Emerging Functions of IL-33 in Homeostasis and Immunity.

Our understanding of the functions of the IL-1 superfamily cytokine and damage-associated molecular pattern IL-33 continues to evolve with our understanding of homeostasis and immunity. The early findings that IL-33 is a potent driver of type 2 immune responses promoting parasite expulsion, but also inflammatory diseases like allergy and asthma, have been further supported. Yet, as the importance of a type 2 response in tissue repair and homeostasis has emerged, so has the fundamental importance of IL-33 to these processes. In this review, we outline an evolving understanding of IL-33 immunobiology, paying particular attention to how IL-33 directs a network of ST2+ regulatory T cells, reparative and regulatory macrophages, and type 2 innate lymphoid cells that are fundamental to tissue development, homeostasis, and repair.

Neuro-epithelial-ILC2 crosstalk in barrier tissues.

Group 2 innate lymphoid cells (ILC2s) contribute to the maintenance of mammalian barrier tissue homeostasis. We review how ILC2s integrate epithelial signals and neurogenic components to preserve the tissue microenvironment and modulate inflammation. The epithelium that overlies barrier tissues, including the skin, lungs, and gut, generates epithelial cytokines that elicit ILC2 activation. Sympathetic, parasympathetic, sensory, and enteric fibers release neural signals to modulate ILC2 functions. We also highlight recent findings suggesting neuro-epithelial-ILC2 crosstalk and its implications in immunity, inflammation and resolution, tissue repair, and restoring homeostasis. We further discuss the pathogenic effects of disturbed ILC2-centered neuro-epithelial-immune cell interactions and putative areas for therapeutic targeting.

Advancing mouse models for transplantation research.

Mouse models have been instrumental in understanding mechanisms of transplant rejection and tolerance, but cross-study reproducibility and translation of experimental findings into effective clinical therapies are issues of concern. The Mouse Models in Transplantation symposium gathered scientists and physician-scientists involved in basic and clinical research in transplantation to discuss the strengths and limitations of mouse transplant models and strategies to enhance their utility. Participants recognized that increased procedure standardization, including the use of prespecified, defined endpoints, and statistical power analyses, would benefit the field. They also discussed the generation of new models that incorporate environmental and genetic variables affecting clinical outcomes as potentially important. If implemented, these strategies are expected to improve the reproducibility of mouse studies and increase their translation to clinical trials and, ideally, new Food and Drug Administration-approved drugs.

Treg-inducing microparticles promote donor-specific tolerance in experimental vascularized composite allotransplantation.

For individuals who sustain devastating composite tissue loss, vascularized composite allotransplantation (VCA; e.g., hand and face transplantation) has the potential to restore appearance and function of the damaged tissues. As with solid organ transplantation, however, rejection must be controlled by multidrug systemic immunosuppression with substantial side effects. As an alternative therapeutic approach inspired by natural mechanisms the body uses to control inflammation, we developed a system to enrich regulatory T cells (Tregs) in an allograft. Microparticles were engineered to sustainably release TGF-ß1, IL-2, and rapamycin, to induce Treg differentiation from naïve T cells. In a rat hindlimb VCA model, local administration of this Treg-inducing system, referred to as TRI-MP, prolonged allograft survival indefinitely without long-term systemic immunosuppression. TRI-MP treatment reduced expression of inflammatory mediators and enhanced expression of Treg-associated cytokines in allograft tissue. TRI-MP also enriched Treg and reduced inflammatory Th1 populations in allograft draining lymph nodes. This local immunotherapy imparted systemic donor-specific tolerance in otherwise immunocompetent rats, as evidenced by acceptance of secondary skin grafts from the hindlimb donor strain and rejection of skin grafts from a third-party donor strain. Ultimately, this therapeutic approach may reduce, or even eliminate, the need for systemic immunosuppression in VCA or solid organ transplantation.

Interleukin-33 contributes to ILC2 activation and early inflammation-associated lung injury during abdominal sepsis.

