Eugenia V. Fedoseyeva

The role of CD154-CD40 versus CD28-B7 costimulatory pathways in regulating allogeneic Th1 and Th2 responses in vivo

We used signal transducer and activator of transcription 4 (STAT4) and STAT6 gene knockout (-/-) mice as recipients of fully mismatched cardiac allografts to study the role of T-cell costimulatory pathways in regulating allogeneic T-helper 1 (Th1) versus Th2 responses in vivo. STAT4(-/-) mice have impaired Th1 responses, whereas STAT6(-/-) mice do not generate normal Th2 responses. Cardiac allografts from C57BL/6 mice were transplanted into normal wild-type (WT), STAT4(-/-), and STAT6(-/-) BALB/c recipients. STAT4(-/-) and STAT6(-/-) mice rejected their grafts with the same tempo as untreated WT recipients. CD28-B7 blockade by a single injection of CTLA4Ig induced long-term engraftment and donor-specific tolerance in all three groups of recipients. CD154 blockade by a single injection of MR1 was effective in prolonging allograft survival and inducing tolerance in STAT4(-/-) mice but was only marginally effective in STAT6(-/-) recipients and WT controls. In addition, a similar protocol of MR1 was ineffective in prolonging graft survival in CD28(-/-) BALB/c recipients, suggesting that the lack of efficacy seen in WT and STAT6(-/-) mice is not due to the presence of a functional CD28-B7 pathway. Furthermore, there was a similar differential effect of CD28-B7 versus CD154-CD40 blockade in inhibiting immune responses in animals immunized with ovalbumin and complete Freunds adjuvant. These novel data indicate that Th1 and Th2 cells are differentially regulated by CD28-B7 versus CD154-CD40 costimulation pathways in vivo and may have potential implications for the development of therapeutic strategies such as T-cell costimulatory blockade in humans.

Modulation of Tissue-Specific Immune Response to Cardiac Myosin Can Prolong Survival of Allogeneic Heart Transplants

The role of immune response to tissue-specific Ags in transplant rejection is poorly defined. We have previously reported that transplantation of cardiac allografts triggers a CD4+ Th1 cell response to cardiac myosin (CM), a major contractile protein of the heart, and that pretransplant activation of proinflammatory CM-specific T cells accelerates rejection. In this study, we show that administration of CM together with IFA (CM/IFA) can prevent acute rejection of an allogeneic heart transplant. Prolongation of cardiac graft survival is associated with activation of CM- and allo-specific T cells secreting type 2 cytokines (IL-4, IL-5) and reduction of the frequency of proinflammatory IFN-γ-secreting (type 1) alloreactive T cells. Blocking of IL-4 cytokine with Abs abrogates the prolongation. CM/IFA treatment prevents acute rejection of MHC class I-mismatched, but not fully mismatched grafts. However, if donor heart is devoid of MHC class II expression, CM-IFA administration delays rejection of fully allogeneic cardiac transplants. This finding suggests that the effect of CM modulation depends on the type (direct vs indirect) and strength of recipient’s CD4+ T cell alloresponse. Our results underscore the important role of host immunity to tissue-specific Ags in the rejection of an allograft. This study demonstrates that modulation of the immune response to a tissue-specific Ag can significantly prolong cardiac allograft survival, an observation that may have important implications for the development of novel selective immune therapies in transplantation.

Indirect T-cell allorecognition: perspectives for peptide-based therapy in transplantation l

Indirect allorecognition is an important component of allotransplant rejection. Although the initial indirect alloresponse is limited to a few dominant determinants on donor major histocompatibility complex (MHC) molecules, subsequent spreading to additional determinants on recipient and donor antigens is common. Gilles Benichou and colleagues discuss the mechanisms by which immunodominance is acquired or disrupted in indirect alloresponses, and examine the implications for the design of peptide-based selective immunotherapy in transplantation.

The relative contribution of direct and indirect antigen recognition pathways to the alloresponse and graft rejection depends upon the nature of the transplant.

