑制剂3 - 甲基腺嘌呤和氯喹，以降低棘阿米巴角膜炎的临床发病率。
