Nam Mi-Hyun, Dhillon Armaan, Nahomi Rooban B, Carrillo Noelle L, Hougen Clarinda S, Nagaraj Ram H
Department of Ophthalmology, UCHealth-Sue Anschutz-Rodgers Eye Centre, School of Medicine, University of Colorado, Aurora, CO, United States.
Department of Radiology, UCHealth University of Colorado Hospital, Aurora, CO, United States.
Front Cell Neurosci. 2024 Aug 1;18:1441924. doi: 10.3389/fncel.2024.1441924. eCollection 2024.
Neurovascular degeneration results in vascular dysfunction, leakage, ischemia, and structural changes that can lead to significant visual impairment. We previously showed the protective effects of peptain-1, a 20 amino acid peptide derived from the αB-crystallin core domain, on retinal ganglion cells in two animal models of glaucoma. Here, we evaluated the ability of peptain-1 to block apoptosis of human retinal endothelial cells (HRECs) and retinal capillary degeneration in mice subjected to retinal ischemia/reperfusion (I/R) injury.
HRECs were treated with either peptain-1 or scrambled peptides (200 μg/mL) for 3 h and a combination of proinflammatory cytokines (IFN-γ 20 ng/mL + TNF-α 20 ng/mL+ IL-1β 20 ng/mL) for additional 48 h. Apoptosis was measured with cleaved caspase-3 formation via western blot, and by TUNEL assay. C57BL/6J mice (12 weeks old) were subjected to I/R injury by elevating the intraocular pressure to 120 mmHg for 60 min, followed by reperfusion. Peptain-1 or scrambled peptide (0.5 μg) was intravitreally injected immediately after I/R injury and 7 days later. One microliter of PBS was injected as vehicle control, and animals were euthanized on day 14 post-I/R injury. Retinal capillary degeneration was assessed after enzyme digestion followed by periodic acid-Schiff staining.
Our data showed that peptain-1 entered HRECs and blocked proinflammatory cytokine-mediated apoptosis. Intravitreally administered peptain-1 was distributed throughout the retinal vessels after 4 h. I/R injury caused retinal capillary degeneration. Unlike scrambled peptide, peptain-1 protected capillaries against I/R injury. Additionally, peptain-1 inhibited microglial activation and reduced proinflammatory cytokine levels in the retina following I/R injury.
Our study suggests that peptain-1 could be used as a therapeutic agent to prevent capillary degeneration and neuroinflammation in retinal ischemia.
神经血管退变会导致血管功能障碍、渗漏、缺血以及结构改变，进而可能导致严重的视力损害。我们之前在两种青光眼动物模型中证明了源自αB-晶状体蛋白核心结构域的20个氨基酸的肽peptain-1对视网膜神经节细胞具有保护作用。在此，我们评估了peptain-1在视网膜缺血/再灌注（I/R）损伤小鼠中阻断人视网膜内皮细胞（HREC）凋亡和视网膜毛细血管退变的能力。
用peptain-1或乱序肽（200μg/mL）处理HREC 3小时，然后用促炎细胞因子组合（IFN-γ 20ng/mL + TNF-α 20ng/mL + IL-1β 20ng/mL）再处理48小时。通过蛋白质印迹法检测裂解的半胱天冬酶-3形成以及通过TUNEL检测法来测量细胞凋亡。12周龄的C57BL/6J小鼠通过将眼压升高至120mmHg持续60分钟，随后再灌注来进行I/R损伤。在I/R损伤后立即以及7天后玻璃体内注射peptain-1或乱序肽（0.5μg）。注射1微升PBS作为溶剂对照，在I/R损伤后第14天对动物实施安乐死。酶消化后进行高碘酸-希夫染色以评估视网膜毛细血管退变。
我们的数据表明peptain-1进入HREC并阻断促炎细胞因子介导的细胞凋亡。玻璃体内注射的peptain-1在4小时后分布于整个视网膜血管。I/R损伤导致视网膜毛细血管退变。与乱序肽不同，peptain-1保护毛细血管免受I/R损伤。此外，peptain-1抑制小胶质细胞活化并降低I/R损伤后视网膜中的促炎细胞因子水平。
我们的研究表明peptain-1可作为一种治疗剂来预防视网膜缺血中的毛细血管退变和神经炎症。