Sepsis is defined as infection with organ dysfunction due to a dysregulated immune response. The lung is one of the most vulnerable organs at the onset of sepsis. Interleukin (IL)-33 can be released by injured epithelial and endothelial cells in the lung and regulate immune activation and infiltration. Therefore, we postulated that IL-33 would contribute to the immune response in the lung during sepsis. Using the cecal ligation and puncture (CLP) sepsis model, we show here that IL-33 contributes significantly to both sepsis-induced inflammation in the lung and systemic inflammatory response in the early phase of sepsis. Despite the higher intra-peritoneal bacterial burden, the absence of IL-33 resulted in less infiltration of neutrophils and monocytes into the lungs in association with lower circulating, lung and liver cytokine levels as well as reduced lung injury at 6 h after sepsis. IL-33 was required for the upregulation of IL-5 in type 2 Innate Lymphoid Cells (ILC2), while IL-5 neutralization suppressed neutrophil and monocyte infiltration in the lungs during CLP sepsis. This reduction in leukocyte infiltration in IL-33-deficient mice was reversed by administration of recombinant IL-5. These results indicate that IL-33 plays a major role in the local inflammatory changes in the lung, in part, by regulating IL-5 and this axis contributes to lung injury early after the onset of sepsis.

Peri-alloHCT IL-33 administration expands recipient T-regulatory cells that protect mice against acute GVHD.

During allogeneic hematopoietic cell transplantation (alloHCT), nonhematopoietic cell interleukin-33 (IL-33) is augmented and released by recipient conditioning to promote type 1 alloimmunity and lethal acute graft-versus-host disease (GVHD). Yet, IL-33 is highly pleiotropic and exhibits potent immunoregulatory properties in the absence of coincident proinflammatory stimuli. We tested whether peri-alloHCT IL-33 delivery can protect against development of GVHD by augmenting IL-33-associated regulatory mechanisms. IL-33 administration augmented the frequency of regulatory T cells (Tregs) expressing the IL-33 receptor, suppression of tumorigenicity-2 (ST2), which persist following total body irradiation. ST2 expression is not exclusive to Tregs and IL-33 expands innate immune cells with regulatory or reparative properties. However, selective depletion of recipient Foxp3(+) cells concurrent with peri-alloHCT IL-33 administration accelerated acute GVHD lethality. IL-33-expanded Tregs protected recipients from GVHD by controlling macrophage activation and preventing accumulation of effector T cells in GVHD-target tissue. IL-33 stimulation of ST2 on Tregs activates p38 MAPK, which drives expansion of the ST2(+) Treg subset. Associated mechanistic studies revealed that proliferating Tregs exhibit IL-33-independent upregulation of ST2 and the adoptive transfer of st2(+) but not st2(-) Tregs mediated GVHD protection. In total, these data demonstrate the protective capacity of peri-alloHCT administration of IL-33 and IL-33-responsive Tregs in mouse models of acute GVHD. These findings provide strong support that the immunoregulatory relationship between IL-33 and Tregs can be harnessed therapeutically to prevent GVHD after alloHCT for treatment of malignancy or as a means for tolerance induction in solid organ transplantation.

IL33-mediated ILC2 activation and neutrophil IL5 production in the lung response after severe trauma: A reverse translation study from a human cohort to a mouse trauma model.

BACKGROUND: The immunosuppression and immune dysregulation that follows severe injury includes type 2 immune responses manifested by elevations in interleukin (IL) 4, IL5, and IL13 early after injury. We hypothesized that IL33, an alarmin released early after tissue injury and a known regulator of type 2 immunity, contributes to the early type 2 immune responses after systemic injury. METHODS AND FINDINGS: Blunt trauma patients admitted to the trauma intensive care unit of a level I trauma center were enrolled in an observational study that included frequent blood sampling. Dynamic changes in IL33 and soluble suppression of tumorigenicity 2 (sST2) levels were measured in the plasma and correlated with levels of the type 2 cytokines and nosocomial infection. Based on the observations in humans, mechanistic experiments were designed in a mouse model of resuscitated hemorrhagic shock and tissue trauma (HS/T). These experiments utilized wild-type C57BL/6 mice, IL33-/- mice, B6.C3(Cg)-Rorasg/sg mice deficient in group 2 innate lymphoid cells (ILC2), and C57BL/6 wild-type mice treated with anti-IL5 antibody. Severely injured human blunt trauma patients (n = 472, average injury severity score [ISS] = 20.2) exhibited elevations in plasma IL33 levels upon admission and over time that correlated positively with increases in IL4, IL5, and IL13 (P < 0.0001). sST2 levels also increased after injury but in a delayed manner compared with IL33. The increases in IL33 and sST2 were significantly greater in patients that developed nosocomial infection and organ dysfunction than similarly injured patients that did not (P < 0.05). Mechanistic studies were carried out in a mouse model of HS/T that recapitulated the early increase in IL33 and delayed increase in sST2 in the plasma (P < 0.005). These studies identified a pathway where IL33 induces ILC2 activation in the lung within hours of HS/T. ILC2 IL5 up-regulation induces further IL5 expression by CXCR2+ lung neutrophils, culminating in early lung injury. The major limitations of this study are the descriptive nature of the human study component and the impact of the potential differences between human and mouse immune responses to polytrauma. Also, the studies performed did not permit us to make conclusions about the impact of IL33 on pulmonary function. CONCLUSIONS: These results suggest that IL33 may initiate early detrimental type 2 immune responses after trauma through ILC2 regulation of neutrophil IL5 production. This IL33-ILC2-IL5-neutrophil axis defines a novel regulatory role for ILC2 in acute lung injury that could be targeted in trauma patients prone to early lung dysfunction.

