The essential roles of parenchymal tissues and passenger leukocytes in the tolerance induced by liver grafting in rats.

Liver allografts in pigs and rodents are uniquely capable of inducing tolerance to themselves and to other grafts of donor tissues, instead of succumbing to the acute rejection that follows transplantation of other allogeneic tissues. We demonstrate here, using normal and chimaeric rat liver grafts, that both the allogeneic liver parenchyma and the intrahepatic leukocytes of donor type contribute to the establishment of long-term tolerance, each component being essential and complementary. The essential role of hepatic parenchyma may be related to its continuous release of soluble transplantation antigens that facilitate tolerogenesis. We suggest that clinical attempts at tolerance induction by the infusion of donor bone marrow-derived leukocytes may likewise be facilitated by the coadministration of soluble transplantation antigens of donor type.

Tolerance of porcine renal allografts induced by donor spleen cells and seven days' treatment with cyclosporine.

Liver allografts in pigs and in rats elicit a substantial cellular immune response that can resolve spontaneously with the induction of donor-specific systemic tolerance. Self-limiting interactions between host and donor (graft)-derived leukocytes may be the basis for tolerogenesis. We have attempted to reproduce this effect of liver grafting in pigs by peroperative infusion of donor leukocytes into kidney graft recipients given an interrupted short course of CsA designed to promote donor leukocyte survival and interaction with host cells. This protocol can secure long-term kidney graft survival resistant to challenge by donor skin grafting. Donor skin is, however, rejected, but more slowly than third-party skin, indicating a degree of systemic specific unresponsiveness in these long-term kidney graft recipients.

Immunosuppression of canine renal allograft recipients by CD4 and CD8 monoclonal antibodies.

A state of tolerance to MHC mismatched allografts can be generated in rodents by treatment with CD4 and CD8 monoclonal antibodies (mAb). In order to transpose this type of therapy to large animals and ultimately to the clinic, a suitable model is required. To this end we have generated a series of mAb to the canine CD4, CD8, and Thy-1 antigens and have tested their ability to prevent rejection of renal allografts. Donor-recipient pairs were selected from a colony of mongrel dogs in which untreated rejection of two haplotype-mismatched kidneys occurred by day 7 (defined as a serum creatinine > 300 mumol/l). Therapy with either the CD4 or the CD8 mAb, using no other immunosuppression, did not prolong graft survival. Depletion of T cells by a Thy-1 mAb prior to surgery only extended graft survival to day 9. However, treating with combinations of mAb up to day 10 (CD4 plus Thy-1; CD4 plus CD8; or CD4 plus CD8 plus Thy-1) prolonged renal allograft function up to 25 days. Combination of the triple mAb therapy with a sub-therapeutic immunosuppressive drug regimen (cyclosporin A plus azathioprine that alone gave a median survival of 15 days) favored survival to a median of 38 days. This protocol also inhibited the antiglobulin response that had curtailed the effects of mAb treatment, opening the way to more extended, and potentially tolerizing, mAb plus drug regimens.

CD4 and CD8 monoclonal antibody therapy: strategies to prolong renal allograft survival in the dog.

The value of CD4 and CD8 monoclonal antibody therapy in tolerance induction has been demonstrated in rodent transplant models. In this paper the immunosuppressive potential of CD4 and CD8 monoclonal antibodies for dog renal allografts was evaluated as a preliminary to tolerogenic studies in this large animal model. Monoclonal antibodies were given for a maximum of 10 days after transplantation. Therapy was stopped prematurely following adverse reactions associated with the recipient developing an antibody response against the foreign (rat) therapeutic monoclonal antibody. Blood trough levels of CD4 and CD8 antibodies indicated that saturating doses were achieved. Although neither CD4 nor CD8 alone prolonged allograft survival (rejection by day 7), combination of CD4 and CD8 antibodies resulted in good graft function for a median of 14 days. The effect of removing circulating T lymphocytes was also assessed using a lytic Thy-1 monoclonal antibody. Alone Thy-1 had little effect but, when combined with CD4, the median allograft survival time was increased to 15.5 days. Reduction of the number of circulating T lymphocytes appears complementary to blockade of CD4 for immunosuppression, while blockade of CD4 combined with removal of CD8 also favours allograft survival.

Immunomodulation of dog islets using a cocktail of monoclonal antibodies.

Islet allografts are particularly vulnerable to rejection, and current immunosuppressive agents are deleterious to their function. They are, however, highly suitable for 'immunomodulation', i.e., the removal or inactivation of passenger leukocytes to reduce their immunogenicity. For this purpose we have used 3 rat anti-dog monoclonal antibodies (Mabs) which are synergistic for leukocytolysis in the presence of autologous dog serum. Spleen cells or purified islets treated with these Mabs together with autologous serum were tested in mixed leukocyte and islet co-culture assays. The stimulatory properties of the Mab-pretreated splenocytes or islets were markedly reduced; moreover, the Mab cytolytic activity was shown to be confined to the leukocyte target cells and did not affect islet secretory function upon glucose stimulation. We conclude that this method of modifying the immunogenicity of dog islets could lead to successful islet grafting in vivo, allowing the reduction of conventional immunosuppression. Successful in vivo studies in this model, which are currently in progress, could have implications for clinical islet transplantation.

TNF staining of graft biopsy in renal transplantation.

Tumour Necrosis Factor (TNF) is a cytokine which may be found in patients' plasma and urine in association with acute rejection in renal transplantation. TNF is produced mainly by macrophage/monocytes and activated lymphocytes and its release in acute rejection may damage the nephron leading to renal dysfunction. However localization of TNF in renal grafts has not yet been demonstrated. We investigated TNF localization in renal graft tissue and the association with acute rejection compared with non-immunological events (cyclosporine toxicity and acute tubular necrosis) in graft biopsy.

Soluble HLA antigens in the circulation of liver graft recipients.

Human liver allografts deliver soluble class I (HLA-A and -B) transplantation antigens into the recipients' circulation. These molecules are detectable in recipient serum shortly after transplantation and they persist at high concentration for as long as the liver graft functions. Levels of graft-derived antigens in the recipient serum and self antigens in donor serum are comparable. Kinetic studies of these soluble antigens in donor and recipient show that donor antigens are continuously exported by the transplanted liver, while in the recipient, self antigens are derived from liver and other sources. At least two molecular forms of soluble HLA-A and -B antigens are present in sera from donors, recipients, and normal individuals. One form with m.w. approximately 50 kd seems to be a soluble monomer associated with beta 2-microglobulin, while the other forms of higher m.w. may be aggregates and/or complexes. Monitoring of these antigens in transplant recipients may be a useful indicator of graft pathology and function.