Abstract
Purpose :
Previous studies reported that a reduction in pore density occurs in eyes with primary open angle glaucoma (POAG), indicating that this reduction may contribute to the elevated outflow resistance characteristic of this disease. This study aimed to determine, under flow conditions, whether release of connections between Schlemm’s canal (SC) inner wall (IW) cells and underlying juxtacanalicular connective tissue (JCT) cells plays a role in GV formation; and whether decreased amount of cell margin overlap between adjacent IW cells promotes paracellular pore formation using advanced 3D EM reconstruction.
Methods :
Normal human eyes (N=3) were either immersion-fixed (0mmHg) or perfusion-fixed (15mmHg). Frontal and radial sections of SC were processed for serial block-face scanning electron microscopy (SBF-SEM). The IW and JCT cells, GVs, and pores were 3D-reconstructed. Morphometric analyses were performed. In each IW cell, total number of connections with underlying JCT cells was determined. Total cell margin length (ML) and zero-overlap length (ZL) of each IW cell were measured to calculate percent zero-overlap length (PZL=ZL/ML). All data was compared between the eyes fixed at 0 and 15mmHg using Student’s t-test.
Results :
Total number of connections between each IW cell with underlying JCT cells significantly decreased in the eye fixed at 15mmHg (33±5, N=4 cells) compared to the eyes fixed at 0mmHg (180±12, N=5 cells, p<0.001). The maximum volume of GVs significantly increased in the eye fixed at 15mmHg (199.05±12.11 µm3) compared to those fixed at 0mmHg (74.12±27.54 µm3, p<0.01). All intracellular pores were found only in the thinnest regions of GV walls. The amount of overlap between adjacent IW cells significantly decreased (0.52±0.19µm vs. 3.71±0.20µm, p<0.0001), and PZL increased 26.68% (p=0.001) in the eye fixed at 15mmHg vs. those fixed at 0mmHg. All paracellular pores were found only in regions where the overlap length was zero µm.
Conclusions :
Our innovative use of SBF-SEM permits 3D ultrastructural analyses of the two types of cellular connections between IW/JCT and IW/IW cells. Our data suggests that these two types of connections each play a role in GV and pore formation in normal human eyes under flow conditions. Results of this study provide a baseline for future comparison with POAG eyes.
This is an abstract that was submitted for the 2016 ARVO Annual Meeting, held in Seattle, Wash., May 1-5, 2016.