IL-33 exacerbates liver sterile inflammation by amplifying neutrophil extracellular trap formation.

BACKGROUND & AIMS: Neutrophils and liver sinusoidal endothelial cells (LSECs) both contribute to sterile inflammatory injury during ischemia/reperfusion (I/R), a well-known liver surgical stress. Interleukin-33 (IL-33) has been shown to drive neutrophil infiltration during inflammatory responses through its receptor ST2. We recently reported that infiltrating neutrophils form neutrophil extracellular traps (NETs), which exacerbate sterile inflammatory injury in liver I/R. Here, we sought to determine the role of IL-33 in NET formation during liver sterile inflammation. METHODS: Evaluation of IL-33 forming NETs was investigated using a partial liver I/R model to generate sterile injury in healthy WT, IL-33 and ST2 knockouts. Serum levels of IL-33 and myeloperoxidase (MPO)-DNA complex were measured in both humans and mice after the first surgery. Liver damage was assessed. Mouse neutrophil depletion was performed by intraperitoneal injection of anti-Ly6G antibody before I/R. RESULTS: Patients undergoing liver resection showed a significant increase in serum IL-33 compared to healthy volunteers. This coincided with higher serum MPO-DNA complexes. NET formation was decreased in IL-33 and ST2 knockout mice compared with control mice, after liver I/R. IL-33 or ST2 deficiency protected livers from I/R injury, whereas rIL-33 administration during I/R exacerbated hepatotoxicity and systemic inflammation. In vitro, IL-33 is released from LSECs to promote NET formation. IL-33 deficient LSECs failed to induce NETs. ST2 deficient neutrophils limited their capacity to form NETs in vitro and adoptive transfer of ST2 knockout neutrophils to neutrophil-depleted WT mice significantly decreased NET formation. CONCLUSIONS: Data establish that IL-33, mainly released from LSECs, causes excessive sterile inflammation after hepatic I/R by inducing NET formation. Therapeutic targeting of IL-33/ST2 might extend novel strategies to minimize organ damage in various clinical settings associated with sterile inflammation. LAY SUMMARY: Liver ischemia and reperfusion injury results in the formation of neutrophil extracellular traps, which contribute to organ damage in liver surgeries. Herein, we show that IL-33 is released from liver sinusoidal endothelial cells to promote NET formation during liver I/R, which exacerbates inflammatory cascades and sterile inflammation.

The IL-33/ST2 axis augments effector T-cell responses during acute GVHD.

Interleukin (IL)-33 binding to the receptor suppression of tumorigenicity 2 (ST2) produces pro-inflammatory and anti-inflammatory effects. Increased levels of soluble ST2 (sST2) are a biomarker for steroid-refractory graft-versus-host disease (GVHD) and mortality. However, whether sST2 has a role as an immune modulator or only as a biomarker during GVHD was unclear. We show increased IL-33 production by nonhematopoietic cells in the gastrointestinal (GI) tract in mice post-conditioning and patients during GVHD. Exogenous IL-33 administration during the peak inflammatory response worsened GVHD. Conversely, GVHD lethality and tumor necrosis factor-α production was significantly reduced in il33(-/-) recipients. ST2 was upregulated on murine and human alloreactive T cells and sST2 increased as experimental GVHD progressed. Concordantly, st2(-/-) vs wild-type (WT) donor T cells had a marked reduction in GVHD lethality and GI histopathology. Alloantigen-induced IL-18 receptor upregulation was lower in st2(-/-) T cells, and linked to reduced interferon-γ production by st2(-/-) vs WT T cells during GVHD. Blockade of IL-33/ST2 interactions during allogeneic-hematopoietic cell transplantation by exogenous ST2-Fc infusions had a marked reduction in GVHD lethality, indicating a role of ST2 as a decoy receptor modulating GVHD. Together, these studies point to the IL-33/ST2 axis as a novel and potent target for GVHD therapy.

