Cytokine Detection and Modulation in Acute Graft vs. Host Disease in Mice

A murine model for acute lethal graft vs. host disease (GVHD) was used to study the role that a number of cytokines play in the development of lethal GVHD. In this study we focused on the role of IL-1, IL-2, IL-4, IL-6, IFN-γ and TNF-α. Lethally irradiated (C57BL × CBA)F1 mice were reconstituted either with 107 allogeneic BALB/c spleen cells or with a similar number of syngeneic cells, as a control. A significant rise in serum levels of IL-6, TNF-α and IFN-γ levels was found in allogeneically reconstituted mice. This is in contrast to the syngeneic control group in which no rise was seen. Serum IL-2 and IL-4 levels were below the detection limit. In the supernatant of Con A stimulated spleen cells from allogeneically reconstituted mice IL-6, IFN-γ and TNF-α concentrations were increased. The expression of mRNA for cytokines as detected by reverse transcription PCR was studied in spleen cells. In the allogeneic reconstituted mice the mRNA expression of IL-1α, IL-2, IL-6, IFN-γ and TNF-α displayed faster kinetics compared with that in syngeneic reconstituted mice. The effect of treatment with recombinant cytokines, antibodies to cytokines and to cytokine receptors on the development of GVHD was investigated. Administration of recombinant IL-2 to allogeneically reconstituted mice strongly increased the morbidity and mortality whereas injection of IL-1α and TNF-α did not influence survival. Administration of antibodies against IL-2 or the IL-2 receptor decreased the morbidity and mortality. Anti-IL-6, anti-IFN-γ, and anti-TNF-α mAB, on the other hand, did not affect the morbidity and mortality of GVHD. The results of this study suggest successive waves of cytokine-secreting cell populations consistent with the induction of an inflammatory response in the development of acute GVH disease

Suppression of graft-versus-host reactivity by a single host-specific blood transfusion to prospective donors of hemopoietic cells

Delayed-type hypersensitivity responses against recipient's histocompatibility antigens can occur early in the course of a graft-versus-host reaction in lethally irradiated allogeneically reconstituted mice. This reactivity could be suppressed by a single host-specific blood transfusion to the prospective donors of allogeneic spleen cells. Maximum suppression was found when the blood transfusion was given 4 or 5 days before the mice were used to reconstitute lethally irradiated hosts. Whole blood and purified white blood cells were capable of inducing suppression, whereas purified red blood cells, plasma, and serum were not. Suppression was already detectable after administration of 1 microliters of whole blood and virtually complete at a dose of 1.0 ml. Irradiation of the blood reduced but did not abrogate its capacity to induce suppression. Purified B and purified T lymphocytes appeared equally effective in inducing suppression. Two helper T cells clones, a Th1 and a Th2 clone, were able to induce suppression as well. A high dose of recombinant IL-2, injected daily for 5 days after reconstitution, did not abrogate or reduce the suppression. Suppression could be induced by H-2 as well as non-H-2 alloantigens, separately or together. A pure H-2-incompatible transfusion was more effective in inducing suppression than a pure non-H-2-incompatible one. Suppression appeared to be a dominant phenomenon and was mediated by a Thy-1 , CD4 , CD8- spleen cell population. This T cell population had its origin in the transfused donor, which excludes the possible involvement of blood-derived "veto-cells.

Prevention of lethal graft-versus-host disease in mice by monoclonal antibodies directed against T cells or their subsets.I.Evidence for the induction of a state of tolerance based on suppression

Lethal GVHD in the fully allogeneic BALB/c (donor)-(C57BL x CBA)F1 (recipient) mouse strain combination could be prevented by a single dose of IgG2b monoclonal antibodies (moAb) directed to T cells. The influence of the time of administration of this moAb after GVHD induction and the effect of anti-T cell subset moAb on the development of GVHD was investigated in this study. Moreover, the state of tolerance in the mice that had become long-term chimeras was examined. Anti-Thy-1 treatment of the recipients 1 day before, 2 h before or 1 day after reconstitution almost completely prevented lethal GVHD. A single dose of 100 micrograms of anti-Thy-1 was as effective as four daily doses of 25 micrograms each. Treatment with a single dose of 25 micrograms or with intervals of 4 days between doses of 25 micrograms was statistically significantly less effective. We injected the recipients with moAb directed to the CD4 or CD8 T cells subsets. Using a dose of 100 micrograms moAb, anti-CD4 treatment appeared to be less effective than anti-Thy-1 treatment whereas anti-CD8 treatment was not effective at all. A double dose of anti-CD4 was equally effective as anti-Thy-1 treatment. All mice that became long term survivors remained free of signs of GVHD and were > 99% repopulated with donor type cells. Injection of spleen cells from these BALB/c into (C57BL x CBA)F1 chimeric mice was used to reconstitute lethally irradiated BALB/c, BALB.K and (C57BL x CBA)F1 recipients. Lethal GVHD developed in the BALB.K and (C57BL x CBA)F1 recipients but not in the BALB/c recipients

Cytokines in lethal graft-versus-host disease

Graft-versus-host disease (GVHD) is caused by donor T lymphocytes that recognize foreign antigens on host tissues. This leads to T cell activation, which involves a cascade of events including the transcription of genes for cytokines and their receptors and the production of cytokines. One of the first cytokines to appear is interleukin 2 (IL-2). IL-2 production enhances the IL-2 receptor expression and leads to T cell proliferation. As a further step, differentation of T cells occurs, which results in the production of a certain pattern of cytokines. These cytokines influence the expression of cell surface antigens and adhesion molecules, and are able to activate other cell types such as cytotoxic T cells, macrophages and natural killer cells, which might act as effector cells in tissue destruction. Insight into the sequential expression of the various cytokines involved might enable a more effective treatment of GVHD. Therefore, we investigated the occurrence of cytokines in a murine model for acute GVHD. We addressed in particular the period early after allogeneic reconstitution

Prevention of lethal graft-versus-host disease in mice by monoclonal antibodies directed against T cells or their subsets.I.Evidence for the induction of a state of tolerance based on suppression

Lethal GVHD in the fully allogeneic BALB/c (donor)-(C57BL x CBA)F1 (recipient) mouse strain combination could be prevented by a single dose of IgG2b monoclonal antibodies (moAb) directed to T cells. The influence of the time of administration of this moAb after GVHD induction and the effect of anti-T cell subset moAb on the development of GVHD was investigated in this study. Moreover, the state of tolerance in the mice that had become long-term chimeras was examined. Anti-Thy-1 treatment of the recipients 1 day before, 2 h before or 1 day after reconstitution almost completely prevented lethal GVHD. A single dose of 100 micrograms of anti-Thy-1 was as effective as four daily doses of 25 micrograms each. Treatment with a single dose of 25 micrograms or with intervals of 4 days between doses of 25 micrograms was statistically significantly less effective. We injected the recipients with moAb directed to the CD4 or CD8 T cells subsets. Using a dose of 100 micrograms moAb, anti-CD4 treatment appeared to be less effective than anti-Thy-1 treatment whereas anti-CD8 treatment was not effective at all. A double dose of anti-CD4 was equally effective as anti-Thy-1 treatment. All mice that became long term survivors remained free of signs of GVHD and were > 99% repopulated with donor type cells. Injection of spleen cells from these BALB/c into (C57BL x CBA)F1 chimeric mice was used to reconstitute lethally irradiated BALB/c, BALB.K and (C57BL x CBA)F1 recipients. Lethal GVHD developed in the BALB.K and (C57BL x CBA)F1 recipients but not in the BALB/c recipients