Mechanisms of lymphocyte adhesion to cultured human corneal epithelial cells

Abstract

PURPOSE. The authors studied the adhesion mechanisms between peripheral blood lymphocytes (PBL) and cultured human corneal epithelial (HCE) cells to investigate the lymphocyte interaction with corneal epithelial cells in the corneal immune response. METHODS. First, the authors examined the expression of inter-cellular adhesion molecule (ICAM)-1 or lymphocyte function-associated antigen (LFA)-3 on the normal human corneal epithelium and cultured HCE cells by an immunostaining technique and flow cytometry. Effects of inflammatory cytokines such as interferon (IFN)-? and tumor necrosis factor (TNF)-a, on ICAM-1 or LFA-3 expression on cultured HCE cells were also examined. Second, the authors performed an adhesion assay with 51 Cr-labeled monocyte-depleted PBL from normal, healthy volunteers and cultured HCE cells, with and without treatment of IFN-? or TNF-a in 96-well-plates for 1 hour at 37°C in 5% CO 2. After unbound PBL were removed, the radioactivity of the sample in each well was counted with a scintillation counter. In addition, the authors evaluated the blocking effects of monoclonal antibodies (mAbs) on the adhesion of PBL to the cultured HCE cells. RESULTS. ICAM-1 expression was not detected in the normal human corneal epithelium. However, the expression of ICAM-1 was detected on the cultured HCE cells with Dulbecco's Modified Eagle Medium supplemented with 10% fetal bovine serum. In addition, both IFN-? and TNF-a increased ICAM-1 expression on the cultured HCE cells dramatically. LFA-3 expression was detected in all cell layers of the normal human corneal epithelia. Neither IFN-? nor TNF-a had any effect on LFA-3 expression on the cultured HCE cells. The PBL adhesion to the HCE cells with and without treatment of IFN-? or TNF-a was blocked dominantly by anti-ICAM-1 or anti-LFA-1a mAb. Anti-LFA-3 mAb also blocked the PBL adhesion but had less blocking effect than anti-ICAM-1 or anti-LFA-1a mAb. Anti-very late activation antigen b, or anti-human leukocyte antigen (HLA)-class I or HLA-class II mAb had no effect on the PBL adhesion to the HCE cells. The adhesion percentile of the PBL applied to the HCE cells pretreated with IFN-? or TNF-a showed a dose-response curve dependent on the concentration of these cytokines. CONCLUSIONS. The results in the present study demonstrate that (i) adhesion of lymphocytes to HCE cells could be mediated by the LFA-1-ICAM-1 pathway and/or the CD2-LFA-3 pathway; (ii) the LFA-1-ICAM-1 pathway could be crucial in lymphocyte adhesion to HCE cells; (iii) IFN-? or TNF-a exerts an enhancing effect not only on the ICAM-1 expression on HCE cells but also on the adhesion of lymphocytes to HCE cells.