McLeod D Scott, Bhutto Imran, Edwards Malia M, Gedam Manasee, Baldeosingh Rajkumar, Lutty Gerard A
Wilmer Ophthalmological Institute, Johns Hopkins Hospital, Baltimore, Maryland, United States.
Invest Ophthalmol Vis Sci. 2017 Nov 1;58(13):5887-5896. doi: 10.1167/iovs.17-22989.
Our previous study demonstrated significantly more degranulating mast cells (MCs) in choroids from subjects with age-related macular degeneration compared to aged controls. This study examined the immunolocalization of tryptase, the most abundant MC secretory granule-derived serine protease, in aged control eyes and eyes with geographic atrophy (GA).
Postmortem human eyes with and without GA were obtained from the National Disease Research Interchange. Tissue was fixed, cryopreserved, sectioned, and immunostained with a monoclonal antibody against tryptase. Sections were imaged on a Zeiss 710 Confocal Microscope.
In the posterior pole of all aged control eyes, tryptase was confined to choroidal MCs, which were located primarily in Sattler's layer. In eyes with GA, many MCs were located in the inner choroid near choriocapillaris and Bruch's membrane (BM). Tryptase was found not only in MCs but also diffusely around them in stroma, suggesting they had degranulated. In contrast with aged control eyes, eyes with GA also had strong tryptase staining in BM. Tryptase was observed within BM in regions of RPE atrophy, at the border of atrophy, and extending well into the nonatrophic region.
Our results demonstrate that tryptase, released during choroidal MC degranulation, binds to BM in GA in advance of RPE atrophy. Tryptase activates MMPs that can degrade extracellular matrix (ECM) and basement membrane components found in BM. ECM modifications are likely to have a profound effect on the function and health of RPE and choroidal thinning in GA.
我们之前的研究表明，与老年对照组相比，年龄相关性黄斑变性患者脉络膜中脱颗粒肥大细胞（MCs）显著更多。本研究检测了老年对照眼和地图样萎缩（GA）眼中，肥大细胞中最丰富的分泌颗粒衍生丝氨酸蛋白酶——组织蛋白酶的免疫定位。
从国家疾病研究交流中心获取有或无GA的死后人类眼球。组织经固定、冷冻保存、切片，并用抗组织蛋白酶的单克隆抗体进行免疫染色。切片在蔡司710共聚焦显微镜下成像。
在所有老年对照眼的后极部，组织蛋白酶局限于脉络膜MCs，主要位于萨特勒层。在GA眼中，许多MCs位于靠近脉络膜毛细血管和布鲁赫膜（BM）的脉络膜内层。不仅在MCs中发现了组织蛋白酶，在其周围的基质中也有弥散分布，提示它们已经脱颗粒。与老年对照眼相比，GA眼中BM也有强烈的组织蛋白酶染色。在RPE萎缩区域、萎缩边界以及延伸至非萎缩区域的BM内均观察到组织蛋白酶。
我们的结果表明，脉络膜MC脱颗粒过程中释放的组织蛋白酶，在RPE萎缩之前就与GA中的BM结合。组织蛋白酶激活基质金属蛋白酶（MMPs），后者可降解BM中的细胞外基质（ECM）和基底膜成分。ECM修饰可能对GA中RPE的功能和健康以及脉络膜变薄产生深远影响。
