SMAD3 prevents graft-versus-host disease by restraining Th1 differentiation and granulocyte-mediated tissue damage

Abstract

Gene expression profiling of human donor T cells before allogeneic hematopoietic cell transplantation revealed that expression of selected genes correlated with the occurrence of graft-versus-host disease (GVHD) in recipients. The gene with the best GVHD predictive accuracy was SMAD3, a core component of the transforming growth factor-␤ signaling pathway, whose expression levels vary more than a 6-fold range in humans. The putative role of SMAD3 in the establishment of graft-host tolerance remained elusive. We report that SMAD3-KO mice present ostensibly normal lymphoid and myeloid cell subsets. However, the lack of SMAD3 dramatically increased the frequency and severity of GVHD after allogeneic hematopoietic cell transplantation into major histocompatibility complex-identical recipients. Lethal GVHD induced by SMAD3-KO donors affected mainly the intestine and resulted from massive tissue infiltration by T-bet ؉ CD4 T cells and granulocytes that caused tissue damage by in situ release of Th1 cytokines and oxidative-nitrosative mediators, respectively. Our report reveals the nonredundant roles of SMAD3 in the development of tolerance to the host. Furthermore, our data support the concept that SMAD3 levels in donor cells dictate the risk of GVHD and that SMAD3 agonists would be attractive for prevention of GVHD. (Blood. 2011;117(5):1734-1744) IntroductionAfter decades of intensive research, graft-versus-host disease (GVHD) remains the unrelenting nemesis of patients and physicians involved in allogeneic hematopoietic cell transplantation (AHCT). 1-3 A better understanding of GVHD pathophysiology could lead to a more widespread use of AHCT, an otherwise potent treatment for hematologic malignancies and other less common diseases. GVHD is a complex immunopathology that hinges on recognition of host alloantigens by donor T cells. [4][5][6] Importantly, although histoincompatibility between donor and recipient is necessary, it is not sufficient to elicit GVHD. Of particular relevance, some donors are more dangerous than others. Indeed, a comprehensive study of CD4 and CD8 T cells from 50 human donor-recipient pairs revealed that pre-AHCT gene expression profiling segregates donors whose recipient did or did not have GVHD. 7 The gene with the best GVHD predictive accuracy was SMAD3. No AHCT recipients had GVHD when their donor cells expressed high levels of SMAD3 transcripts. 7 SMAD3 mRNA levels are stable over time in a given person but present substantial interindividual differences as they were found to vary more than a 6-fold range in a cohort of 397 subjects. 8 At least in transfected cells, SMAD3 protein levels correlate with expression of SMAD3 transcripts and with the strength of SMAD3 signaling. 9-11 Moreover, TCR signaling leads to commensurate decreases in expression of SMAD3 transcripts and phosphorylated SMAD3 protein. 12 Proteins of the SMAD family are homologous to proteins encoded by the Caenorhabditis elegans gene Sma and the Drosophila gene Mad. SMAD3 is one of the 2 "receptor-SMADs" that transduce tr...

Smad3-Deficient CD11b+Gr1+ Myeloid-Derived Suppressor Cells Prevent Allograft Rejection via the Nitric Oxide Pathway

“…Consistent with these results, our studies and others have shown that deletion of Smad3-dependent TGF-b signaling in T cells promotes severe graft-versus-host disease (our unpublished data and Ref. 38). In our investigation, Smad3-deficient CD4 + CD25 2 T cells were resistant to TGF-b-induced differentiation into CD4 + CD25 + Tregs in vitro, as expected.…”

Development and Function of Innate Polyclonal TCRαβ+ CD8+ Thymocytes

“…Analysis of cell surface markers and intracellular cytokines was performed by flow cytometry, as described (18). For in vitro-proliferation assays, sorted thymocytes were labeled with 5 mg/ml eFluor 670 for 10 min at 37˚C, washed once with complete medium, and washed three times with PBS.…”

Altered homeostatic regulation of innate and adaptive immunity in lower gastrointestinal tract GVHD pathogenesis

“…High IDO expression depletes tryptophan, resulting in apoptosis of alloreactive donor T effectors and generation of iTregs (107,108). Aside from these direct immune regulatory activities, IDO-generated tryptophan by-products may also suppress GVHD directly (89). Clinical studies showed that lower levels of urinary 3-indoxyl sulfate, an indole metabolite influenced by commensal bacteria, correlated with decreased patient survival after allo-HCT (109).…”

“…Transfer of TLR-expressing neutrophils enhanced GVHD mortality compared with the decreased mortality found after the transfer of TLR-deficient neutrophils. The transfer of BM cells lacking SMAD3, a critical signaling component of the TGF-β axis, greatly increased GVHD lethality, which correlated with enhanced Th1 polarization of donor T cells and neutrophil recruitment to the GI tract, mesenteric LNs, and spleen (89). Depleting myeloid cells using anti-Gr-1 mAb therapy greatly improved the outcome of mice receiving SMAD-deficient BM (89).…”