Vision CRC, Sydney, Australia ; School of Optometry and Vision Science, University of New South Wales, Sydney, Australia.
PLoS One. 2013 Aug 19;8(8):e71948. doi: 10.1371/journal.pone.0071948. eCollection 2013.
This study investigated ocular surface components that contribute to matrix-metalloproteinase (MMP)-2 and MMP-9 found in tears following corneal epithelial wounding.
Laboratory short-haired cats underwent corneal epithelial debridement in one randomly chosen eye (n = 18). Eye-flush tears were collected at baseline and during various healing stages. Procedural control eyes (identical experimental protocol as wounded eyes except for wounding, n = 5) served as controls for tear analysis. MMP activity was analyzed in tears using gelatin zymography. MMP staining patterns were evaluated in ocular tissues using immunohistochemistry and used to determine MMP expression sites responsible for tear-derived MMPs.
The proMMP-2 and proMMP-9 activity in tears was highest in wounded and procedural control eyes during epithelial migration (8 to 36 hours post-wounding). Wounded eyes showed significantly higher proMMP-9 in tears only during and after epithelial restratification (day 3 to 4 and day 7 to 28 post-wounding, respectively) as compared to procedural controls (p<0.05). Tears from wounded and procedural control eyes showed no statistical differences for pro-MMP-2 and MMP-9 (p>0.05). Immunohistochemistry showed increased MMP-2 and MMP-9 expression in the cornea during epithelial migration and wound closure. The conjunctival epithelium exhibited highest levels of both MMPs during wound closure, while MMP-9 expression was reduced in conjunctival goblet cells during corneal epithelial migration followed by complete absence of the cells during wound closure. The immunostaining for both MMPs was elevated in the lacrimal gland during corneal healing, with little/no change in the meibomian glands. Conjunctival-associated lymphoid tissue (CALT) showed weak MMP-2 and intense MMP-9 staining.
Following wounding, migrating corneal epithelium contributed little to the observed MMP levels in tears. The major sources assessed in the present study for tear-derived MMP-2 and MMP-9 following corneal wounding are the lacrimal gland and CALT. Other sources included stromal keratocytes and conjunctiva with goblet cells.
本研究调查了角膜上皮损伤后泪液中基质金属蛋白酶（MMP）-2 和 MMP-9 的眼表成分。
实验室短毛猫随机选择一只眼进行角膜上皮清创术（n=18）。在基线和各种愈合阶段采集眼冲洗液。程序对照眼（与受伤眼相同的实验方案，但不受伤，n=5）作为泪液分析的对照。使用明胶酶谱法分析泪液中的 MMP 活性。使用免疫组织化学评估眼组织中的 MMP 染色模式，并用于确定负责泪源性 MMP 的 MMP 表达部位。
受伤眼和程序对照眼的 proMMP-2 和 proMMP-9 活性在上皮迁移期间（受伤后 8 至 36 小时）最高。与程序对照相比，受伤眼仅在上皮再分层期间和之后（分别为受伤后第 3 至 4 天和第 7 至 28 天）显示出更高的泪液 proMMP-9（p<0.05）。受伤眼和程序对照眼的 pro-MMP-2 和 MMP-9 之间无统计学差异（p>0.05）。免疫组织化学显示上皮迁移和伤口闭合过程中角膜中 MMP-2 和 MMP-9 的表达增加。在伤口闭合期间，结膜上皮表达最高水平的两种 MMP，而在角膜上皮迁移过程中结膜杯状细胞中的 MMP-9 表达减少，随后在伤口闭合过程中完全消失。在角膜愈合过程中，泪腺的免疫染色均升高，而睑板腺几乎没有变化。结膜相关淋巴组织（CALT）显示 MMP-2 弱阳性和 MMP-9 强阳性染色。
受伤后，迁移的角膜上皮对泪液中观察到的 MMP 水平贡献不大。本研究评估的角膜创伤后泪源性 MMP-2 和 MMP-9 的主要来源是泪腺和 CALT。其他来源包括基质成纤维细胞和结膜伴杯状细胞。
