Abstract
Introduction
To elucidate the specific functions of the primary cilia in corneal endothelial cells (CECs) by investigating the histological changes of corneal endothelium exposed at low temperature.
Study design
Experimental study.
Methods
This study involved corneas freshly obtained from Japanese white rabbits preserved in Optisol™-GS (Bausch & Lomb) corneal storage medium at 4 °C for 0, 1, and 7 days. Corneas preserved for 7 days were also incubated at 37 °C in culture media for an additional 2 days. A rabbit CEC line was also preserved in Optisol™-GS at 4 °C for 0 and 1 day. The corneal endothelium specimens and CECs were then assessed by immunostaining and scanning electron-microscopy (SEM).
Results
Immediately post isolation, the CECs of the specimens showed positive immunostaining for primary cilia (i.e., approximately 20%) via anti-acetylated alpha Tubulin antibody and SEM observation. Primary cilia were found to have attenuated/disappeared on the corneal endothelium specimens preserved for 1 or 7 days at 4 °C. After an additional 2-day incubation at 37 °C, primary cilia reappeared on the corneal endothelium specimens (approximately 20%). The disappearance of cilia during the preservation period was also observed in the immortalized CECs.
Conclusion
The findings in this study using rabbit corneas indicate that the primary cilia of corneal endothelium preserved at low temperature disappeared, then reappeared after returning to body temperature, suggesting that temperature has a direct effect on the primary cilia of corneal endothelium.
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Acknowledgements
The authors wish to thank John Bush for reviewing the manuscript and Hideto Deguchi, MD for conducting additional experiments on the 34 °C organ culture. This work was supported by Grants-in-Aid for Scientific Research from the Japan Society for the Promotion of Science (JSPS) (Grant No. JP90171834 and No. JP20K09832).
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H. Tanioka, Employee (Santen); K. Shinomiya, Employee (Santen); S. Kinoshita, None; C. Sotozono, None.
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Corresponding Author: Hidetoshi Tanioka
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10384_2022_933_MOESM2_ESM.pptx
Supplemental Figure 2 The primary cilia of rabbit corneal endothelium preserved in organ culture media at 34 °C. a The corneal endothelium immediately after 7-day preservation in organ culture media. Endothelium showed anti-acetylated alpha Tubulin-positive primary cilia. b Arrows indicate the primary cilia at a. The fluorescein-stained specimens were inspected via the use of a confocal laser microscope (FV1200IX83; OLYMPUS, Tokyo, Japan), and confocal laser-scanning microscopy images were captured. Scale Bar = 50 μm (PPTX 729 KB)
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Tanioka, H., Shinomiya, K., Kinoshita, S. et al. Temperature effects on the disappearance and reappearance of corneal-endothelium primary cilia. Jpn J Ophthalmol 66, 481–486 (2022). https://doi.org/10.1007/s10384-022-00933-0
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DOI: https://doi.org/10.1007/s10384-022-00933-0