A physiological perspective on the swelling properties of the mammalian corneal stroma

Abstract

Purpose: The present studies were designed to assess whether measurement of corneal stroma swelling in the laboratory, especially in non-physiological solutions, was associated with a measurable effect on the keratocytes.

Methods: Complete corneal stroma preparations were made from quality- and age-selected recent post-mortem cattle eyes. These were either assessed immediately or incubated in three different solutions, namely a balanced salts solution with glucose (BSSG), isotonic phosphate buffered saline (PBS) or pure water. Incubations were carried out at 37 degrees C for 9h, and repeated measures of wet mass made so that the rates and extent of swelling could be determined. After incubation, an aqueous extract was made of the stroma for measurements of the levels the enzymes lactate dehydrogenase (LDH), aldehyde dehydrogenase (ALDH) and N-acetyl-beta-glucosaminidase.

Results: The initial rates of swelling were lowest in BSSG, marginally faster in PBS and much faster in water. The secondary rates of swelling showed the same sequence being 10.0%/h in BSSG, 14.8%/h in PBS and 34.2%/h in water. Compared to non-incubated preparations, reductions in all three enzyme activities occurred. For LDH, these were 15% with BSSG, 40% in PBS and 80% with water. Similar results were seen with ALDH activity when comparing the three incubation solutions, while incubation in BSSG also resulted in a substantial (40%) reduction in N-acetyl-glucosaminidase activity.

Conclusions: When immersed in an isotonic BSSG with added glucose at 37 degrees C, the swelling of a complete bovine corneal stroma is much less than smaller pieces of stroma, and also slightly less than if isotonic PBS was used. With the use of BSSG, little or no change in cytoplasmic enzyme activities occurred, but measurable decreases were noted with PBS and very substantial decreases when water was used, indicating a toxic effect on the keratocytes. The observation that substantial decreases in a lysosomal enzyme activity could occur even with the use of BSSG indicate substantial stress is imposed on the stroma during these types of experiments. Notwithstanding, the data collectively indicate that the keratocyte cells within the collagen matrix of the stroma can be substantially damaged and this needs to be taken into account in future experiments on the true physiology of the corneal stroma.