Effector memory T cells (T(EM)) do not cause graft-versus-host disease (GVHD), though why this is has not been elucidated. To compare the fates of alloreactive naive (T(N)) or memory (T(M)) T cells, we developed a model of GVHD in which donor T cells express a transgene-encoded TCR specific for an antigenic peptide that is ubiquitously expressed in the recipient. Small numbers of naive TCR transgenic (Tg) T cells induced a robust syndrome of GVHD in transplanted recipients. We then used an established method to convert TCR Tg cells to T(M) and tested these for GVHD induction. This allowed us to control for the potentially different frequencies of alloreactive T cells among T(N) and T(M), and to track fates of alloreactive T cells after transplantation. T(EM) caused minimal, transient GVHD whereas central memory T cells (T(CM)) caused potent GVHD. Surprisingly, T(EM) were not inert: they, engrafted, homed to target tissues, and proliferated extensively, but they produced less IFN-γ and their expansion in target tissues was limited at later time points, and local proliferation was reduced. Thus, cell-intrinsic properties independent of repertoire explain the impairment of T(EM), which can initiate but cannot sustain expansion and tissue damage.
Effector memory T cells (T) are less capable of inducing graft-versus-host disease (GVHD) compared with naive T cells (T). Previously, in the TS1 TCR transgenic model of GVHD, wherein TS1 CD4 cells specific for a model minor histocompatibility Ag (miHA) induce GVHD in miHA-positive recipients, we found that cell-intrinsic properties of TS1 T reduced their GVHD potency relative to TS1 T Posttransplant, TS1 T progeny expressed higher levels of PD-1 than did TS1 T progeny, leading us to test the hypothesis that T induce less GVHD because of increased sensitivity to PD-ligands. In this study, we tested this hypothesis and found that indeed TS1 T induced more severe skin and liver GVHD in the absence of PD-ligands. However, lack of PD-ligands did not result in early weight loss and colon GVHD comparable to that induced by TS1 T, indicating that additional pathways restrain alloreactive T TS1 T also caused more severe GVHD without PD-ligands. The absence of PD-ligands on donor bone marrow was sufficient to augment GVHD caused by either T or T, indicating that donor PD-ligand-expressing APCs critically regulate GVHD. In the absence of PD-ligands, both TS1 T and T induced late-onset myocarditis. Surprisingly, this was an autoimmune manifestation, because its development required non-TS1 polyclonal CD8 T cells. Myocarditis development also required donor bone marrow to be PD-ligand deficient, demonstrating the importance of donor APC regulatory function. In summary, PD-ligands suppress both miHA-directed GVHD and the development of alloimmunity-induced autoimmunity after allogeneic hematopoietic transplantation.
NFAT activating protein with ITAM motif 1 (NFAM1) is an ITAM bearing-transmembrane receptor that has been reported to play a role in B cell signaling and development. We performed expression analysis of NFAM1 using publicly available gene expression data sets and found that NFAM1 expression is significantly induced in intestinal biopsies from Crohn’s disease (CD) and ulcerative colitis (UC) patients. At the cellular level, we further observed high expression of NFAM1 in monocytes and neutrophils, and low expression in B and T cells. To explore the role of NFAM1 in multiple immune cells and its potential role in IBD, we generated NFAM1-/- mice. In contrast with previous reports using NFAM1-transgenic mice, NFAM1-/- mice have no obvious defects in immune cell development, or B cell responses. Interestingly, NFAM1-/- monocytes produce reduced levels of TNF-α in response to activation by multiple IBD-relevant stimuli, including CD40L, TLR ligands and MDP. Additional cytokines and chemokines such as IL-6, IL-12, CCL3 and CCL4 are also reduced in CD40L stimulated NFAM1-/- monocytes. Collectively, these findings indicate that NFAM1 promotes monocyte activation, thereby amplifying the response to diverse stimuli. Similarly, we observed that deletion of NFAM1 in human monocytes reduces expression of CD40L-induced CCL4. Lastly, to assess the role of NFAM1 in IBD, we compared development of anti-CD40 induced colitis in NFAM1+/+ and NFAM1-/- mice. We found that although NFAM1 deletion had no impact on development of gut pathology, we did observe a decrease in serum TNF-α, confirming that NFAM1 promotes pro-inflammatory cytokine production in vivo. Taken together, we conclude that NFAM1 functions to amplify cytokine production and should be further evaluated as a therapeutic target for treatment of autoimmune disease.
