Montmorillonite (Mt) and its derivatives are now widely used in industrial and biomedical fields. Therefore, safety assessments of these materials are critical to protect human health after exposure; however, studies on the ocular toxicity of Mt are lacking. In particular, varying physicochemical characteristics of Mt may greatly alter their toxicological potential. To explore the effects of such characteristics on the eyes, five types of Mt were investigated in vitro and in vivo for the first time, and their underlying mechanisms studied. The different types of Mt caused cytotoxicity in human HCEC-B4G12 corneal cells based on analyses of ATP content, lactate dehydrogenase (LDH) leakage, cell morphology, and the distribution of Mt in cells. Among the five Mt types, Na-Mt exhibited the highest cytotoxicity. Notably, Na-Mt and chitosan-modified acidic Na-Mt (C-H-Na-Mt) induced ocular toxicity in vivo, as demonstrated by increases corneal injury area and the number of apoptotic cells. Na-Mt and C-H-Na-Mt also induced reactive oxygen species (ROS) generation in vitro and in vivo, as indicated by 2′,7′-dichlorofluorescin diacetate and dihydroethidium staining. In addition, Na-Mt activated the mitogen-activated protein kinase signaling pathway. The pretreatment of HCEC-B4G12 cells with N-acetylcysteine, an ROS scavenger, attenuated the Na-Mt-induced cytotoxicity and suppressed p38 activation, while inhibiting p38 activation with a p38-specific inhibitor decreased Na-Mt-induced cytotoxicity. The results indicate that Mt induces corneal toxicity in vitro and in vivo. The physicochemical properties of Mt greatly affect its toxicological potential. Furthermore, ROS generation and p38 activation contribute at least in part to Na-Mt-induced toxicity.

中文翻译：

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ROS 生成和 p-38 激活有助于蒙脱石诱导的体外和体内角膜毒性

蒙脱石（Mt）及其衍生物现已广泛应用于工业和生物医学领域。因此，对这些材料的安全评估对于保护暴露后的人体健康至关重要；然而，缺乏关于 Mt 眼部毒性的研究。特别是，Mt 的不同物理化学特性可能会极大地改变它们的毒理学潜力。为了探索这些特征对眼睛的影响，首次在体外和体内研究了五种类型的 Mt，并研究了它们的潜在机制。基于 ATP 含量、乳酸脱氢酶 (LDH) 渗漏、细胞形态和 Mt 在细胞中的分布分析，不同类型的 Mt 在人 HCEC-B4G12 角膜细胞中引起细胞毒性。在五种 Mt 类型中，Na-Mt 表现出最高的细胞毒性。尤其，Na-Mt 和壳聚糖修饰的酸性 Na-Mt (CH-Na-Mt) 在体内诱导眼毒性，表现为角膜损伤面积增加和凋亡细胞数量增加。Na-Mt 和 CH-Na-Mt 还在体外和体内诱导活性氧 (ROS) 的产生，如 2',7'-二氯荧光素二乙酸酯和二氢乙锭染色所示。此外，Na-Mt 激活丝裂原活化蛋白激酶信号通路。用 ROS 清除剂 N-乙酰半胱氨酸预处理 HCEC-B4G12 细胞可减弱 Na-Mt 诱导的细胞毒性并抑制 p38 活化，同时用 p38 特异性抑制剂抑制 p38 活化可降低 Na-Mt 诱导的细胞毒性。结果表明 Mt 在体外和体内诱导角膜毒性。Mt 的物理化学性质极大地影响其毒理学潜力。此外，