Alexander T. Phythian-Adams

CD11c depletion severely disrupts Th2 induction and development in vivo.

Although dendritic cells (DCs) are adept initiators of CD4+ T cell responses, their fundamental importance in this regard in Th2 settings remains to be demonstrated. We have used CD11c–diphtheria toxin (DTx) receptor mice to deplete CD11c+ cells during the priming stage of the CD4+ Th2 response against the parasitic helminth Schistosoma mansoni. DTx treatment significantly depleted CD11c+ DCs from all tissues tested, with 70–80% efficacy. Even this incomplete depletion resulted in dramatically impaired CD4+ T cell production of Th2 cytokines, altering the balance of the immune response and causing a shift toward IFN-γ production. In contrast, basophil depletion using Mar-1 antibody had no measurable effect on Th2 induction in this system. These data underline the vital role that CD11c+ antigen-presenting cells can play in orchestrating Th2 development against helminth infection in vivo, a response that is ordinarily balanced so as to prevent the potentially damaging production of inflammatory cytokines.

A dominant role for the methyl-CpG-binding protein Mbd2 in controlling Th2 induction by dendritic cells

Dendritic cells (DCs) direct CD4+ T-cell differentiation into diverse helper (Th) subsets that are required for protection against varied infections. However, the mechanisms used by DCs to promote Th2 responses, which are important both for immunity to helminth infection and in allergic disease, are currently poorly understood. We demonstrate a key role for the protein methyl-CpG-binding domain-2 (Mbd2), which links DNA methylation to repressive chromatin structure, in regulating expression of a range of genes that are associated with optimal DC activation and function. In the absence of Mbd2, DCs display reduced phenotypic activation and a markedly impaired capacity to initiate Th2 immunity against helminths or allergens. These data identify an epigenetic mechanism that is central to the activation of CD4+ T-cell responses by DCs, particularly in Th2 settings, and reveal methyl-CpG-binding proteins and the genes under their control as possible therapeutic targets for type-2 inflammation.

The adult murine heart has a sparse, phagocytically active macrophage population that expands through monocyte recruitment and adopts an 'M2' phenotype in response to Th2 immunologic challenge.

Tissue resident macrophages have vital homeostatic roles in many tissues but their roles are less well defined in the heart. The present study aimed to identify the density, polarisation status and distribution of macrophages in the healthy murine heart and to investigate their ability to respond to immune challenge. Histological analysis of hearts from CSF-1 receptor (csf1-GFP; MacGreen) and CX3CR1 (Cx3cr1GFP/+) reporter mice revealed a sparse population of GFP positive macrophages that were evenly distributed throughout the left and right ventricular free walls and septum. F4/80+CD11b+ cardiac macrophages, sorted from myocardial homogenates, were able to phagocytose fluorescent beads in vitro and expressed markers typical of both ‘M1’ (IL-1β, TNF and CCR2) and ‘M2’ activation (Ym1, Arg 1, RELMα and IL-10), suggesting no specific polarisation in healthy myocardium. Exposure to Th2 challenge by infection of mice with helminth parasites Schistosoma mansoni, or Heligmosomoides polygyrus, resulted in an increase in cardiac macrophage density, adoption of a stellate morphology and increased expression of Ym1, RELMα and CD206 (mannose receptor), indicative of ‘M2’ polarisation. This was dependent on recruitment of Ly6ChighCCR2+ monocytes and was accompanied by an increase in collagen content. In conclusion, in the healthy heart resident macrophages are relatively sparse and have a phagocytic role. Following Th2 challenge this population expands due to monocyte recruitment and adopts an ‘M2’ phenotype associated with increased tissue fibrosis.

Th2 Responses to Helminth Parasites Can Be Therapeutically Enhanced by, but Are Not Dependent upon, GITR–GITR Ligand Costimulation In Vivo

The immune suppression that characterizes human helminth infections can hinder the development of protective immunity or help to reduce pathogenic inflammation. Signaling through the T cell costimulator glucocorticoid-induced TNFR-related protein (GITR) counteracts immune downregulation by augmenting effector T cell responses and abrogating suppression by Foxp3+ regulatory T cells. Thus, superphysiological Ab-mediated GITR costimulation represents a novel therapy for promoting protective immunity toward parasitic helminths, whereas blocking physiological GITR–GITR ligand (GITRL) interactions may provide a mechanism for dampening pathogenic Th2 inflammation. We investigated the superphysiological and physiological roles of the GITR–GITRL pathway in the development of protective and pathogenic Th2 responses in murine infection models of filariasis (Litomosoides sigmodontis) and schistosomiasis (Schistosoma mansoni). Providing superphysiological GITR costimulation using an agonistic anti-GITR mAb over the first 12 d of L. sigmodontis infection initially increased the quantity of Th2 cells, as well as their ability to produce Th2 cytokines. However, as infection progressed, the Th2 responses reverted to normal infection levels, and parasite killing remained unaffected. Despite the Th2-promoting role of superphysiological GITR costimulation, Ab-mediated blockade of the GITR–GITRL pathway did not affect Th2 cell priming or maintenance during L. sigmodontis infection. Blockade of GITR–GITRL interactions during the acute egg phase of S. mansoni infection resulted in reduced Th2 responses, but this effect was confined to the spleen and did not lead to changes in liver pathology. Thus, although superphysiological GITR costimulation can therapeutically enhance Th2 responses, physiological GITR–GITRL interactions are not required for the development of Th2-mediated resistance or pathology in murine models of filariasis and schistosomiasis.

