Diverse T Cell Responses Lead To The Different Manifestations Of Cutaneous Graft-Versus-Host Disease

Graft-versus-host disease (GVHD) is a major complication of allogeneic hematopoietic stem cell transplantation (HCT). GVHD is classified into an acute (aGVHD) and a chronic form (cGVHD) based on NIH-defined, clinical criteria. In skin, aGVHD presents as a maculopapular rash, while cGVHD shows at the beginning the signs of lichenoid tissue inflammation (chronic inflammatory GVHD, ciGVHD) and, later, exhibits scleroderma-like features (chronic sclerotic GVHD, csGVHD).

Mechanisms involved in the initiation and propagation of the different forms of GVHD are a main focus of HCT research, but remain controversially discussed. Acute GVHD has long been thought to mainly involve a type 1 T cell response, but more recent data have shown an involvement of Th2, Th17 and Treg cells. These T cell subsets, and particularly an altered regulatory T cell homeostasis, have also been reported to play a role in chronic GVHD, the pathogenesis of which is much less understood. Taken together, it is unclear which exact mechanisms are operative in the T cell immunopathogenesis of cutaneous GVHD.

In this study, we searched for cellular and molecular patterns that would allow us to distinguish between the different forms of cutaneous GVHD and to draw conclusions about the pathomechanisms involved. To address this issue, we collected lesional skin biopsies from patients diagnosed with aGVHD (n=25), ciGVHD (n=17) and csGVHD (n=7). Clinical grading of acute and chronic GVHD was performed according to the modified Glucksberg criteria and the NIH consensus criteria, respectively. 14 HCT recipients underwent serial skin biopsies at different time points prior/after HCT (prior/after conditioning, day 20-30 post-HCT, day 100-120 post-HCT). Skin biopsies obtained were subjected to phenotypic and functional analyses. The cellular infiltrate was assessed by immunofluorescence stainings; interleukins and chemokines were measured by real-time RT-PCR. Cytokine secretion profiles of stimulated T cells from collagenase-digested GVHD skin biopsies were analyzed by intracellular flow cytometry.

While CD4⁺ and CD8⁺ T cells dominated the inflammatory infiltrate in both acute and chronic GVHD skin lesions, the analysis of the quality of the T cell-mediated immune response revealed striking differences. In acute cutaneous GVHD, we found a “Th2 signature” as evidenced by highly increased transcripts of Th2 cytokines (IL-4, IL-5 and IL-13) and chemokines (CCL17, CCL20). Surprisingly, we also observed a strong up-regulation of IL-22, but not IL-17 mRNA, which was paralleled by an increase of IL-22-single-producing CD4⁺ T cells. IL-17/IL-22 double-positive T cells were almost absent. In sharp contrast to aGVHD, the immune response occurring in ciGVHD showed a mixed Th1/Th17 signature. This was evidenced by an up-regulation of Th1/Th17 cytokine (IFN-γ, IL-12/IL-23p40 and IL23p19) and chemokine transcripts (CCL5, CCR5, CXCL9, CXCL10). In accordance with this finding, increased numbers of IFN-ү- and IL-17-single-producing CD8⁺ T cells infiltrated ciGVHD skin lesions.Chronic sclerotic GVHD exhibited distinct features, including an abundance of mast cells and a down-regulation of T cell cytokines and chemokines relative to normal skin.

In an attempt to identify a factor predicting the occurrence of acute cutaneous GVHD, we analyzed the cellular and molecular signatures in skin of HCT recipients before GVHD onset. In accordance with the finding of a Th2 response occurring in aGVHD, levels of TSLP, a keratinocyte-derived cytokine skewing the immune response towards a Th2 direction, were increased at day 20 after HCT in non-lesional skin of patients who would later develop aGVHD.

In conclusion, the present study provides new insights into the diverse pathomechanisms operative in the acute and chronic form of cutaneous GVHD, respectively. Our findings allow to more accurately distinguish aGVHD from ciGVHD based on different cellular and molecular patterns. The analysis of TSLP expression in skin (before disease onset) could be helpful in identifying HCT recipients at risk for developing acute cutaneous GVHD.