Allogeneic BMT is a potentially curative therapeutic option for many malignant and non-malignant diseases. However, the beneficial effects of the donor immune system on engraftment and elimination of residual malignant cells are closely linked to the occurrence of aGvHD which leads to treatment related mortality. Treg have been shown to reduce the incidence and severity of aGvHD in rodent models [21
] and increased Foxp3+
Treg cells in the hematopoietic graft were shown to predict a reduced risk for aGvHD in human transplant recipients [32
]. Expansion and function of Treg relies on IL-2 [25
] while the role of other IL-2Rγ-chain dependent cytokines such as IL-7 and IL-15 are still under investigation [36
]. Insight into the cytokine milieu that can favor Treg function may help to manipulate the function of this cell population. In this report we evaluated the role of IL-18 on aGvHD and the differential impact on Tconv and Treg expansion. Interestingly we found that expression of the IL-18Rα was similar in Treg and Tconv isolated from different organs. Since in the in vitro studies IL-18 from the stimulators and irradiated non-Treg responders was present we did not address the functional role of IL-18 on Treg suppressor function but showed that Treg which developed in the absence of IL-18 are equally suppressive as their wt counterparts.
Our in vivo data indicate that ontogeny, function and homeostasis of Treg remain intact in the absence of IL-18 in healthy mice. In a model of oral tolerance induction, IL-18 deficient recipients did not develop Treg in Peyer’s patches after feeding with BLG [23
]. The observation of Tsuji et al. that IL-18 was relevant for Treg mediated oral tolerance induction could be due to the requirement of the cytokine for Treg in the situation where an immune response requires regulation [23
]. We therefore investigated the relevance of IL-18 for Treg and Tconv during aGvHD as an aberrant immune response. Our observation that donor deficiency for IL-18 did not affect CD4 T cell expansion and the course of aGvHD made us hypothesize that production of IL-18 by the host may be more relevant for the previously reported increased IL-18 levels after BMT [3
Indeed we found IL-18 levels to be significantly decreased when the recipient but not the donor animals were IL-18 deficient. This suggested that in the early phase after transplantation residual host hematopoietic cells such as macrophages, Kupffer cells and dendritic cells, that can survival the first week after the myeloablative irradiation, produce significant amounts of IL-18. Other recipient type cells with known ability to produce IL-18 are intestinal epithelial cells and keratinocytes [4
]. These recipient cells may secrete IL-18 triggered by tissue damage due to the conditioning regimen. Interestingly, IL-18 in contrast to other cytokines is stored as biologically inactive precursor (pro-IL-18) and is secreted when appropriate cleaving enzymes are present [37
]. Previous studies have demonstrated the involvement of a caspase-1-like molecule and caspase-1 for cleavage of IL-18 [4
]. Lipopolysaccharide (LPS), that enhances aGvHD severity and leaks into the blood stream after conditioning induced bowel wall damage [40
], has been shown to induce caspase-1 which is critical in the process of IL-18 secretion [41
]. Our observation that increased IL-18 serum levels after BMT are host derived is compatible with the finding that caspase-1-deficient recipients engrafted with wildtype H-2 disparate splenocytes displayed decreased IL-18 production [4
]. Our study is the first to delineate the relative contribution of the host on increased IL-18 cytokine levels during the early phase after BMT that has a functional impact on the aGvHD course. With respect to the clinical situation, the manipulation of GvHD through the cytokine Il-18 may be difficult to achieve based on its opposing effects on CD4 as compared to CD8 T cells [17
] and due to the fact that human GvHD is not restricted to either subset as it can be studied in murine models with selective MHC class I or class II differences.
Since IL-18 has been demonstrated to induce IFN-γ production [7
], a cytokine with known relevance for downmodulation of aGvHD in the early phase after BMT [8
] and for long-term allograft survival [42
] we studied the impact of host IL-18 deficiency on IFN-γ secretion. We found that reduced IL-18 production in IL-18 deficient recipients was paralleled by diminished IFN-γ serum levels. This finding may explain why the absence of host IL-18 resulted in more aggressive aGvHD, since IFN-γ was shown to induce death of activated donor CD4 cells [17
]. IL-18 may mediate its effects not only directly by augmentation of IFN-γ production [43
] but also by increasing the expression of the IL-12R complex, thereby enhancing the effects of IL-12 [45
]. Interestingly, IL-12 has been demonstrated to be critical for tolerance induction in solid organ transplantation [46
] and to reduce aGvHD [12
]. A recent in vivo study demonstrated a unique role for IFN-γ in the functional activity of alloantigen-reactive Treg cells during the development of operational tolerance to donor skin allografts [47
]. In our studies the absence of host IL-18 was paralleled by reduced IFN-γ and we did not find any change in Treg proliferation under these conditions. Further studies are needed to address the role of reduced IFN-γ levels on Treg suppressor function in the BMT model.
Since the expansion kinetics of Treg were independent of host IL-18 the protective effect of host IL-18 on aGvHD was not through immunoregulation by increased Treg expansion. In contrast to data from the oral tolerance model [23
], we did not find an essential role of IL-18 for the induction of Treg cells which may be due to model specific differences.
We conclude that Treg and Tconv express comparable levels of IL-18Rα which is upregulated in response to alloantigen. Absence of host but not donor IL-18 production translated into lower IFN-γ levels in the early phase after transplantation and resulted in a more rapid course of aGvHD. Host derived IL-18 has a protective effect in CD4+ mediated aGvHD, possibly through increased IFN-γ production. IL-18 is non-essential for Treg expansion in an allogeneic environment however downmodulates CD4+ Tconv expansion. In the absence of IL-18 increased CD4+ T cell proliferation results in more aggressive aGvHD.