Effects of benzalkonium chloride and cyclosporine applied topically to rabbit conjunctiva: a histomorphometric study

ABSTRACT Purpose: Chronic instillation of benzalkonium chloride, a preservative, has inflammatory effects on the ocular surface. However, addition of the anti-inflammatory agent cyclosporine to a therapeutic protocol may mitigate these effects. This study compared the toxic effects of a 0.1% benzalkonium chloride solution and the possible protective effect of 0.05% cyclosporine when applied topically to the rabbit conjunctiva. Methods: Fifteen age- and weight-matched, female New Zealand white rabbits were categorized into three groups and treated for 30 consecutive days. Group 1, 2, and 3 - benzalkonium chloride received 0.1% every 24 h, 0.05% cyclosporine every 6 h, and both treatments, respectively. In each rabbit, the left eye was subjected to treatment and the right eye was a control. The rabbits were euthanized at after the experiment. Goblet cells and blood vessels were then enumerated in conjunctival tissues stained with periodic acid-Schiff and hematoxylin-eosin, respectively. Differences between treated and untreated eyes and between groups were compared using the t-test and analysis of variance. Results: Benzalkonium chloride treatment, with and without cyclosporine, significantly reduced (p≤0.05) in the number of goblet cells in treatment eyes compared with that in respective control eyes. Alternatively, adding cyclosporine to benzalkonium chloride did not prevent the loss of conjunctival goblet cells, and a significant reduction in the number of goblet cells was noted. Benzalkonium chloride-induced significant increase in the number of new blood vessels was mitigated significantly by the addition of cyclosporine. Conclusion: This study demonstrated the magnitude of conjunctival injury caused by chronic instillation of benzalkonium chloride. Although cyclosporine did not mitigate the effects on goblet cells, its addition minimized inflammatory angiogenesis induced by benzalkonium chloride.

摘要
目的：防腐剂苯扎氯铵（benzalkonium chloride）长期滴眼会对眼表产生炎性损伤。然而，在治疗方案中加入抗炎药物环孢素（cyclosporine）或可缓解此类损伤。本研究旨在比较0.1%苯扎氯铵溶液的毒性效应，以及局部滴用0.05%环孢素对兔结膜的潜在保护作用。
方法：选取15只年龄、体重匹配的雌性新西兰白兔，分为3组，连续给药30天。第1、2、3组分别为：每24小时滴用0.1%苯扎氯铵、每6小时滴用0.05%环孢素，以及联合使用上述两种疗法。每只家兔的左眼为给药眼，右眼为对照眼。实验结束后对家兔实施安乐死，分别采用过碘酸-雪夫染色（periodic acid-Schiff）和苏木精-伊红染色（hematoxylin-eosin）对结膜组织中的杯状细胞和血管进行计数。采用t检验（t-test）和方差分析（analysis of variance）比较给药眼与对照眼、以及各组间的差异。
结果：无论是否联合使用环孢素，苯扎氯铵给药组的给药眼杯状细胞数量均较对应对照眼显著降低（p≤0.05）。值得注意的是，在苯扎氯铵给药方案中加入环孢素并未阻止结膜杯状细胞的丢失，仍可见杯状细胞数量显著减少。而苯扎氯铵诱导的新生血管数量显著增加的效应，可通过联合环孢素得到显著缓解。
结论：本研究证实了长期滴用苯扎氯铵所造成的结膜损伤程度。尽管环孢素未能缓解其对杯状细胞的损伤，但可有效减轻苯扎氯铵诱导的炎性血管新生。