Role of CD4+ T cells in immunobiology of orthotopic corneal transplants in mice.

To determine, with the use of mice genetically deficient in expression of CD4 or CD8 molecules, which T cells are responsible for rejection of orthotopic corneal allografts in mice. Corneas were prepared from major histocompatibility complex (MHC)-only incompatible, minor histocompatibility (H)- only incompatible, and MHC-plus-minor H incompatible donors and grafted orthotopically to eyes of CD4 knockout (KO), CD8KO, and wild-type control mice. Graft survival patterns were assessed clinically and compared. Mice that retained healthy corneal allografts beyond 8 weeks were evaluated for evidence of donor-specific tolerance and anterior-chamber-associated immune deviation (ACAID) using local adoptive transfer reactions and challenge with orthotopic skin allografts. Corneas grafted to CD8KO mice were rejected with an incidence and tempo indistinguishable from that in wild-type control animals. By contrast, MHC-only, and minor-H-only incompatible corneal grafts survived indefinitely in eyes of CD4KO mice. Approximately 50% of corneal grafts that confronted CD4KO recipients with both MHC and minor H alloantigens experienced delayed rejection, whereas similar grafts in wild-type recipients were rejected acutely. CD4KO mice with long-accepted grafts displayed neither donor-specific ACAID nor allograft tolerance. CD8+ T cells play little or no role in acute rejection of orthotopic corneal allografts. Instead, acute rejection is mediated almost exclusively by CD4+ T cells. Moreover, when corneal allografts survive for 8 weeks without acute rejection, CD4+ T cells promote donor-specific ACAID thereby insuring long-term graft acceptance.

Yamada J ，Kurimoto I ，Streilein JW 《INVESTIGATIVE OPHTHALMOLOGY & VISUAL SCIENCE》

Characterization of cell-mediated immune responses elicited by orthotopic corneal allografts in mice.

Corneal allografts placed orthotopically induce a unique and unusual response in recipient mice. More orthotopic corneal allografts are accepted indefinitely than similar skin allografts. Of the rejected corneal grafts, class I major histocompatibility complex (MHC)-incompatible grafts are rejected less frequently than grafts that express only minor histocompatibility complex (minor H) or MHC plus minor H alloantigens. To describe the spectrum of T cells activated (or not) by orthotpic corneal grafts, the authors examined the development of delayed hypersensitivity (DH) to donor-specific alloantigens. Recipient BALB/c mice received orthotopic corneal allografts from donor mice that were MHC incompatible at MHC loci only, multiple minor H loci only, or MHC plus multiple minor H loci. These groups of mice were examined to determine when alloantigen-specific DH developed. The authors report that all mice, whether they accept or reject grafts, acquire donor-specific DH within 4 weeks of engraftment. This reactivity is primarily directed at minor H, rather than MHC-encoded, alloantigens. Through time, spontaneous DH reactivity disappears in all mice, and thereafter, donor-specific DH can be induced by cognate immunization only in mice that have rejected their cornea grafts. These results can be explained in the context of "direct" and "indirect" pathways of allorecognition. Because normal corneas lack passenger leukocytes, the potential for direct recognition of alloantigens on orthotopic corneal grafts is small. Therefore, T cells activated by orthotopic corneal allografts must recognize donor-derived antigens primarily on recipient antigen presenting cells, that is, through the indirect pathway of allorecognition. Because minor H antigens are the dominant cellular proteins in grafts, it is proposed that minor H determinants are the most immunogenic alloantigens in orthotopic corneal grafts because they are the major source of peptides that will be loaded onto recipient class II molecules for T-cell recognition. We further predict that long-term acceptance of corneal allografts is promoted when recipient mice acquire anterior chamber associated immune deviation (impaired and suppressed DH) directed at minor H alloantigens of the grafts.

Sonoda Y ，Sano Y ，Ksander B ，Streilein JW ... -  《INVESTIGATIVE OPHTHALMOLOGY & VISUAL SCIENCE》

Cytotoxic T cells play no essential role in acute rejection of orthotopic corneal allografts in mice.

