Comparison of tight junction protein expression in the ciliary epithelia of mouse, rabbit, cat and human eyes

Abstract

Tight junctions in the nonpigmented epithelium (NPE) of the ciliary processes and the iris vascular endothelium form the ocular blood aqueous barrier that prevents leakage of proteins, immune cells and non-immune cells of blood into the anterior chamber. We attempted to determine whether ultrastructural differences in tight junctions reported in earlier studies are reflected in the expression pattern of tight junction proteins (TJP) and whether the TJP in mice, rabbits and cats resemble those of humans. For immunohistochemistry, 10 μm thick cryosections were rehydrated in PBS and fixed in 50 mM ammonium chloride at room temperature. After rinses in PBS, the sections were incubated twice in 0.1% Triton X-100, 10% goat serum, specific primary antibody or in PBS. After rinses in PBS, the sections were incubated in FITC-conjugated secondary antibody. After rinses in PBS, the sections were mounted with Vectashield mounting medium with propidium iodide, examined and photographed using a confocal microscope. The expression patterns of TJP in ocular ciliary epithelium of human, rabbit, cat and mouse were similar. Occludin immunoreactivity was observed as a sharp line along the junction between pigmented epithelium (PE) and NPE, and along the apico-lateral surfaces of NPE. Very light staining of the ciliary stroma was observed in cat and mouse. Claudin-1 was expressed along the entire boundaries of NPE and was more distinct between PE and NPE in rabbit. The ciliary stroma showed faint staining in cat and mouse. ZO-1 showed staining between PE and NPE, and at the adjacent membrane. Moderate staining was seen in PE in cat and mouse, which suggests that claudin-1, occludin and ZO-1 are expressed along the junction between PE and NPE, and the apico-lateral border of NPE. Lack of major difference in the expression patterns among the different species is important for validating the use of rabbit, mouse and cat in studies of intraocular inflammation in humans.

Key words::

• ciliary epithelium
• claudin-1
• histochemistry
• occludin
• tight junction
• ZO-1

Acknowledgments

Supported by National Institutes of Health, National Eye Institute Grant EY-06918, the Kentucky Lions Eye Foundation and an unrestricted grant from Research to Prevent Blindness, Inc., New York, NY. We thank Kathy Shaeffer for her technical assistance.

Declaration of interest: The authors report no conflicts of interest. The authors alone are responsible for the content and writing of the paper.