Influence of temperature on corneas stored in culture medium. A comparative study using functional and morphological methods

Abstract

ABSTRACT.Purpose: To investigate the influence of storage temperature on corneal swelling and on endothelial morphology in cultured corneas. Material and methods: Forty-eight rabbit corneas were separated into four groups of 12. The corneas were stored in culture medium at 37 (group 37), 34 (group 34), 31 (group 31) and 23 (room temperature) (group 23), respectively. All the corneas were monitored by weight recordings on days 0, 1, 2, 3, 4 and 7. On day 7, corneas were prepared for scanning electron microscopy and endothelial cell counts were performed. Results: Lowering the temperature of the culture medium resulted in less swelling. Both temperature and storage time had significant effects on corneal swelling (p < 0.001). On day 7, the observed mean weight increase was 131.2%, 143.0%, 172.5% and 199.7% in groups 23, 31, 34 and 37, respectively. The estimated mean daily weight increase for the corneas were 2.6%, 4.0%, 9.1% and 16.0% in groups 23, 31, 34 and 37, respectively. Scanning electron microscopy showed an intact endothelial layer in all groups after 7 days and there were no statistically significant differences in endothelial counts between groups 23, 31 and 34. In group 37, the cell borders were difficult to distinguish after 7 days and no meaningful count could be performed. Conclusions: The swelling rate of cultured corneas is significantly less at 23 and 31than it is at 34 and 37 during the first week. This is most likely the result of a greatly increased barrier effect at lower temperatures. Whereas weight recording revealed profound differences between the groups, scanning electron microscopy and endothelial cell counting did not. The results support the hypothesis that storage at 37is not optimal in culturing corneas. Lowering the temperature below body temperature, and even lower than 31, results in less corneal swelling.

Ex Vivo Adenovirus-Mediated Gene Transfer to Corneal Graft Endothelial Cells in Mice

“…Secondly, 4°C is the standard temperature for human corneal storage in eye banks; indeed many investigators have established that temperatures of 31°C or lower are appreciably better than higher temperatures (including 37°C) for preserving corneal endothelial morphology and function-without which the transfection would not have been successful. 30 In the aggregate therefore, we believe that the lower temperatures are far preferable for gene transfer to the corneal endothelium ex vivo. Our findings provide relevant information for delivery of exogenous gene(s) during corneal storage of eye bank tissue similar to the effective gene transfer that has been shown during cold organ preservation for other solid organ transplants, such as liver and heart.…”

A novel method for preserving cultured limbal epithelial cells

“…A few reports have been published that consider the influence of room temperature (23-25˚C) on corneas stored in culture media such as McCarey-Kaufman medium, 12 K-Sol medium, 13 TC 199 medium 14 and RPMI 1640 organ culture medium. 15 However, in these studies, the corneal endothelium has been the main focus of attention.…”

Upregulation of chemokine expression in murine cornea due to mechanical trauma or endotoxin

“…We have compared RPMI containing 10% FCS at 37°C (more extreme than conditions in general use) with Optisol at 4°C. We found that there was some upregulation of mRNA encoding chemokines using both storage conditions; however it was more marked following incubation at 37°C, which may not be optimal for corneal organ culture 14. The chemokines IP-10, RANTES and MIP2 showed the highest level of upregulation.…”

“…At present, there are two methods used for ex vivo corneal storage: storage in a dextran and chondroitin sulphate-based medium at 4°C or organ culture 14. Typically corneas are organ-cultured at 31–34°C in Eagle’s MEM containing 2–8% fetal bovine serum.…”