mTORC2 Deficiency in Myeloid Dendritic Cells Enhances Their Allogeneic Th1 and Th17 Stimulatory Ability after TLR4 Ligation In Vitro and In Vivo.

The mammalian/mechanistic target of rapamycin (mTOR) is a key integrative kinase that functions in two independent complexes, mTOR complex (mTORC) 1 and mTORC2. In contrast to the well-defined role of mTORC1 in dendritic cells (DC), little is known about the function of mTORC2. In this study, to our knowledge, we demonstrate for the first time an enhanced ability of mTORC2-deficient myeloid DC to stimulate and polarize allogeneic T cells. We show that activated bone marrow-derived DC from conditional Rictor(-/-) mice exhibit lower coinhibitory B7-H1 molecule expression independently of the stimulus and enhanced IL-6, TNF-α, IL-12p70, and IL-23 production following TLR4 ligation. Accordingly, TLR4-activated Rictor(-/-) DC display augmented allogeneic T cell stimulatory ability, expanding IFN-γ(+) and IL-17(+), but not IL-10(+) or CD4(+)Foxp3(+) regulatory T cells in vitro. A similar DC profile was obtained by stimulating Dectin-1 (C-type lectin family member) on Rictor(-/-) DC. Using novel CD11c-specific Rictor(-/-) mice, we confirm the alloreactive Th1 and Th17 cell-polarizing ability of endogenous mTORC2-deficient DC after TLR4 ligation in vivo. Furthermore, we demonstrate that proinflammatory cytokines produced by Rictor(-/-) DC after LPS stimulation are key in promoting Th1/Th17 responses. These data establish that mTORC2 activity restrains conventional DC proinflammatory capacity and their ability to polarize T cells following TLR and non-TLR stimulation. Our findings provide new insight into the role of mTORC2 in regulating DC function and may have implications for emerging therapeutic strategies that target mTOR in cancer, infectious diseases, and transplantation.

Tumoral expression of IL-33 inhibits tumor growth and modifies the tumor microenvironment through CD8+ T and NK cells.

Cancer immunotherapy has shown great promise as a new standard cancer therapeutic modality. However, the response rates are limited for current approach that depends on enhancing spontaneous antitumor immune responses. Therefore, increasing tumor immunogenicity by expressing appropriate cytokines should further improve the current immunotherapy. IL-33 is a member of the IL-1 family of cytokines and is released by necrotic epithelial cells or activated innate immune cells and is thus considered a "danger" signal. The role of IL-33 in promoting type 2 immune responses and tissue inflammation has been well established. However, whether IL-33 drives antitumor immune responses is controversial. Our previous work established that IL-33 promoted the function of CD8(+) T cells. In this study, we showed that the expression of IL-33 in two types of cancer cells potently inhibited tumor growth and metastasis. Mechanistically, IL-33 increased numbers and IFN-γ production by CD8(+) T and NK cells in tumor tissues, thereby inducing a tumor microenvironment favoring tumor eradication. Importantly, IL-33 greatly increased tumor Ag-specific CD8(+) T cells. Furthermore, both NK and CD8(+) T cells were required for the antitumor effect of IL-33. Moreover, depletion of regulatory T cells worked synergistically with IL-33 expression for tumor elimination. Our studies established "alarmin" IL-33 as a promising new cytokine for tumor immunotherapy through promoting cancer-eradicating type 1 immune responses.

IL-33 is an unconventional Alarmin that stimulates IL-2 secretion by dendritic cells to selectively expand IL-33R/ST2+ regulatory T cells.

