Abstract
Purpose :
Abnormality of the cornea can lead to blindness and vision loss. The corneal endothelial cells (CECs) play a significant role in maintaining corneal function. We developed a multi-scale imaging workflow to visualize the mouse cornea at the microscale and the underlining subcellular variations of CECs in corneal flatmount at the nanoscale. We used a visible-light optical coherence tomography (vis-OCT) to image a mouse model of intraocular hypertension in vivo and revealed the nanoscopic cytoskeleton changes in the CECs using single-molecule localization microscopy (SMLM).
Methods :
The left eyes of wild-type mice were maintained at 40 mmHg intraocular pressure (IOP) for 1 hour using anterior chamber puncture. The right eyes were used as controls without any procedures. The vis-OCT images of cornea were acquired using a home-built vis-OCT system. The mice were sacrificed and the cornea was isolated, fixed, and immunostained to visualize the ZO-1 tight junction structure variations using indirect immunofluorescence labeling. The cornea sample was flat-mounted for SMLM imaging in a commercially available STORM imaging buffer in a home-built SMLM system.
Results :
The vis-OCT images of corneal tissues revealed the typical edema after maintaining 40 mmHg IOP in the epithelium and stroma layers of the cornea. The SMLM images showed characteristic hexagonal CECs and revealed significant changes in tight junction proteins’ filament density and thickness after exposure to high IOP. Such disruptions of cell tight junctions are likely related to the blurred vision in diseases associated with elevated IOP. Further, it reveals the native cell responses in tissue that would be challenging for imaging cell cultures.
Conclusions :
We developed a multi-scale imaging workflow to correlate microscopic variations in corneal tissue and nanoscopic variations in CECs of corneal flatmount in response to high IOP in mouse models. These results demonstrate the utility of multi-scale imaging for studying tissue- and molecular level variations for corneal diseases and glaucoma.
This is a 2021 ARVO Annual Meeting abstract.