A new method of detecting changes in corneal health in response to toxic insults.

The size and arrangement of stromal collagen fibrils (CFs) influence the optical properties of the cornea and hence its function. The spatial arrangement of the collagen is still questionable in relation to the diameter of collagen fibril. In the present study, we introduce a new parameter, edge-fibrillar distance (EFD) to measure how two collagen fibrils are spaced with respect to their closest edges and their spatial distribution through normalized standard deviation of EFD (NSDEFD) accessed through the application of two commercially available multipurpose solutions (MPS): ReNu and Hippia. The corneal buttons were soaked separately in ReNu and Hippia MPS for five hours, fixed overnight in 2.5% glutaraldehyde containing cuprolinic blue and processed for transmission electron microscopy. The electron micrographs were processed using ImageJ user-coded plugin. Statistical analysis was performed to compare the image processed equivalent diameter (ED), inter-fibrillar distance (IFD), and EFD of the CFs of treated versus normal corneas. The ReNu-soaked cornea resulted in partly degenerated epithelium with loose hemidesmosomes and Bowman's collagen. In contrast, the epithelium of the cornea soaked in Hippia was degenerated or lost but showed closely packed Bowman's collagen. Soaking the corneas in both MPS caused a statistically significant decrease in the anterior collagen fibril, ED and a significant change in IFD, and EFD than those of the untreated corneas (p<0.05, for all comparisons). The introduction of EFD measurement in the study directly provided a sense of gap between periphery of the collagen bundles, their spatial distribution; and in combination with ED, they showed how the corneal collagen bundles are spaced in relation to their diameters. The spatial distribution parameter NSDEFD indicated that ReNu treated cornea fibrils were uniformly distributed spatially, followed by normal and Hippia. The EFD measurement with relatively lower standard deviation and NSDEFD, a characteristic of uniform CFs distribution, can be an additional parameter used in evaluating collagen organization and accessing the effects of various treatments on corneal health and transparency.

Swelling studies of camel and bovine corneal stroma.

In the present study we investigated the swelling characteristics of fresh camel and bovine cornea in sodium salt solutions. Swelling studies were carried out at 20 minutes, 14 hours, and 46 hours on five fresh camel and 5 five fresh bovine corneas. During the 20-minute hydration of fresh corneal stroma was investigated using sodium chloride (NaCl), sodium bicarbonate (NaHCO(3)), sodium acetate (CH(3)COONa), sodium thiocyanate (NaSCN), and sodium floride (NaF) at 2-minute time intervals. During a 46-hour time period, the hydration study was carried out using NaCl (150, 300 mM) and NaF (150 mM) at random intervals. The 14-hour study was carried out to assess the rehydration of corneal stroma after 6 hours of drying. During the 20-minute swelling studies in the first 2 minutes the rate of hydration in both camel and bovine corneas was high but gradually reduced in the 2-20-minute period. The rates and levels of hydration of camel and bovine cornea were not significantly different from each other in all the strengths of solutions. During the 46-hour swelling studies, the initial rate of hydration (0-2 hours) of camel and bovine stroma, in all solutions was significantly higher (Z = 0.056) compared to hydration during later hours (2-46 hours). Camel stromal hydration (high) in 150 mM NaCl was significantly higher compared to bovine stromal hydration in the same solution during the 10-24, and 24-46-hour time periods. Rehydration in camel stroma was significantly lower than bovine in 150 mM NaF. The 20-minute study showed that there was no selective affinity for particular ions in camel or bovine corneal stroma. Initial swelling in both corneal and bovine stroma is faster and more prominant compared to later swelling. The swelling in camel cornea is more prominant compared to bovine corneal stroma. This could be due to higher negatively charged keratin sulfate-proteoglycans in the stroma. Lower rehydration in camel cornea suggests stronger leaching of proteoglycans from stroma in NaF.