IL-33 is a recently characterized IL-1 family member that is proposed to function as an alarmin, or endogenous signal of cellular damage, as well as act as a pleiotropic cytokine. The ability of IL-33 to potentiate both Th1 and Th2 immunity supports its role in pathogen clearance and disease immunopathology. Yet, IL-33 restrains experimental colitis and transplant rejection by expanding regulatory T cells (Treg) via an undefined mechanism. We sought to determine the influence of IL-33 on hematopoietic cells that drives Treg expansion and underlies the therapeutic benefit of IL-33 administration. In this study, we identify a feedback loop in which conventional mouse CD11c(+) dendritic cells (DC) stimulated by IL-33 secrete IL-2 to selectively expand IL-33R(ST2(+))- suppressive CD4(+)Foxp3(+) Treg. Interestingly, this occurs in the absence of classical DC maturation, and DC-derived (innate) IL-2 increases ST2 expression on both DC and interacting Treg. ST2(+) Treg represent an activated subset of Foxp3(+) cells, demonstrated to be ICOS(high)CD44(high) compared with their ST2(-) counterparts. Furthermore, although studies have shown that IL-33-exposed DC promote Th2 responses, we reveal that ST2(+) DC are required for IL-33-mediated in vitro and in vivo Treg expansion. Thus, we have uncovered a relationship between IL-33 and innate IL-2 that promotes the selective expansion of ST2(+) Treg over non-Treg. These findings identify a novel regulatory pathway driven by IL-33 in immune cells that may be harnessed for therapeutic benefit or for robust expansion of Treg in vitro and in vivo.

Cutting edge: Flt3 ligand mediates STAT3-independent expansion but STAT3-dependent activation of myeloid-derived suppressor cells.

The Flt3-Flt3 ligand (Flt3L) pathway is critically involved in the differentiation and homeostasis of myeloid cells, including dendritic cells (DC); however, its role in the expansion and function of myeloid-derived suppressor cells (MDSC) has not been determined. In this article, we describe the ability of Flt3L to expand and activate murine MDSC capable of suppressing allograft rejection upon adoptive transfer. Although Flt3L expands and augments the stimulatory capacity of myeloid DC, MDSC expanded by Flt3L have increased suppressive activity. Although STAT3 is considered the central transcription factor for MDSC expansion, inhibition and genetic ablation of STAT3 did not block, but rather augmented, Flt3L-mediated MDSC expansion. MDSC suppressive function, preserved when STAT3 inhibition was removed, was reduced by genetic STAT3 deletion. Both STAT3 inhibition and deletion reduced Flt3L-mediated DC expansion, signifying that STAT3 had reciprocal effects on suppressive MDSC and immunostimulatory DC expansion. Together, these findings enhance our understanding of the immunomodulatory properties of Flt3L.

Controlling the burn and fueling the fire: defining the role for the alarmin interleukin-33 in alloimmunity.

PURPOSE OF REVIEW: The purpose of this review is to provide a general update on recent developments in the immunobiology of IL-33 and IL-33-targeted immune cells. We also discuss emerging concepts regarding the potential role IL-33 appears to play in altering alloimmune responses mediating host-versus-graft and graft-versus-host alloresponses. RECENT FINDINGS: Stromal cells and leukocytes display regulated expression of IL-33 and may actively or passively secrete this pleotropic cytokine. Type 2 innate lymphoid cells and a large proportion of tissue resident regulatory T cells (Treg) express membrane-bound suppressor of tumorigenicity 2 (ST2), the IL-33 receptor. Although Treg are appreciated suppressors of the inflammatory function of immune cells, both type 2 innate lymphoid cells and tissue resident Treg could play key roles in tissue repair and homeostasis. The functions of IL-33 in transplantation are poorly understood. However, like other disease models, the functions of IL-33 in alloimmunity appear to be quite pleiotropic. IL-33 is associated with immune regulation and graft protection in cardiac transplant settings. Yet, it is highly proinflammatory and stimulates lethal graft-versus-host disease through its capacity to stimulate type 1 immunity. SUMMARY: Intensive studies on IL-33/ST2 signaling pathways and ST2 cell populations in solid organ and cell transplantation are warranted. A better understanding of this important pathway will provide promising therapeutic targets controlling pathogenic alloimmune responses, as well as potentially facilitating the function of regulatory and reparative immune cells posttransplantation.

