Downregulation of intragraft IFN-gamma expression correlates with increased IgG1 alloantibody response following intrathymic immunomodulation of sensitized rat recipients.

Abstract

A single intrathymic injection of donor-specific spleen cells (2 x 10(7)) abrogates accelerated (24 hr) rejection of LBNF1 cardiac allografts in presensitized LEW rats and prolongs graft survival to about 11 days. This effect is donor-specific, gamma-irradiation-sensitive, thymus-dependent, and requires no concomitant therapy. We have recently shown that following intrathymic alloantigen administration, there is an earlier and increased systemic production of alloreactive IgM, and subsequently a premature isotype switching to IgG with the predominant IgG1 and IgG2a alloantibody responses. There is also a preferential binding of these IgG subclasses to the endothelium of well-functioning allografts. In this work, we analyzed the early cell activation and related cytokine elaboration patterns at the mRNA and protein levels by competitive template RT-PCR and immunohistochemistry, respectively. We found that prolonged cardiac allograft survival following intrathymic administration of donor spleen cells in presensitized rats was associated with markedly depressed intragraft IFN gamma mRNA and protein expression. Moreover, intrathymic allostimulation has led to a defect in the IL-2 pathway as the expression of IL-2 and IL-2R protein at the graft site was inhibited despite stable IL-2 mRNA levels. The inhibition of cell activation was also demonstrated by reduced MHC class II and the lack of ICAM-1, and TNF-alpha expression by immunohistochemistry. The expression of biologically active IL-12 (p70) by mononuclear and endothelial cells was detected in rejecting grafts between 3 and 12 hr. The well-functioning grafts after intrathymic allostimulation were devoid of IL-12 (p70), which in turn may have contributed to the downregulation of IFN-gamma mRNA and protein. Treatment with r.IFN-gamma, but not with r.IL-2, recreated the rejection response, and the characteristic IgG subclass pattern associated with accelerated graft loss. Hence, intrathymic immunomodulation with alloantigen results in selective inhibition of IFN-gamma-producing cells and a preferential upregulation of IgG1 alloantibodies. These data support the notion of the interlocked immunoregulatory roles of cytokine and alloantibody networks in rat allograft recipients.