Lee Jong Heon, Lee Jong Soo, Kim Sujin, Lee Ji Eun
Department of Ophthalmology, School of Medicine, Pusan National University, Busan 49241, Korea.
Department of Ophthalmology, Gyeongsang National University Changwon Hospital, School of Medicine, Gyeongsang National University, Changwon 51472, Korea.
Korean J Physiol Pharmacol. 2017 Mar;21(2):189-195. doi: 10.4196/kjpp.2017.21.2.189. Epub 2017 Feb 21.
This study aimed to compare the cellular toxicities of three clinically used dry eye treatments; 3% diquafosol tetrasodium and hyaluronic acid at 0.3 and 0.18%. A methyl thiazolyltetrazoiun (MTT)-based calorimetric assay was used to assess cellular proliferation and a lactate dehydrogenase (LDH) leakage assay to assess cytotoxicity, using Human corneal epithelial cells (HCECs) exposed to 3% diquafosol tetrasodium, 0.3% hyaluronic acid (HA), or 0.18% HA or 1, 6 or 24 h. Cellular morphology was evaluated by inverted phase-contrast light microscopy and electron microscopy, and wound widths were measured 24 h after confluent HCECs were scratched. Diquafosol had a significant, time-dependent, inhibitory effect on HCEC proliferation and cytotoxicity. HCECs treated with diquafosol detached more from the bottoms of dishes and damaged cells showed degenerative changes, such as, reduced numbers of microvilli, vacuole formation, and chromatin of the nuclear remnant condensed along the nuclear periphery. All significantly stimulated reepithelialization of HCECs scratched, which were less observed in diquafosol. Therefore, epithelial toxicity should be considered after long-term usage of diquafosol and in overdose cases, especially in dry eye patients with pre-existing punctated epithelial erosion.
本研究旨在比较三种临床使用的干眼症治疗药物的细胞毒性;3%的四水二喹富罗钠以及0.3%和0.18%的透明质酸。采用基于甲基噻唑基四唑(MTT)的比色法评估细胞增殖,采用乳酸脱氢酶(LDH)泄漏试验评估细胞毒性,将人角膜上皮细胞(HCECs)分别暴露于3%的四水二喹富罗钠、0.3%的透明质酸(HA)、0.18%的HA中1、6或24小时。通过倒置相差显微镜和电子显微镜评估细胞形态,并在汇合的HCECs划痕后24小时测量伤口宽度。二喹富罗对HCEC增殖和细胞毒性具有显著的、时间依赖性的抑制作用。用二喹富罗处理的HCECs从培养皿底部脱落得更多,受损细胞表现出退行性变化,如微绒毛数量减少、空泡形成以及核残余物的染色质沿核周边浓缩。所有药物均显著刺激了划痕的HCECs的再上皮化,而在二喹富罗中观察到的较少。因此,长期使用二喹富罗后以及过量使用时,尤其是在已有点状上皮糜烂的干眼症患者中,应考虑上皮毒性。