Lebrun Labs LLC, Anaheim, CA, United States of America.
Gavin Herbert Eye Institute, University of California, Irvine, CA, United States of America.
Toxicol In Vitro. 2019 Dec;61:104628. doi: 10.1016/j.tiv.2019.104628. Epub 2019 Aug 13.
The purpose of this study was to develop Globally Harmonized System (GHS) and U.S. Environmental Protection Agency (EPA) prediction models for classifying irritant materials based on histopathologic in vitro depth of injury (DoI) measurements. Sixteen different materials were selected, representing all classes of toxicity, according to the GHS and EPA classification systems. Food-source rabbit eyes, similar to eyes used for the widely accepted Bovine Corneal Opacity and Permeability and Isolated Chicken Eye ocular irritation tests, were used. Tissues were exposed to test material for 1 min, and corneas were collected at 3- and 24-hours post-exposure. Tissues were then fixed and processed for live/dead biomarker fluorescent staining using phalloidin. DoI was then measured, and the percent DoI values for the epithelium and stroma were compared to the EPA and GHS classifications. Excluding surfactants, EPA nonclassified (category IV) materials showed no stromal and very slight epithelial damage (≤10%) to the cornea, whereas EPA corrosive (category I) materials showed significantly greater damage (P < 0.001), ranging from 39% to 100% of the stromal depth. Importantly, EPA reversible (categories II and III) materials showed significant damage to the epithelium (>10%, P < 0.005) but significantly less severe damage to the corneal stroma (P < 0.001), ranging from 1% to 38% of the stromal depth. GHS nonclassified (category NC) irritants caused damage to the epithelium but not to the stroma. All GHS class 2 materials showed damage to the stroma (1-11%), whereas GHS corrosives caused significantly greater damage to the stroma (38-100%; P < 0.001). Additionally, one corrosive material, which produced a stromal DoI of 99% at 24 h, produced no apparent damage at 3-hours post-exposure. Based on these findings, histopathologic EPA and GHS prediction models are proposed that appear to separate and identify reversible irritants from other irritant classes. Furthermore, GHS classification appears to require stromal damage, whereas NC materials may or may not damage the corneal epithelium.
本研究旨在开发全球协调系统 (GHS) 和美国环境保护署 (EPA) 的预测模型，根据组织病理学体外损伤深度 (DoI) 测量结果对刺激性材料进行分类。根据 GHS 和 EPA 的分类系统，选择了 16 种不同的材料，代表了所有毒性类别。使用类似于广泛接受的牛角膜混浊和通透性以及鸡眼眼刺激性试验中使用的兔眼，这些兔眼来自食物来源。组织暴露于测试材料 1 分钟，在暴露后 3 小时和 24 小时收集角膜。然后用鬼笔环肽对组织进行活/死生物标志物荧光染色固定和处理。然后测量 DoI，并将上皮和基质的百分比 DoI 值与 EPA 和 GHS 分类进行比较。不包括表面活性剂，EPA 未分类 (IV 类) 材料对角膜无基质和极轻微的上皮损伤 (≤10%)，而 EPA 腐蚀性 (I 类) 材料则显示出明显更大的损伤 (P < 0.001)，范围从 39%到 100%的基质深度。重要的是，EPA 可逆 (II 和 III 类) 材料对上皮有明显的损伤 (>10%，P < 0.005)，但对角膜基质的损伤明显较轻 (P < 0.001)，范围从 1%到 38%的基质深度。GHS 未分类 (NC) 刺激性物质对上皮造成损伤，但对基质无损伤。所有 GHS 2 类材料均对基质造成损伤 (1-11%)，而 GHS 腐蚀性物质对基质造成的损伤明显更大 (38-100%；P < 0.001)。此外，一种腐蚀性物质在 24 小时时基质 DoI 为 99%，但在暴露后 3 小时时没有明显损伤。基于这些发现，提出了组织病理学 EPA 和 GHS 预测模型，似乎可以将可逆刺激性物质与其他刺激性物质区分开来。此外，GHS 分类似乎需要基质损伤，而 NC 材料可能会或可能不会损伤角膜上皮。
