Analysis of Ag-presenting cells in the conjunctiva during the development of experimental immune-mediated blepharoconjunctivitis.
OBJECTIVE
The aim of this study was to investigate the phenotypes of antigen (Ag) presenting cells (APCs) in the conjunctiva during the development of experimental immune-mediated blepharoconjunctivitis (EC), which serves as a model for investigation of severe types of human allergic conjunctivitis.


METHODS
Brown Norway rats treated by ovalbumin (OVA) were used in this study. To confirm the restriction of MHC class II by OVA-specific T cells, monoclonal Abs against MHC class II were added to the conventional proliferation assay. To evaluate the MHC class II expression in the conjunctiva during the development of EC, an immunohistochemical analysis, either as the single or double staining, was performed. Conjunctival fibroblast cell lines were established from naive rats and the MHC class II expression was evaluated by flow cytometric analysis. To examine the roles of costimulatory molecules, OVA-specific T cells were stimulated with anti-TcR Ab and anti-CD28 Ab and then subjected for Western blotting to evaluate the ERK phosphorylation. Finally, in vivo expression of B7 molecules was examined immunohistochemically.


RESULTS
OVA-specific T cells recognized OVA in the context of MHC class II. MHC class II was expressed in conjunctival macrophages but not in fibroblasts. EC induction was accompanied by abundant infiltration of macrophages positive for MHC class II. MHC class II was also expressed in conjunctival epithelial cells by EC induction. Stimulation from CD28 was necessary for ERK phosphorylation. B7-2, but not B7-1, was expressed in the conjunctiva.


CONCLUSIONS
Conjunctival macrophages may represent a major source of APCs for the induction of EC in the conjunctiva.
Ozaki
Akemi
A
Department of Ophthalmology, Kochi Medical School, Kohasu, Oko-cho, Nankoku City 783-8505, Japan.
Fukushima
Atsuki
A
Ishida
Waka
W
Zinchuk
Olga
O
Fukata
Kazuyo
K
Hayashi
Yoshihiro
Y
Enzan
Hideaki
H
Takata
Masaru
M
Yagita
Hideo
H
Ueno
Hisayuki
H
D000818:Animals; D000938:Antigen-Presenting Cells / Q000276:immunology*; D015703:Antigens, CD / Q000378:metabolism; D051940:B7-2 Antigen; D001762:Blepharitis / Q000276:immunology*; D015153:Blotting, Western; D002452:Cell Count; D003228:Conjunctiva / Q000276:immunology*; D003233:Conjunctivitis, Allergic / Q000276:immunology*; D048049:Extracellular Signal-Regulated MAP Kinases / Q000378:metabolism; D005434:Flow Cytometry; D000949:Histocompatibility Antigens Class II / Q000378:metabolism; D007124:Immunoenzyme Techniques; D016130:Immunophenotyping; D008213:Lymphocyte Activation; D008264:Macrophages / Q000276:immunology*; D008297:Male; D008562:Membrane Glycoproteins / Q000378:metabolism; D018613:Microscopy, Confocal; D008962:Models, Theoretical; D010047:Ovalbumin / Q000276:immunology; D010766:Phosphorylation; D051381:Rats; D011914:Rats, Inbred BN; D013601:T-Lymphocytes / Q000276:immunology
D015703:Antigens, CD; D051940:B7-2 Antigen; C497966:Cd86 protein, rat; D000949:Histocompatibility Antigens Class II; D008562:Membrane Glycoproteins; D010047:Ovalbumin; D048049:Extracellular Signal-Regulated MAP Kinases