Murine dendritic cell rapamycin-resistant and rictor-independent mTOR controls IL-10, B7-H1, and regulatory T-cell induction.

Mammalian target of rapamycin (mTOR) is an important, yet poorly understood integrative kinase that regulates immune cell function. mTOR functions in 2 independent complexes: mTOR complex (mTORC) 1 and 2. The immunosuppressant rapamycin (RAPA) inhibits mTORC1 but not mTORC2 and causes a paradoxical reduction in anti-inflammatory interleukin (IL) 10 and B7-homolog 1 (B7-H1) expression by dendritic cells (DCs). Using catalytic mTOR inhibitors and DCs lacking mTORC2, we show that restraint of signal transducer and activator of transcription 3-mediated IL-10 and B7-H1 expression during DC maturation involves a RAPA-insensitive and mTORC2-independent mTOR mechanism. Relatedly, catalytic mTOR inhibition promotes B7-H1-dependent and IL-1ß-dependent DC induction of regulatory T cells (Tregs). Thus, we define an immunoregulatory pathway in which RAPA-sensitive mTORC1 in DCs promotes effector T-cell expansion and RAPA-insensitive mTORC1 restrains T(reg) induction. These findings identify the first known RAPA-insensitive mTOR pathway that is not mediated solely by mTORC2 and have implications for the use of catalytic mTOR inhibitors in inflammatory disease settings.

Low TCR signal strength induces combined expansion of Th2 and regulatory T cell populations that protect mice from the development of type 1 diabetes.

AIMS/HYPOTHESIS: Weak stimulation of CD4(+) T cells induces expansion of CD4(+) forkhead box P3(+) regulatory T cells (Tregs) and can also promote T helper (Th) 2 responses, which have demonstrable beneficial effects on autoimmune diabetes. This study explored the feasibility of combined Treg/Th2 expansion for immunotherapy of type 1 diabetes in NOD mice. METHODS: We compared Treg and Th responses to dendritic cells (DC) presenting scaled antigen doses to islet-specific NOD CD4(+) T cells. Flow cytometric and Luminex analyses were performed to determine the phenotype and cytokine profile of expanded T cells. The ability of expanded T cells to prevent type 1 diabetes was tested in an adoptive transfer model. RESULTS: In vitro studies revealed a hierarchical, selective expansion of Treg and T effector (Teff) populations at different antigen doses. Thus, a single low dose produced a mixture of Tregs Th2 and type 1 regulatory (Tr1) cells, which prevented diabetes in NOD-SCID mice and increased the ratio of Treg/Teff cells infiltrating the pancreatic islets. Subcutaneous injection of DC, previously shown to prevent diabetes in NOD mice, induced expansion of the same mixture of Tregs Tr1 and Th2 cells. Low-dose expansion of Treg required MHC-T cell receptor interaction and was partly dependent on T cell derived TGF-ß and IL-2. Autocrine IFN-γ was required for the promotion of diabetogenic Th1 cells at high antigen doses. CONCLUSIONS/INTERPRETATION: Weak stimulation of CD4(+) T cells with DC and low-dose antigen expands a combination of antigen-specific Tregs Th2 and Tr1 cells that prevent autoimmunity, without the need to target or purify specific Treg populations.

A window into immunosuppressant immunoregulation: recipient conversion to rapamycin increases potentially tolerogenic immune cells.

Mechanistic target of rapamycin inhibitors (mTORi) have a complex immunoregulatory profile in both animal models and transplant patients. Studies suggest that mTORi act as tolerance-supporting and regulatory T cell (Treg)-promoting immunosuppressants. Yet proinflammatory influences on myeloid dendritic cells have been established. Insight is needed into the impact of mTORi on immune cells. Stallone et al. describe a clinical study identifying a potential immunoregulatory pathway involving plasmacytoid dendritic cells and Tregs in renal transplant patients on mTORi.

IL-12hi rapamycin-conditioned dendritic cells mediate IFN-γ-dependent apoptosis of alloreactive CD4+ T cells in vitro and reduce lethal graft-versus-host disease.

