School of Medicine, Nankai University, Tianjin, 300071, China; Nankai University Affiliated Eye Hospital, Tianjin, 300020, China; Tianjin Key Laboratory of Ophthalmology and Visual Science, Tianjin Eye Institute, Tianjin Eye Hospital, Tianjin, 300020, China.
Tianjin Key Laboratory of Ophthalmology and Visual Science, Tianjin Eye Institute, Tianjin Eye Hospital, Tianjin, 300020, China; Clinical College of Ophthalmology, Tianjin Medical University, Tianjin, 300020, China.
Exp Eye Res. 2024 Oct;247:110030. doi: 10.1016/j.exer.2024.110030. Epub 2024 Aug 8.
Benzalkonium chloride (BAC) is commonly used as a preservative in ophthalmic medications, despite its potential to induce chemical injury. Extensive research has demonstrated that BAC can lead to adverse effects, including injuries to the ocular surface. Our study aimed to elucidate the underlying mechanism of necroptosis induced by BAC.
Human corneal epithelial (HCE) cells and mouse corneas were subjected to chemical injury, and the necrostatin-1 (Nec1) group was compared to the dimethylsulfoxide (DMSO) group. The extent of damage to HCE cells was assessed using CCK-8 and flow cytometry. Hematoxylin and eosin staining, as well as fluorescein sodium staining, were used to detect and characterize corneal injury. The activation of inflammatory cytokines and necroptosis-related proteins and genes was evaluated using Western blotting, immunofluorescence staining, and quantitative RT‒PCR.
In our study, the induction of necroptosis by a hypertonic solution was not observed. However, necroptosis was observed in HCE cells exposed to NaOH and BAC, which activated the receptor-interacting protein kinase 1 (RIPK1) - receptor-interacting protein kinase 3 (RIPK3) - mixed lineage kinase domain-like protein (MLKL) signaling pathway. In mouse corneal tissues, BAC could induce necroptosis and inflammation. The administration of Nec1 mitigated the inflammatory response and ocular surface damage caused by BAC-induced necroptosis in our experimental models. Furthermore, our in vivo experiments revealed that the severity of necroptosis was greater in the 3-day group than in the 7-day group.
Necroptosis plays a role in the pathological development of ocular surface injury caused by exposure to BAC. Furthermore, our study demonstrated that the administration of Nec1 could mitigate the pathological effects of necroptosis induced by BAC in clinical settings.
苯扎氯铵（BAC）常用于眼科药物的防腐剂，尽管其有引发化学损伤的潜在风险。大量研究表明，BAC 可导致不良反应，包括眼表损伤。我们的研究旨在阐明 BAC 诱导细胞坏死的潜在机制。
用化学物质损伤人角膜上皮（HCE）细胞和小鼠角膜，并用 necrostatin-1（Nec1）组与二甲基亚砜（DMSO）组进行比较。通过 CCK-8 和流式细胞术评估 HCE 细胞的损伤程度。用苏木精和伊红染色以及荧光素钠染色来检测和描述角膜损伤。用 Western blot、免疫荧光染色和实时定量 RT-PCR 评估炎症细胞因子和坏死相关蛋白和基因的激活。
在我们的研究中，未观察到高渗溶液诱导的细胞坏死。然而，NaOH 和 BAC 暴露于 HCE 细胞中观察到细胞坏死，其激活受体相互作用蛋白激酶 1（RIPK1）-受体相互作用蛋白激酶 3（RIPK3）-混合谱系激酶结构域样蛋白（MLKL）信号通路。在小鼠角膜组织中，BAC 可诱导细胞坏死和炎症。在我们的实验模型中，Nec1 的给药减轻了 BAC 诱导的细胞坏死引起的炎症反应和眼表损伤。此外，我们的体内实验表明，3 天组的细胞坏死程度比 7 天组更严重。
细胞坏死在暴露于 BAC 引起的眼表损伤的病理发展中起作用。此外，我们的研究表明，在临床环境中，Nec1 的给药可以减轻 BAC 诱导的细胞坏死的病理效应。
