Abstract
Abstract: :
Purpose: To test the hypothesis that within 3D reconstructions of the monkey ONH, detectable changes in the size and shape of Bruch’s membrane opening occur early in experimental glaucoma. Methods: Both eyes of three young adult monkeys (one eye given early glaucoma (EG) via a moderate, laser–induced IOP elevation, then followed to the onset of CSLT–detected ONH surface change) were set to an IOP of 10 mm Hg for 1 hour. Each monkey was than perfusion–fixed to isolate the plastic (permanent) deformation of the ONH connective tissues. The trephinated ONH and peripapillary sclera from each eye was embedded in paraffin, and microtome serial sectioned at 3 µm thickness, with the embedded tissue block face stained and imaged after each cut. The images were then aligned and stacked to form a 3D volumetric reconstruction of the ONH connective tissues (2.5 x 2.5 x 3.0 µm/voxel). Custom software served radial voxel data planes from the 3D volume at 4.5° intervals, and the termination of Bruch’s membrane within the scleral canal was marked within each data plane (a total of 80 points per opening). Each ONH was marked on three separate days to assess observer variability. The coordinates of the data points for each measurement of each ONH were analyzed with custom MATLAB routines which fitted a least–squares ellipse to the opening in Bruch’s membrane. The major (vertical) axis, minor (horizontal) axis, and the area of the fitted ellipses were calculated and compared by ANOVA. Results: Observer variability was not significant (p>0.05) for any parameter within any animal. The mean area of Bruch’s membrane opening (BMO) in the EG eyes was 24% larger than in the normal (N) eyes (p<0.001). The major (vertical) axis of the ellipse fitted to BMO in the EG eyes was 23% larger than in the normal eyes (p<0.001). There was no overall significant difference between EG and N eyes in the minor (horizontal) axis. Individually, all three monkeys demonstrated similar enlargement of the area and elongation of the vertical axis of the BMO in their EG eyes compared to their contralateral normal eyes (p<0.01). In monkey 1, the horizontal axis of BMO increased by 8% in the EG eye (p<0.01), while in monkey 3 the horizontal axis decreased by 11% in the EG eye (p<0.01). Conclusions: At the onset of CSLT–detected ONH surface change in young adult monkey eyes, the clinically visible opening in Bruch’s membrane is vertically elongated by histologic measurement. Clinical detection of this change may be a marker of early glaucomatous connective tissue damage.
Keywords: Bruch's membrane • optic disc • imaging/image analysis: non–clinical