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Johan R. Germundsson, Per Fagerholm, Neil S. Lagali; Comparison Of Bowman’s Layer Thickness And Morphology By In Vivo And Ex Vivo Techniques. Invest. Ophthalmol. Vis. Sci. 2011;52(14):305.
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To assess Bowman’s layer (BL) by in vivo confocal microscopy (IVCM) and by light microscopy in the same tissue samples, to compare anatomical thickness and pathologic morphology between the two methods.
Following ethical approval and informed consent, we examined 17 consecutive corneal transplant patients preoperatively by IVCM. Removed, pathologic corneal buttons were stored in 2% glutaraldehyde then prepared and cut in 4µm sections for light microscopy. 13 of 17 corneas were suitable for in vivo and ex vivo comparison. 7 of 13 of the prepared corneas had a uniformly thick BL in light microscopy and were selected for in vivo/ex vivo thickness comparison. BL thickness was measured between 4 to 12 times per cornea at various central locations by light microscopy. 5 to 7 IVCM scans through BL from each cornea were selected, from which BL thickness was measured by two observers. We then compared the thickness of BL by the two methods. The remaining 6 of 13 corneas showed abnormal morphology in BL by light microscopy and were further characterized and compared with the IVCM images.
The 95% limits of agreement for interobserver differences in BL thickness measurement was ± 11.5% for light microscopy and ± 11.9% for IVCM measurement. The mean BL thickness in a given central cornea ranged from 7.8-12.8µm by light microscopy and from 10.4-14.2µm by IVCM. BL thickness in vivo was significantly greater than in tissue sections (p< 0.017). In the same cornea, in vivo BL thickness was significantly greater than ex vivo values in 5 of 7 cases (p<0.016). No significant correlation, however, between in vivo and ex vivo thickness could be found in our small series. Of the 6 corneas with abnormal BL morphology, 4 had stromal cells, stromal tissue or scar tissue anterior to BL, 3 had focal absences, thinning, or folding of BL, and 1 had an abnormal-appearing basement membrane by light microscopy. Interestingly, these morphologic findings were also observed in vivo, in an en face orientation.
We believe this to be the first report to compare BL thickness in vivo and ex vivo in the same tissue samples. The thickness of BL depended on the method used, with the in vivo measurements generally yielding a larger value. This study is important as there is very little data on BL thickness in vivo, and laser surgery done today is based on assumed BL thickness values. Additionally, correlation of BL morphology in vivo with light microscopy sections could help us to identify signs of pathology in patients in the future by in vivo observation alone.
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