Induction of Transplantation Tolerance Using Monoclonal Antibodies to CD4: Experimental Studies Using a Rat Heterotopic Cardiac Allograft Model

Ahmiedat, Hamdi H (1997) Induction of Transplantation Tolerance Using Monoclonal Antibodies to CD4: Experimental Studies Using a Rat Heterotopic Cardiac Allograft Model. PhD thesis, University of Glasgow.

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A major goal in clinical organ transplantation is to devise new strategies for the induction of specific immunological tolerance to the allograft. This would obviate the need for long-term immunosuppressive drugs and leave the immune system of the recipient otherwise intact to protect the recipient from the development of infection and malignancy. CD4 T cells play a central role in allograft rejection and the CD4 molecule has therefore attracted considerable interest as a molecular target for manipulating the allograft response. In this thesis, mouse monoclonal antibodies (mAbs) directed against the rat CD4 molecule were used in an attempt to prolong allograft survival and possibly to promote tolerance to a heterotopic cardiac allograft in the rat. In preliminary studies, administration of the mouse IgG2a anti-CD4 mAb MRC OX38 (directed against an epitope on the distal domains of the rat CD4 molecule) for a brief period immediately prior to transplantation [10 mg/kg on day -3 and 2 mg/kg on day -2, -1 and day 0 (day of transplantation)] prolonged heart allograft survival indefinitely (>200 days) in the Lewis to DA rat strain combination. Long-standing allograft recipients displayed donor-specific tolerance by rejecting a second "third party" (PVG) heart graft but permanently accepting a second donor-specific (Lewis) heart graft. The ability of OX38 mAb to induce transplant tolerance was highly dependent on the rat strain combination studied. Although 0X38 mAb treatment consistently increased cardiac allograft survival in other fully allogeneic rat strains tested (e.g. DA to PVG and DA to Lewis), long-term survival was not observed. Further studies were performed with additional anti-CD4 mAbs directed against epitopes on the proximal (OX70, OX71 and OX73) or distal domains (W3/25) of the CD4 molecule. All of the mAb's tested, with the exception of W3/25, led to a marked increase in the survival of Lewis heart allografts in DA recipients and many recipients accepted their grafts permanently. The mAb W3/25 mAb depleted CD4 T cells to a lesser extent than the other anti-CD4 mAb tested, which may explain, at least in part, its inability to prolong graft survival indefinitely. Further studies were performed to investigate some of the possible mechanisms involved in the induction and maintenance of anti-CD4 induced transplant tolerance. The ability of anti-CD4 mAbs to prolong cardiac allograft survival was significantly dependent on the presence of the thymus gland, since thymectomised DA recipients treated with OX38 or OX70 mAb did not become tolerant to Lewis strain heart grafts. Administration of exogenous recombinant interleukin-2 (rIL-2) during the induction of tolerance by anti-CD4 mAbs (OX38 and OX73) failed to abrogate induction of tolerance, suggesting that a lack of endogenous IL-2 alone does not account for allograft survival after anti-CD4 therapy. On the other hand, treatment with a neutralising anti-IL-4 mAb (MRC OX81) during the induction of tolerance with 0X38 mAb reduced the efficacy of tolerance induction. Although not conclusive, this finding suggests a possible role for IL-4 during the induction phase of tolerance following anti-CD4 mAbs therapy. The role of CDS cells in the induction and maintenance of transplant tolerance was also examined. Administration of the depleting anti-CD8 mAb (OX8) during the induction of tolerance abrogated the ability of anti-CD4 mAb to increase cardiac allograft survival but when OX8 mAb was given to animals bearing long-term tolerant heart grafts it did not prevent such animals from accepting a second donor strain heart indefinitely. Thus CD8 cells appear to be necessary for induction but not maintenance of the tolerant state following anti-CD4 therapy. In further experiments adoptive transfer studies using splenocytes obtained from OX38 treated tolerant recipients provided evidence for the existence of immunoregulatory (or suppressor) lymphocytes. Thus, transfer of 2x108 spleen cells from tolerant animals to secondary unmodified graft recipients enabled them (in 2 of 4 recipients) to maintain their graft indefinitely. Finally, purified CD4 T cells were obtained from animals bearing long-standing Lewis heart allografts following anti-CD4 mAb treatment (OX38 or OX70) to determine their in vitro proliferative response and cytokine production. CD4 T cells from OX38 treated recipients showed near normal levels of proliferation to irradiated donor (Lewis) stimulators in the in vitro MLR. Moreover, CD4 T cells from OX38 treated tolerant animals released levels of IFNgamma and EL-4 into the culture supernatants which were comparable to those produced in response to third party stimulators or released by CD4 T cells obtained from normal DA strain rats in response to Lewis stimulators cells.

Item Type: Thesis (PhD)
Qualification Level: Doctoral
Additional Information: Adviser: Andrew Bradley
Keywords: Medicine, Immunology
Date of Award: 1997
Depositing User: Enlighten Team
Unique ID: glathesis:1997-75856
Copyright: Copyright of this thesis is held by the author.
Date Deposited: 19 Dec 2019 09:15
Last Modified: 19 Dec 2019 09:15

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