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R Ickes, DL Harris, NC Joyce; "Classical" Cadherin Expression in Corneal Endothelium . Invest. Ophthalmol. Vis. Sci. 2002;43(13):3190.
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© ARVO (1962-2015); The Authors (2016-present)
Purpose: The "classical cadherins, E-(epithelial), N-(neuronal), P-(placental), and R-(retinal) cadherin, mediate cell contact in many cell types. The current studies investigated "classical" cadherin isoform expression and localization in rat and human corneal endothelium and form the basis for future study of the mechanisms of contact-dependent inhibition of proliferation in these cells. Methods: Corneas from adult Sprague-Dawley rats and donor human corneas from National Disease Research Interchange were used in these studies. Expression of E, N, P and R cadherins was analyzed in transverse corneal sections via immunocytochemistry (ICC) using established protocols. Whole mounts of donor human and rat corneas were utilized for N-cadherin localization. For both sudies, secondary antibody alone was used as a negative control. All tissues were mounted in medium containing propidium iodide to detect nuclei. Staining was visualized by fluorescence confocal microscopy. The kinetics of N-cadherin protein expression were studied in rat corneal endothelial cell cultures. Primary cultures of explanted cells were subcultured at low density into medium containing 10% FBS and harvested at specific time points for Western blot detection of N-cadherin. Relative N-cadherin expression was determined by densitometry using non-muscle myosin for normalization. ICC determined N-cadherin localization at the same time points. Results: Rat and human endothelium stained intensely for N- and E-cadherins. A low level of staining for P and R-cadherins was observed in human endothelium. Rat endothelium did not stain for P or R-cadherin over background levels. The expression and localization of N-cadherin was futher studied. Corneal whole mounts showed intense lateral and basolateral staining in rat and human corneal endothelium. Western blots of cultured rat cells showed a density-dependent expression of N-cadherin, which increased as cells approached confluence. At confluence protein levels remained at detectable, though slightly reduced, levels. ICC showed gradual accumulation of intense membrane-associated staining as cells formed cell-cell contacts. Conclusions: N- and E-cadherin are the major "classical" cadherin isoforms expressed in human and rat corneal endothelium. ICC in corneal whole mounts suggests that N-cadherin is most likely involved in both cell-cell and cell-substrate contacts. Protein expression of N-cadherin is density-dependent. Of the four "classical" cadherins, N-cadherin is a strong candidate for mediating cell-cell contact and contact-dependent inhibition of proliferation in corneal endothelium.
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