Cardiac allograft tolerance: failure to develop in interleukin-4-deficient mice correlates with unusual allosensitization patterns.

BACKGROUND: The development of long-term allograft survival and understanding the mechanism(s) by which it is induced are major goals of experimental transplantation. Studies by several different investigators have provided conflicting evidence for the role of interleukin (IL)-4 in the process of allograft rejection or long-term allograft survival. These studies examine the role of IL-4 in experimental cardiac allograft rejection and in inducing long-term allograft survival. A possible mechanism for long-term allograft maintenance via alternative allosensitization is discussed. METHODS: Adult IL-4-intact or IL-4-deficient (knockout, KO) C57BL/6 (B6) mice were transplanted with heterotopic DBA/2 cardiac allografts and immunosuppressed either with gallium nitrate (GN), or the anti-CD4 monoclonal antibody, GK1.5. Cellular allosensitization was assessed by testing the allograft recipients for donor-reactive delayed-type hypersensitivity (DTH) responses. The presence of antigen-driven suppressive mechanisms was assessed using a linked unresponsiveness (bystander suppression) DTH assay. RESULTS: In general, the results were the same with either GN or GK1.5. We observed that (1) IL-4 is not required for acute allograft rejection or allogeneic DTH responses in nonsuppressed mice, (2) IL-4 is required for long-term allograft survival in immunosuppressed mice, (3) immunosuppression creates a requirement for IL-4 in major histocompatibility complex self-restricted, but not allorestricted, DTH responses, and (4) the development of alloantigen-dependent linked DTH unresponsiveness (bystander suppression) in allograft recipients requires IL-4. CONCLUSION: In summary, these studies demonstrate a common requirement for IL-4 during the development of long-term allograft survival and the concurrent development of alloantigen-dependent DTH down-regulation in cardiac allograft recipients after immunosuppression.

Patterns of allosensitization in allograft recipients: long-term cardiac allograft acceptance is associated with active alloantibody production in conjunction with active inhibition of alloreactive delayed-type hypersensitivity.

BACKGROUND: The immunologic characteristics of experimental allograft acceptance remain ill-defined. This study evaluates humoral and cell-mediated immunity in transiently immunosuppressed mice that have accepted cardiac allografts. METHODS: DBA/2-->C57BL/6 heterotopic cardiac allograft recipients were immunosuppressed with either GK1.5 monoclonal antibody or gallium nitrate and monitored for donor-reactive delayed-type hypersensitivity (DTH) assessed by ear challenge and for alloantibody production detected by flow cytometry. RESULTS: Cardiac allograft function continued for >90 days in approximately 50% of GK1.5-treated and 97% of gallium nitrate-treated transplant recipients. All nonsuppressed recipients lost graft function within 7 to 10 days. Among mice that accepted allografts, donor-reactive IgG was produced by about 50% of GK1.5 monoclonal antibody-treated mice and 80% of gallium nitrate-treated mice. None of the these mice exhibited donor-reactive DTH responses, and all could down-regulate third-party DTH responses in a donor alloantigen-dependent manner. This down-regulation is not found in nonsuppressed allograft recipients or in naive mice. Importantly, transfer into SCID mice of splenocytes from mice that accepted allografts, but not naive splenocytes, provided them with a similar ability to accept cardiac allografts, even if the grafts co-expressed third-party alloantigens. CONCLUSIONS: IgG alloantibody production by murine cardiac allograft recipients is not a precise indicator of allosensitization leading to either cardiac allograft rejection or acceptance. However, expression of alloreactive DTH is a reliable indicator of allosensitization leading to acute rejection, and the absence of DTH in association with active DTH down-regulatory mechanisms is a reliable indicator of allograft acceptance in this experimental model. Thus, DTH analysis may hold more promise than alloantibody detection for clinical assessment of posttransplant immune status.

Alloreactive delayed-type hypersensitivity in graft recipients: complexity of responses and divergence from acute rejection.

BACKGROUND: Immunocompetent allograft recipients typically exhibit evidence of sensitization to graft antigens through alloantibody production and allograft rejection, as well as delayed-type hypersensitivity (DTH) reactivity to donor antigens. Most previous studies have relied on whole donor splenocytes, which primarily elicit allorestricted allogeneic responses, to test specific DTH responses and overlook the independent element of self-restricted responses in host-allograft interactions. METHODS: We tested expression of self-MHC-restricted versus allo-MHC-restricted allogeneic DTH responses in both nonimmunosuppressed and tolerized C57BL/6 mice. Mice were sensitized for allogeneic DTH either by rejection of skin or cardiac allografts, or by subcutaneous injection of intact allogeneic splenocytes. Patterns of alloreactive DTH were compared in allosensitized, tolerant, and naive hosts. RESULTS: All three methods of allosensitization resulted in equivalent self-restricted and allorestricted allogeneic DTH responses in nonimmunosuppressed mice. Gallium nitrate blocked acute rejection of cardiac allografts, and also blocked allosensitization of both self-restricted and allorestricted DTH responses, but did not influence the expression of DTH responses in presensitized mice. Gallium nitrate treatment could not block acute rejection of skin allografts, but interfered with sensitization for self-restricted, but not allorestricted, DTH responses in these recipients. This divergence of self- versus allo-MHC-restricted allosensitization for DTH was observed in two additional situations: the rates of allosensitization for self-restricted versus allorestricted DTH, and the acquisition of allorestricted, but not self-restricted, alloreactive DTH responses in cardiac allograft tolerant mice subsequently challenged with a skin allograft. CONCLUSIONS: These studies demonstrate that acute rejection correlates generally with allogeneic DTH, whereas tolerance is associated with a lack of alloreactive DTH. However, self-restricted and allorestricted allosensitization can operate independently in allograft recipients. Thus, the relationships between alloreactive DTH and graft-induced allosensitization, acute rejection, or tolerance are more complicated than previously appreciated.