Rapamycin (RAPA) inhibits the mechanistic target of rapamycin (mTOR), a crucial immune system regulator. Dendritic cells (DC) generated in RAPA (RAPA-DC) enrich for CD4(+) forkhead box p3 (FoxP3(+)) regulatory T cells and induce T cell apoptosis by an unknown mechanism. RAPA-DC also promote experimental allograft survival, yet paradoxically secrete increased IL-12, crucial for the generation of IFN-γ(+) CD4(+) T cells. However, IFN-γ is pro-apoptotic and IL-12-driven IFN-γ inhibits experimental graft-versus-host disease (GVHD). We hypothesized that IL-12(hi) RAPA-DC would facilitate IFN-γ-mediated apoptosis of alloreactive T cells and, unlike control (CTR)-DC, would reduce lethal GVHD. Following LPS stimulation, RAPA-DC exhibited decreased MHCII and co-stimulatory molecules and contained a significant population of CD86(lo) IL-12(hi) cells. Consistent with our hypothesis, both unstimulated and LPS-stimulated RAPA-DC enhanced alloreactive CD4(+) T cell apoptosis in culture. Augmented T cell apoptosis was ablated by IFN-γ neutralization or using T cells lacking the IFN-γ receptor, and it was associated with increased expression of Fas and cleaved caspase 8. DC production or responses to IFN-γ were not important to increased apoptotic functions of RAPA-DC. LPS-stimulated IL-12p40(-/-) RAPA-DC induced lower levels of T cell apoptosis in culture, which was further decreased with addition of anti-IFN-γ. Finally, whereas CTR-DC accelerated mortality from GVHD, LPS-treated RAPA-DC significantly prolonged host survival. In conclusion, increased apoptosis of allogeneic CD4(+) T cells induced by LPS-stimulated IL-12(hi) RAPA-DC is mediated in vitro through IFN-γ and in part by increased IL-12 expression. Enhanced production of IL-12, the predominant inducer of IFN-γ by immune cells, is a probable mechanism underlying the capacity of LPS-treated RAPA-DC to reduce GVHD.

Dendritic cells and regulation of graft-versus-host disease and graft-versus-leukemia activity.

Hematopoietic stem cell transplantation is the only curative treatment for many malignant hematologic diseases, with an often critical graft-versus-leukemia effect. Despite peritransplant prophylaxis, GVHD remains a significant cause of posthematopoietic stem cell transplantation morbidity and mortality. Traditional therapies have targeted T cells, yet immunostimulatory dendritic cells (DCs) are critical in the pathogenesis of GVHD. Furthermore, DCs also have tolerogenic properties. Monitoring of DC characteristics may be predictive of outcome, and therapies that target DCs are innovative and promising. DCs may be targeted in vivo or tolerogenic (tol) DCs may be generated in vitro and given in the peritransplant period. Other cellular therapies, notably regulatory T cells (T(reg)) and mesenchymal stem cells, mediate important effects through DCs and show promise for the prevention and treatment of GVHD in early human studies. Therapies are likely to be more effective if they have synergistic effects or target both DCs and T cells in vivo, such as tolDCs or T(reg). Given the effectiveness of tolDCs in experimental models of GVHD and their safety in early human studies for type 1 diabetes, it is crucial that tolDCs be investigated in the prevention and treatment of human GVHD while ensuring conservation of graft-versus-leukemia effects.

Implications for Interleukin-33 in solid organ transplantation.

Interleukin(IL)-33 is a member of the IL-1 cytokine family that has been attributed T helper (Th) type 2 immunity-promoting capacity. However, new studies indicate that IL-33 is a multifunctional protein that acts as transcriptional/signaling repressor, functions as an alarmin alerting the immune system to necrosis, as well as serves as a cytokine that targets cells expressing ST2, the IL-33 receptor. Interestingly, IL-33 is also emerging as a pleiotropic cytokine. Depending on the innate or adaptive immune cells targeted by IL-33, it can not only promote type 2, but also IFN-γ dominated type 1 immunity. In addition, IL-33 expands regulatory T cells. In this review, we assimilate the current knowledge of IL-33 immunobiology and discuss how IL-33 may mediate such diverse roles in the immune response to pathogens and development of immune-mediated pathologies. The function of IL-33 in shaping alloimmune responses to transplanted organs is poorly explored, but a particularly beneficial role of IL-33 in experimental heart transplant models is summarized. Finally, given the implication of IL-33 in pathologies of the lung and intestine, we discuss how IL-33 may contribute to the comparatively poor outcomes following transplantation of these two organs.