In this study, we measured direct and indirect T-cell alloresponses mediated by CD4(+) and CD8(+) T cells in three mouse transplantation models: skin, cornea, and retina. We show that the contribution of direct and indirect antigen recognition pathways to the alloresponse to fully allogeneic grafts varies depending upon the nature of the tissue/organ transplanted. The implications of this finding for understanding the cellular mechanisms by which rejection is mediated in different transplant models are discussed.

Induction of T cell responses to a self-antigen following allotransplantation.

T cell tolerance to self-antigens is established through the recognition by immature T cells of dominant self-peptides presented in association with self-MHC molecules in the developing thymus (negative selection). The self-peptide Dd 61-80 is dominant in syngeneic BALB/c mice (H2d). T cell tolerance to Dd 61-80 in this mouse strain resulted in the absence of T cell proliferation following in vivo priming with Dd 61-80 peptide. Here, we show that transplantation of BALB/c mice with allogeneic B10.A (H2a) splenocytes led to an autoimmune T cell response toward the dominant self-peptide Dd 61-80. No T cell responses to Dd 61-80 peptide were observed after transplantation of C57BL/6 (H2b) splenocytes into BALB/c recipients. In addition, we provide evidence indicating that the breakdown of tolerance to Dd 61-80 self-peptide resulted from the presentation of the donor crossreactive peptide Kk 61-80 at the surface of recipient antigen-presenting cells. Taken together, our results suggest that following allotransplantation, T cell responses to donor antigens could spread to crossreactive determinants on self-proteins, thus perpetuating and amplifying the rejection process and presumably initiating tissue-specific autoimmune disorders.

T-cell response to cardiac myosin persists in the absence of an alloimmune response in recipients with chronic cardiac allograft rejection

BACKGROUND Immune-mediated injury to the graft has been implicated in the pathogenesis of chronic rejection. However, little is known regarding the nature of the antigen(s) involved in this immune process. We demonstrated that cardiac transplantation in mice induces an autoimmune T-cell response to a heart tissue-specific protein, cardiac myosin (CM). This response contributes to transplant rejection in that its modulation affects cardiac graft survival. This study investigates whether anti-CM T cells undergo activation and expansion in mice with chronic cardiac allograft rejection. METHODS The frequency of CM- and donor major histocompatibility complex (MHC)-specific interferon (IFN)-gamma-producing T cells were assessed by ELISPOT in BALB/c mice, which were injected with anti-CD40L (MR1) mAb (chronic rejection group) or CTLA4Ig fusion protein (tolerant group) and transplanted with C57BL/6 cardiac allografts. RESULTS AND CONCLUSIONS MR1-treated BALB/c recipients of C57BL/6 hearts with chronic rejection displayed a high frequency of activated CM-specific T cells, whereas the frequency of activated alloreactive T cells were similar to naïve, nontransplanted mice. In contrast, no activation of CM-reactive T cells was detected in tolerant recipients after CTLA4Ig treatment. Therefore, in the absence of alloimmunity, chronic rejection is associated with persistence of a T-cell response against CM. Our data indicate that anti-CM autoimmunity may be involved in the immune mechanisms of chronic rejection and suggest that tolerance strategies should target both allo- and autoimmune responses to prevent this process.

Development of autoimmunity after skin graft rejection via an indirect alloresponse.

Background. T cell allorecognition occurs through direct contact with donor peptide: MHC complexes on graft cells and through indirect recognition of donor-derived determinants expressed by recipient MHC molecules. As both indirect allorecognition and autoantigen recognition are self-restricted, we hypothesized that chronic activation of indirectly primed T cells might result in determinant spreading to involve autoantigens, analogous to that which occurs during chronic autoimmune diseases. Methods. We placed C57BL/6 MHC II knockout (B6 II-/-) skin grafts onto BALB/c SCID mice reconstituted with wild-type (WT) CD4+ T cells. Under these conditions the CD4+ cells could not recognize any antigen on the graft, but could respond through the indirect pathway. CD4+ cell-mediated rejection of WT B6 skin was studied to determine if autoreactivity was induced after direct allorecognition. Recall immune responses against donor- and self-stimulator cells were determined by ELISPOT and animals were tested for their ability to reject second isografts. Results. WT allografts were rejected by day 14 although B6 II-/- grafts underwent delayed rejection over 4–5 weeks. CD4+ cells reisolated from the recipients of the MHC II-/- grafts, but not from the recipients of WT grafts, vigorously produced interferon-&ggr; and interleukin-2 in response to self, BALB/c stimulators. These autoreactive CD4+ T cells mediated rejection of a second isogenic BALB/c skin graft, demonstrating that the autoimmune response was pathogenic. Conclusion. Autoreactivity can develop after transplant rejection via the indirect pathway. Although the direct alloresponse is likely to be the driving force in acute graft rejection, posttransplantation induced autoimmune responses may be important elements of delayed or chronic rejection.

