Nguyen Cathy, Midgett Dan, Kimball Elizabeth C, Steinhart Matthew R, Nguyen Thao D, Pease Mary E, Oglesby Ericka N, Jefferys Joan L, Quigley Harry A
The Glaucoma Center of Excellence, Wilmer Ophthalmological Institute, Baltimore, Maryland, United States.
Department of Mechanical Engineering, Johns Hopkins University, Baltimore, Maryland, United States.
Invest Ophthalmol Vis Sci. 2017 Feb 1;58(2):721-733. doi: 10.1167/iovs.16-20620.
To develop an ex vivo explant system using multiphoton microscopy and digital volume correlation to measure the full-field deformation response to intraocular pressure (IOP) change in the peripapillary sclera (PPS) and in the optic nerve head (ONH) astrocytic structure.
Green fluorescent protein (GFP)-glutamate transporter-GLT1 (GLT1/GFP) mouse eyes were explanted and imaged with a laser-scanning microscope under controlled inflation. Images were analyzed for regional strains and changes in astrocytic lamina and PPS shape. Astrocyte volume fraction in seven control GLT1/GFP mice was measured. The level of fluorescence of GFP fluorescent astrocytes was compared with glial fibrillary acidic protein (GFAP) labeled astrocytes using immunohistochemistry.
The ONH astrocytic structure remained stable during 3 hours in explants. Control strain-globally, in the central one-half or two-thirds of the astrocytic lamina-was significantly greater in the nasal-temporal direction than in the inferior-superior or anterior-posterior directions (each P≤ 0.03, mixed models). The PPS opening (perimeter) in normal eye explants also became wider nasal-temporally than superior-inferiorly during inflation from 10 to 30 mm Hg (P = 0.0005). After 1 to 3 days of chronic IOP elevation, PPS area was larger than in control eyes (P = 0.035), perimeter elongation was 37% less than controls, and global nasal-temporal strain was significantly less than controls (P = 0.007). Astrocyte orientation was altered by chronic IOP elevation, with processes redirected toward the longitudinal axis of the optic nerve.
The explant inflation test measures the strain response of the mouse ONH to applied IOP. Initial studies indicate regional differences in response to both acute and chronic IOP elevation within the ONH region.
开发一种离体组织块系统，利用多光子显微镜和数字体积相关技术来测量视乳头周围巩膜（PPS）和视神经乳头（ONH）星形细胞结构对眼内压（IOP）变化的全场变形反应。
将绿色荧光蛋白（GFP）-谷氨酸转运体-GLT1（GLT1/GFP）小鼠的眼睛取出并在控制充气的情况下用激光扫描显微镜成像。分析图像以获取区域应变以及星形细胞层和PPS形状的变化。测量了7只对照GLT1/GFP小鼠的星形细胞体积分数。使用免疫组织化学方法将GFP荧光星形细胞的荧光水平与胶质纤维酸性蛋白（GFAP）标记的星形细胞进行比较。
在离体组织中，ONH星形细胞结构在3小时内保持稳定。对照应变——整体上，在星形细胞层的中央二分之一或三分之二区域——在鼻颞方向上显著大于上下方向或前后方向（各P≤0.03，混合模型）。在从10至30 mmHg充气过程中，正常眼离体组织中的PPS开口（周长）在鼻颞方向上也比上下方向更宽（P = 0.0005）。在慢性眼压升高1至3天后，PPS面积大于对照眼（P = 0.035），周长伸长比对照少37%，整体鼻颞应变显著小于对照（P = 0.007）。慢性眼压升高会改变星形细胞的方向，其突起重新指向视神经的纵轴。
离体组织充气试验可测量小鼠ONH对施加的IOP的应变反应。初步研究表明，ONH区域内对急性和慢性眼压升高的反应存在区域差异。
