Objectives. In glaucomatous eyes, the main aqueous humor (AH) outflow pathway is damaged by accumulated oxidative stress arising from the microenvironment, vascular dysregulation, and aging, which results in increased outflow resistance and ocular hypertension. Schlemm’s canal (SC) serves as the final filtration barrier of the main AH outflow pathway. The present study is aimed at investigating the possible regulation of vasoactive intestinal peptide (VIP) on the cytoskeleton by stabilizing ZO-1 in SC. Methods. Model of chronic ocular hypertension (COH) induced by episcleral venous cauterization was treated with topical VIP. The ultrastructure of junctions, ZO-1 levels, and permeability of the SC inner wall to FITC-dextran (70 kDa) were detected in the COH models. The F-actin distribution, F/G-actin ratio, and ZO-1 degradation pathway in human umbilical vein endothelial cells (HUVECs) and HEK 293 cells were investigated. Results. ZO-1 in the outer wall of the SC was less than that in the inner wall. COH elicited junction disruption, ZO-1 reduction, and increased permeability of the SC inner wall to FITC-dextran in rats. ZO-1 plays an essential role in maintaining the F/G-actin ratio and F-actin distribution. VIP treatment attenuated the downregulation of ZO-1 associated with COH or H₂O₂-induced oxidative damage. In H₂O₂-stimulated HUVECs, the caspase-3 inhibitor prevents ZO-1 disruption. Caspase-3 activation promoted endolysosomal degradation of ZO-1. Furthermore, a decrease in caspase-3 activation and cytoskeleton redistribution was demonstrated in VIP + H₂O₂-treated cells. The knockdown of ZO-1 or the overexpression of caspase-3 blocked the effect of VIP on the cytoskeleton. Conclusion. This study provides insights into the role of VIP in stabilizing the interaction between the actin cytoskeleton and cell junctions and may provide a promising targeted strategy for glaucoma treatment.

中文翻译：

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VIP 通过减少 Caspase-3 介导的 ZO-1 内溶酶体降解来稳定施累姆管内皮细胞的细胞骨架

目标。在青光眼眼中，主要的房水流出通路被微环境、血管失调和衰老引起的累积氧化应激破坏，导致流出阻力增加和高眼压。施累姆管 (SC) 是主要 AH 流出通路的最终过滤屏障。本研究旨在通过稳定 SC 中的 ZO-1 来研究血管活性肠肽 (VIP) 对细胞骨架的可能调节作用。方法. 用局部VIP治疗由巩膜外静脉烧灼引起的慢性高眼压（COH）模型。在 COH 模型中检测到连接的超微结构、ZO-1 水平和 SC 内壁对 FITC-葡聚糖 (70 kDa) 的渗透性。研究了人脐静脉内皮细胞 (HUVECs) 和 HEK 293 细胞中的 F-肌动蛋白分布、F/G-肌动蛋白比率和 ZO-1 降解途径。结果。SC外壁的ZO-1小于内壁。COH 在大鼠中引起连接中断、ZO-1 减少和 SC 内壁对 FITC-葡聚糖的渗透性增加。ZO-1 在维持 F/G-肌动蛋白比率和 F-肌动蛋白分布中起重要作用。_{VIP 治疗减弱了与 COH 或 H 2} O相关的 ZO-1 的下调_2-诱导的氧化损伤。在 H ₂ O ₂刺激的 HUVEC 中，caspase-3 抑制剂可防止 ZO-1 破坏。Caspase-3 激活促进了 ZO-1 的内溶酶体降解。此外，在 VIP + H ₂ O ₂处理的细胞中证实了 caspase-3 活化和细胞骨架再分布的减少。ZO-1 的敲低或 caspase-3 的过表达阻断了 VIP 对细胞骨架的影响。结论。这项研究提供了对 VIP 在稳定肌动蛋白细胞骨架和细胞连接之间相互作用中的作用的见解，并可能为青光眼治疗提供有希望的靶向策略。