To determine whether cytotoxic T cells of the direct alloreactive type are activated and responsible for early, acute failure of orthotopic corneal allografts observed in eyes of C57BL/6 but not of BALB/c mice. Corneas from BALB/c and BALB.B mice were placed orthotopically in eyes of C57BL/6 and beta-2 microglobulin knockout mice (deficient in CD8(+) cytotoxic T cells). Graft fates were assessed clinically, and the T lymphocytes of recipients were assayed for the capacity to lyse target cells bearing donor major (MHC) and/or minor histocompatibility (minor H) antigens (direct and indirect pathways, respectively). RESULTS. Similar to BALB/c recipients, C57BL/6 mice with rejected cornea allografts acquired donor minor H-specific T cells. Unlike BALB/c recipients, C57BL/6 mice-both rejectors and acceptors-acquired donor MHC-specific T cells. beta-2 Microglobulin knockout mice showed rejection of corneal allografts in a manner indistinguishable from C57BL/6 mice, including early, acute rejection, yet T cells from beta-2 microglobulin knockout recipients of corneal allografts displayed no cytotoxic T cells specific for either donor MHC or minor H alloantigens. Although C57BL/6 mice acquired donor MHC-specific cytotoxic T cells (direct alloreactive cells), neither these cells nor donor minor H-specific cytotoxic T cells (indirect alloreactive cells) play any essential role in corneal allograft rejection, including the early acute failure uniquely observed in C57BL/6 eyes.

Yamada J ，Ksander BR ，Streilein JW 《INVESTIGATIVE OPHTHALMOLOGY & VISUAL SCIENCE》

Fate of orthotopic corneal allografts in eyes that cannot support anterior chamber-associated immune deviation induction.

Corneal allografts placed in human eyes at high risk often fail, and immune rejection is thought to be a major pathogenic factor. To understand the immunologic factors responsible for rejection in this instance, the authors have created "high-risk" eyes in mice by inducing corneal neovascularization. The authors then examined the fate of orthotopic corneal grafts placed in these beds and assessed the development of donor-specific delayed hypersensitivity (DH) in recipient mice. Three interrupted sutures were placed in the central cornea of recipient BALB/c mice to induce corneal neovascularization. Two weeks later, when corneal vessels occupied more than two quadrants of the cornea, mice received orthotopic corneal grafts from donor mice expressing alloantigens encoded by major and minor histocompatibility loci. Corneal allografts were evaluated by slit-lamp examination after grafting, and recipient mice were examined at the time of the rejection to determine whether they had acquired DH to alloantigens expressed on the corneal grafts. Compared to grafts placed in normal eyes, a much higher incidence of rejection was observed among corneal allografts placed in neovascularized eyes (96.7% versus 46.7%). Moreover, grafts in neovascularized beds were rejected much more swiftly (2 weeks versus > 3 to 4 weeks). Rejection of corneal allografts in high-risk eyes coincided temporally with development of intense donor-specific DH, and the specificity of this immune response was directed solely at minor H antigens (not major histocompatibility complex-encoded antigens) on the graft. These results indicate that eyes rendered high risk by virtue of corneal neovascularization fail to provide immune privilege for orthotopic corneal allografts. In this circumstance, the grafts rapidly induce intense donor-specific DH that is readily detectable within 2 weeks of engraftment, at which time the grafts are acutely and universally rejected. The recipient DH response is directed exclusively at minor H antigens on the graft, which is consistent with the view that neovascularization creates graft beds in which recipient antigen-presenting cells infiltrate the graft and carry antigenic information by lymphatics to draining lymph nodes. In this manner, anterior chamber-associated immune deviation is avoided, and potentially allodestructive DH is promoted.

Sano Y ，Ksander BR ，Streilein JW 《INVESTIGATIVE OPHTHALMOLOGY & VISUAL SCIENCE》

An immunodominant minor histocompatibility alloantigen that initiates corneal allograft rejection.

Haskova Z ，Sproule TJ ，Roopenian DC ，Ksander AB ... -  《TRANSPLANTATION》