Induction of T-cell response to cryptic MHC determinants during allograft rejection.

The presentation of MHC peptides by recipient and donor antigen presenting cells is an essential element in allorecognition and allograft rejection. MHC proteins contains two sets of determinants: the dominant determinants that are efficiently processed and presented to T cells, and the cryptic determinants that are not presented sufficiently enough to induce T-cell responses in vivo. In transplanted mice, initial T-cell response to MHC peptides is consistently limited to a single or a few immunodominant determinants on donor MHC molecule. However, in this article we show that under appropriate circumstances the hierarchy of determinants on MHC molecules can be disrupted. First, we observed that gamma IFN can trigger de novo presentation of cryptic self-MHC peptides by spleen cells. Moreover, we showed that allotransplantation is associated with induction of T-cell responses to formerly cryptic determinants on both syngeneic and allogeneic MHC molecules. Our results suggest that cross-reactivity and inflammation are responsible for the initiation of these auto- and alloimmune responses after transplantation.

CD4+ T Cell Responses to Self- and Mutated p53 Determinants During Tumorigenesis in Mice

We analyzed CD4+ T helper responses to wild-type (wt) and mutated (mut) p53 protein in normal and tumor-bearing mice. In normal mice, we observed that although some self-p53 determinants induced negative selection of p53-reactive CD4+ T cells, other p53 determinants (cryptic) were immunogenic. Next, BALB/c mice were inoculated with J774 syngeneic tumor cell line expressing mut p53. BALB/c tumor-bearing mice mounted potent CD4+ T cell responses to two formerly cryptic peptides on self-p53. This response was characterized by massive production of IL-5, a Th2-type lymphokine. Interestingly, we found that T cell response was induced by different p53 peptides depending upon the stage of cancer. Mut p53 gene was shown to contain a single mutation resulting in the substitution of a tyrosine by a histidine at position 231 of the protein. Two peptides corresponding to wt and mutated sequences of this region were synthesized. Both peptides bound to the MHC class II-presenting molecule (Ed) with similar affinities. However, only mut p53.225–239 induced T cell responses in normal BALB/c mice, a result strongly suggesting that high-affinity wt p53.225–239 autoreactive T cells had been eliminated in these mice. Surprisingly, CD4+ T cell responses to both mut and wt p53.225–239 peptides were recorded in J774 tumor-bearing mice, a phenomenon attributed to the recruitment of low-avidity p53.225–239 self-reactive T cells.

THE PRESENTATION OF SELF AND ALLOGENEIC MHC PEPTIDES TO T LYMPHOCYTES

The presentation of donor-derived MHC peptides by recipient APCs to T cells is an essential component of the rejection of allografts (indirect allorecognition). Initial alloreactive T cell response is confined to a few well processed and presented dominant determinants on donor MHC. However, during long-term graft rejection, T cell response spreads to formerly poorly presented cryptic allogeneic MHC peptides. This phenomenon is likely to play an important role in the amplification and the perpetuation of the rejection process. Additionally, we present evidence that T cell repertoire selection to allogeneic MHC peptides is acquired via recognition of self-MHC peptides presented in the thymus during ontogeny. Supporting this view, we have shown that indirect alloresponses can lead to self-T cell tolerance breakdown to cross-reactive determinants on self-MHC molecules or alternatively that sensitization of recipients to self-MHC peptides can lead to accelerated graft rejection. It is therefore essential to determine the factors which govern the processing and presentation of self and allogeneic MHC molecules and to elucidate the mechanisms regulating subsequent T cell responses in order to design antigen-specific based immune therapies in transplantation.