结合组织学评分的牛角膜浑浊和渗透性方法研究

Research on bovine corneal opacity and permeability test integrating with histopathological evaluation

为了优化替代眼刺激动物试验的牛角膜浑浊和渗透性(BCOP)实验的方法,提高其适用范围和预测性,选取新鲜牛眼角膜,直接暴露液体化学受试物作用10 min,后孵育2 h并用荧光素钠处理90 min,采用浊度仪检测角膜浑浊度,用酶标仪检测荧光素钠渗透性,同时将角膜常规固定和制备组织学切片,观察组织学病理变化并评分,基于上述3个指标建立预测模型。结果显示,将角膜浑浊度和渗透性2个定量指标,与角膜上皮组织、基质组织和内皮细胞的组织学变化评分进行组合,可对轻微刺激到中等刺激的范围进行准确细分。研究表明,结合组织学观察的优化BCOP方法可提高其预测性,适用于眼刺激机制研究和化妆品评估。

In order to refine the alternative method for animal eye irritation test——the bovine corneal opacity and permeability（ BCOP） test,and to enhance its application scope and predictability,the fresh bovine corneas were chosen and directly exposed to liquid chemical material to be tested for 10 min,post- incubated for 2 h and further treated with fluorescein sodium for 90 min. Then opacity of the treated corneas was measured with opacitometer; the permeability was determined with the multiskan spectrum microplate spectrophotometer and the histopathological changes of the cornea were observed and graded after routinely fixed and paraffin embedding. Finally,the predictive models based on the above three indices were established. The results showed that the corneal opacity and permeability,integrated with histopathological changes of epithelium,stroma and endothelium,the scope of irritation from mild level to moderate level can be accurately distinguished.In conclusion,the refined BCOP method integrated with histological observation could improve its predictability and suitable for study of eye irritation mechanism and cosmetic product evaluation.