Antibody blockade of junctional adhesion molecule-A in rabbit corneal endothelial tight junctions produces corneal swelling.
OBJECTIVE
The ultrastructure of tight junctions in the corneal endothelium has been studied extensively, yet little is known about their molecular composition. Junctional adhesion molecule-A (JAM-A) is a tight junction-associated adhesion protein previously implicated in tight junction assembly and regulation of barrier function. In this study, we sought to investigate the expression and function of JAM-A in the corneal endothelium.


METHODS
Immunofluorescence confocal microscopy was used to investigate expression of JAM-A and the related proteins JAM-C, CAR, and AF-6 in the rabbit corneal endothelium. Corneal endothelial perfusion specular microscopy was then used to test the effects of antibodies to JAM-A on corneal swelling.


RESULTS
The expression of JAM-A was observed in the tight junctions of rabbit corneal endothelium in a localization pattern identical with that of ZO-1, a known marker of the tight junction and binding partner of JAM-A. Expression of related proteins JAM-C and CAR (Coxsackie and adenovirus receptor) was also observed in the corneal endothelium, but their distribution was diffuse and not limited to the tight junction. Expression of AF-6, a known binding partner of JAM-A, was also observed in the tight junction in a pattern similar to ZO-1. Last, functional experiments were performed in which a monoclonal antibody to JAM-A was shown to increase rabbit corneal swelling by 63% compared with the control.


CONCLUSIONS
The results provide new evidence that JAM-A and its binding partner AF-6 are expressed in tight junctions of the corneal endothelium and that JAM-A has a major role in maintaining the corneal endothelial barrier function.
Mandell
Kenneth J
KJ
Emory Eye Center, Department of Pathology and Laboratory Medicine, Emory University School of Medicine, Atlanta, GA 30322, USA. kjmande@emory.edu
Holley
Glenn P
GP
Parkos
Charles A
CA
Edelhauser
Henry F
HF
D000818:Animals; D019138:Antibodies, Blocking / Q000502:physiology; D000911:Antibodies, Monoclonal; D015153:Blotting, Western; D015815:Cell Adhesion Molecules / Q000502:physiology*; D002463:Cell Membrane Permeability; D015715:Corneal Edema / Q000378:metabolism*; D004728:Endothelium, Corneal / Q000378:metabolism*; D005260:Female; D019084:Fluorescent Antibody Technique, Indirect; D007136:Immunoglobulins / Q000502:physiology*; D016547:Kinesins / Q000378:metabolism; D008565:Membrane Proteins / Q000378:metabolism; D051379:Mice; D008807:Mice, Inbred BALB C; D018613:Microscopy, Confocal; D009218:Myosins / Q000378:metabolism; D010750:Phosphoproteins / Q000378:metabolism; D011817:Rabbits; D011956:Receptors, Cell Surface; D019108:Tight Junctions / Q000378:metabolism*; D062826:Zonula Occludens-1 Protein
C095516:AFDN protein, human; D019138:Antibodies, Blocking; D000911:Antibodies, Monoclonal; D015815:Cell Adhesion Molecules; C407243:F11R protein, human; D007136:Immunoglobulins; D008565:Membrane Proteins; D010750:Phosphoproteins; D011956:Receptors, Cell Surface; C569042:TJP1 protein, human; C569043:Tjp1 protein, mouse; D062826:Zonula Occludens-1 Protein; D009218:Myosins; D016547:Kinesins