Graft-versus-host disease (GVHD) limits the broader application of allogeneic hematopoietic stem cell transplantation. In prior studies we defined roles for both host and donor-derived antigen presenting cells (APCs) in the activation of alloreactive donor T cells and in promotion of GVHD. While initial T cell activation in GVHD occurs predominantly in secondary lymphoid organs, we have consistently observed MHCII+ donor-derived APCs, including dendritic cells (DCs), in histopathologic GVHD lesions, frequently adjacent to infiltrating T cells, suggesting they have a role in local GVHD reactions. Donor-derived tissue APCs (t-APCs), including tissue-DCs (t-DCs) could activate donor T cells through indirect or cross-presentation of host antigens, produce chemokines that recruit other effectors, and elaborate inflammatory mediators or suppressors of inflammation. We first characterized t-DC subsets in the skin and bowel of GVHD-affected mice. 129 (H-2b) hosts were irradiated and reconstituted with B6 (H-2b) BM with or without CD4+ and CD8+ T cells to induce GVHD and analyzed mononuclear cells from skin and bowel approximately 4 weeks post transplant. In skin, both main dermal DC populations (CD11b+ and CD103+) were significantly increased in GVHD mice as compared to BM alone controls, though the ratios of CD11b+: CD103+ DCs were similar. In the bowel lamina propria, the ratios of CD11b+CD103- to CD11b+CD103+ were increased in GVHD mice in the colon but were similar to that in BM alone controls in the small bowel. We next studied the roles of CCR6 and CCR2 in the recruitment of donor-derived APCs to skin and bowel. We transplanted mice with CCR6-/- BM in competition with wild type (wt) BM and found that the contribution of each to skin and bowel APCs matched their contributions to myeloid hematopoiesis in BM, spleen and blood, indicating that CCR6 is not required. To study the role of CCR2 we first compared mice transplanted with either wt or CCR2-/- BM with wt T cells. Despite having a profound reduction in blood monocytes, all skin and bowel t-APC subsets were present in CCR2-/- recipients, indicating that CCR2 is not required for t-APC recruitment in contrast to its role in many other models of inflammation. However, CD103+ DCs were more prevalent relative to CD11b+ DCs, consistent with a pre-cDC origin. Despite monocytopenia, recipients of CCR2-/- BM developed clinical GVHD; histology data is being analyzed and will be presented. To better define the contributions of CCR2 to t-APC recruitment and to determine monocyte versus pre-cDC origin of t-DCs, we transplanted mice with CCR2-/- BM in competition with wt BM and compared ratios of BM and blood precursors (pre-cDCs and monocytes) to t-DC ratios. For CD103+ DCs, wt/KO ratios matched the ratios of general myeloid hematopoiesis and pre-cDCs, indicating a pre-cDC origin. For CD11b+CD103- DCs, the ratio of wt/KO matched that in blood monocytes. We further subsetted CD11b+ t-DCs based on the expression of Ly6C, MAR1, CD64 and CD24, used to differentiate pre-cDC from mono-derived DCs in other organs, and did not identify any population with wt/KO ratios that did not match that of the general CD11b+ DC population, suggesting that most if not all CD11b+ t-DCs are of monocyte origin. Experiments are underway examining the role of CX3CR1 in t-APC recruitment and these data will be presented. Disclosures: No relevant conflicts of interest to declare.
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