Downie Laura E, Choi Janet, Lim Jeremiah K H, Chinnery Holly R
Invest Ophthalmol Vis Sci. 2016 Jun 1;57(7):3477-84. doi: 10.1167/iovs.15-19005.
We previously reported that applying toll-like receptor (TLR) ligands to an injured cornea induces corneal edema at 24 hours, which subsides by 1 week. We tested the hypotheses that endothelial expression of the tight-junction protein, zonula occludens-1 (ZO-1), would be altered during experimental sterile corneal inflammation and that endothelial cell density (ECD) would remain unaffected.
Anesthetized C57BL/6J mice received central 1-mm corneal abrasions followed by topical application of saline or cytosine-phosphate-guanosine oligodeoxynucleotide (CpG-ODN, TLR-9 agonist). At 24 hours, 1 week and 4 weeks post treatment, spectral-domain optical coherence tomography images were captured. Eyes were enucleated and processed for zonula occludens-1 (ZO-1) immunofluorescent staining. Corneal flatmounts were analyzed for endothelial ZO-1 expression, cell density, polymegethism, and polymorphism. Corneal stromal inflammatory cell infiltration was evaluated at 4 weeks by immunostaining for CD45.
Central corneal thickness (CCT) was increased in CpG-ODN treated eyes at 24 hours, had normalized by 1 week, but was again thickened by 4 weeks. In eyes with CpG-ODN, endothelial cell ZO-1 expression was reduced at 24 hours but returned to normal levels by 1 week. Endothelial cell density was not altered at 24 hours or 1 week. By 4 weeks, only CpG-ODN eyes showed relatively reduced ECD, as well as large numbers of CD45+ cells in the stroma. Changes to ECD correlated with CCT (r = -0.53, P < 0.01). Compared with naïve controls, more saline- and CpG-ODN-treated eyes exhibited polymegethism.
This study provides novel insights into the interplay between endothelial cell integrity, corneal edema, and chronic stromal leukocyte activation during sterile corneal inflammation in mice.
我们之前报道过，将 Toll 样受体（TLR）配体应用于受伤的角膜会在 24 小时时诱导角膜水肿，这种水肿在 1 周时消退。我们检验了以下假设：在实验性无菌性角膜炎症期间，紧密连接蛋白小带闭合蛋白 -1（ZO-1）的内皮表达会发生改变，而内皮细胞密度（ECD）将不受影响。
对麻醉的 C57BL/6J 小鼠进行中央 1 毫米角膜擦伤，然后局部应用生理盐水或胞嘧啶 - 磷酸 - 鸟嘌呤寡脱氧核苷酸（CpG-ODN，TLR-9 激动剂）。在治疗后 24 小时、1 周和 4 周时，采集光谱域光学相干断层扫描图像。摘除眼球并进行小带闭合蛋白 -1（ZO-1）免疫荧光染色处理。对角膜平铺片进行分析，以检测内皮 ZO-1 表达、细胞密度、大小不均一性和多形性。在 4 周时通过对 CD45 进行免疫染色评估角膜基质炎性细胞浸润情况。
CpG-ODN 处理的眼睛在 24 小时时中央角膜厚度（CCT）增加，1 周时恢复正常，但 4 周时再次增厚。在使用 CpG-ODN 的眼睛中，内皮细胞 ZO-1 表达在 24 小时时降低，但 1 周时恢复到正常水平。内皮细胞密度在 24 小时或 1 周时未改变。到 4 周时，只有 CpG-ODN 处理的眼睛显示 ECD 相对降低，并且基质中有大量 CD45 + 细胞。ECD 的变化与 CCT 相关（r = -0.53，P < 0.01）。与未处理的对照组相比，更多经生理盐水和 CpG-ODN 处理的眼睛表现出大小不均一性。
本研究为小鼠无菌性角膜炎症期间内皮细胞完整性、角膜水肿和慢性基质白细胞激活之间的相互作用提供了新的见解。
