Effect of hydration on the ultra‐structural distribution of collagen fibrils and proteoglycans in human corneal stroma

Abstract Purpose We investigated the ultra‐structural changes in the distribution of collagen fibrils (CF) and proteoglycans (PGs) due to hydration in the human corneal stroma. Methods Six fresh human scleral corneal rings were hydrated separately in deionized water for 2hrs and 48hrs. Corneas were divided into two parts. The first part of each cornea was fixed in 2.5% glutaraldehyde containing cuprolinic blue in sodium acetate buffer and processed for electron microscopy to assess proteoglycans. A second part of each cornea was fixed in 2.5% glutaraldehyde in 0.1M phosphate buffer and post fixed in 1% osmium tetroxide to assess collagen fibril diameter. Results In the hydrated corneas the mean diameter of CF in the anterior stroma (23.40±2.61nm), middle stroma (22.48±3.28nm) and posterior stroma (23.58±2.46nm) was less than the CF diameter in the anterior (28.27±4.05nm), middle (28.22±3.09nm) and posterior stroma (29.73±3.72nm) of the normal cornea. Micro‐fibrils within the CF in the hydrated cornea were degenerated. In the hydrated cornea, the mean area of the PGs in the anterior stroma was reduced whereas it increased in the middle stroma compared to those in the normal cornea. The density of the PGs in the hydrated cornea was reduced in the anterior, middle and posterior stroma compared to those in the normal cornea. Conclusion The effect of hydration on the uniform organisation of CF and PGs was higher in the middle and posterior stroma than in the anterior stroma. This could be due to less penetration of water in the interlacing lamellae of the anterior stroma. The reduction in the collagen fibrils diameter could be due to degeneration of microfibrils within the collagen fibrils.