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J. Hori, M. Miyashita, H. Takahashi, T. Takemori, H. Yagita, M. Azuma; B7–H1–Mediated Protection of Corneal Endothelial Cells From Killing by Allo–Reactive T Cells in vitro . Invest. Ophthalmol. Vis. Sci. 2006;47(13):5866.
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© ARVO (1962-2015); The Authors (2016-present)
Programmed Death–1 (PD–1) / B7–H1 costimulatory pathway has been demonstrated to play a role in the regulation of immune responses and peripheral tolerance. We have demonstrated that cornea of normal mouse eyes constitutively express B7–H1, and that PD–1/B7–H1 pathway has a role in corneal allograft survival by inducing apoptosis of infiltrating cells. To further substantiate the B7–H1–mediated protection of corneal allografts from effector T cells, we evaluated the corneal endothelial cell (CEC) destruction by allo–reactive T cells in vitro.
Fresh normal cornea from C57BL/6 eyes was incubated with anti–B7–H1 mAb or control rat IgG for 2 h. Magnetic cell sorting and separation was used to purify T cells from the spleen of BALB/c mice that were pre–sensitized by subcutaneous immunization with C57BL/6 spleen cells or with third–party (C3H/He) spleen cells, or from the spleen of naive BALB/c, C57BL/6, or C3H/He mice. The cornea pre–treated with anti–B7–H1 monoclonal antibody (mAb) or control rat IgG was incubated with 2.5x105 T cells. The unfixed corneal samples were incubated with 50 ug/ml propidium iodide (PI) to stain nuclei of dead CEC. PI–positive cells were counted at three randomly selected areas in the corneal endothelium of each corneal sample using confocal microscopy.
The killing of CEC by allo–reactive T cells in vitro was significantly enhanced in the corneas pre–treated with anti–B7–H1mAb as compared to those pre–treated with control IgG (p=0.0016). Interestingly, B7–H1–mediated protection was also observed after incubation with third–party–reactive T cells (p=0.0248). Moreover, the B7–H1–mediated protection was still observed even after incubation with naive allogeneic T cells (p=0.0051). In contrast, no significant difference was observed between the anti–B7–H1 mAb and control rat IgG–treated corneas when syngeneic C57BL/6 T cells were used as the effector.
B7–H1 expressed on CEC plays a substantial role in the protection of CEC from destruction by allo–reactive effector T cells. B7–H1 protects cornea from bystander injury by activated T cells. Moreover, constitutive expression of B7–H1 in cornea may inhibit the peripheral sensitization, since naive T cells were sensitized by B7–H1–deficient allogeneic cornea to promote their injury.
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