A unique DNA methylation signature defines a population of IFN-γ/IL-4 double-positive T cells during helminth infection

Th1 and Th2 cell fates are traditionally viewed as mutually exclusive, but recent work suggests that these lineages may be more plastic than previously thought. When isolating splenic CD4+ T cells from mice infected with the parasitic helminth Schistosoma mansoni, we observed a defined population of IFN‐γ/IL‐4 double‐positive cells. These IFN‐γ+IL‐4+ cells showed differences in DNA methylation at the Ifng and Il4 loci when compared with IFN‐γ+IL‐4− (Th1) and IFN‐γ−IL‐4+ (Th2) cells, demonstrating that they represent a distinct effector cell population. IFN‐γ+IL‐4+ cells also displayed a discrete DNA methylation pattern at a CpG island within the body of the Gata3 gene, which encodes the master regulator of Th2 identity. DNA methylation at this region correlated with decreased Gata3 levels, suggesting a possible role in controlling Gata3 expression. These data provide important insight into the molecular mechanisms behind the co‐existence of Th1 and Th2 characteristics.

Optimal effector functions in human natural killer cells rely upon autocrine bone morphogenetic protein signaling

Natural killer (NK) cells are critical for innate tumor immunity due to their specialized ability to recognize and kill neoplastically transformed cells. However, NK cells require a specific set of cytokine-mediated signals to achieve optimal effector function. Th1-associated cytokines promote effector functions that are inhibited by the prototypic Th2 cytokine IL4 and the TGFβ superfamily members TGFβ1 and activin-A. Interestingly, the largest subgroup of the TGFβ superfamily are the bone morphogenetic proteins (BMP), but the effects of BMP signaling on NK cell effector functions have not been evaluated. Here, we demonstrate that blood-circulating NK cells express type I and II BMP receptors, BMP-2 and BMP-6 ligands, and phosphorylated isoforms of Smad-1/-5/-8, which mediate BMP family member signaling. In opposition to the inhibitory effects of TGFβ1 or activin-A, autocrine BMP signaling was supportive to NK cell function. Mechanistic investigations in cytokine and TLR-L-activated NK cells revealed that BMP signaling optimized IFNγ and global cytokine and chemokine production, phenotypic activation and proliferation, and autologous dendritic cell activation and target cytotoxicity. Collectively, our findings identify a novel auto-activatory pathway that is essential for optimal NK cell effector function, one that might be therapeutically manipulated to help eradicate tumors. Cancer Res; 74(18); 5019-31. ©2014 AACR.

A central role for hepatic conventional dendritic cells in supporting Th2 responses during helminth infection

Dendritic cells (DCs) are the key initiators of T‐helper (Th) 2 immune responses against the parasitic helminth Schistosoma mansoni. Although the liver is one of the main sites of antigen deposition during infection with this parasite, it is not yet clear how distinct DC subtypes in this tissue respond to S. mansoni antigens in vivo, or how the liver microenvironment might influence DC function during establishment of the Th2 response. In this study, we show that hepatic DC subsets undergo distinct activation processes in vivo following murine infection with S. mansoni. Conventional DCs (cDCs) from schistosome‐infected mice upregulated expression of the costimulatory molecule CD40 and were capable of priming naive CD4+ T cells, whereas plasmacytoid DCs (pDCs) upregulated expression of MHC class II, CD86 and CD40 but were unable to support the expansion of either naive or effector/memory CD4+ T cells. Importantly, in vivo depletion of pDCs revealed that this subset was dispensable for either maintenance or regulation of the hepatic Th2 effector response during acute S. mansoni infection. Our data provides strong evidence that S. mansoni infection favors the establishment of an immunogenic, rather than tolerogenic, liver microenvironment that conditions cDCs to initiate and maintain Th2 immunity in the context of ongoing antigen exposure.

Modulation of dendritic cell alternative activation and function by the vitamin A metabolite retinoic acid.

Retinoic acid modulates the functions of IL-4 in alternatively activated DCs

Type I interferon is required for T helper (Th) 2 induction by dendritic cells

Type 2 inflammation is a defining feature of infection with parasitic worms (helminths), as well as being responsible for widespread suffering in allergies. However, the precise mechanisms involved in T helper (Th) 2 polarization by dendritic cells (DCs) are currently unclear. We have identified a previously unrecognized role for type I IFN (IFN‐I) in enabling this process. An IFN‐I signature was evident in DCs responding to the helminth Schistosoma mansoni or the allergen house dust mite (HDM). Further, IFN‐I signaling was required for optimal DC phenotypic activation in response to helminth antigen (Ag), and efficient migration to, and localization with, T cells in the draining lymph node (dLN). Importantly, DCs generated from Ifnar1−/− mice were incapable of initiating Th2 responses in vivo. These data demonstrate for the first time that the influence of IFN‐I is not limited to antiviral or bacterial settings but also has a central role to play in DC initiation of Th2 responses.

SOX9 predicts progression toward cirrhosis in patients while its loss protects against liver fibrosis

Fibrosis and organ failure is a common endpoint for many chronic liver diseases. Much is known about the upstream inflammatory mechanisms provoking fibrosis and downstream potential for tissue remodeling. However, less is known about the transcriptional regulation in vivo governing fibrotic matrix deposition by liver myofibroblasts. This gap in understanding has hampered molecular predictions of disease severity and clinical progression and restricted targets for antifibrotic drug development. In this study, we show the prevalence of SOX9 in biopsies from patients with chronic liver disease correlated with fibrosis severity and accurately predicted disease progression toward cirrhosis. Inactivation of Sox9 in mice protected against both parenchymal and biliary fibrosis, and improved liver function and ameliorated chronic inflammation. SOX9 was downstream of mechanosignaling factor, YAP1. These data demonstrate a role for SOX9 in liver fibrosis and open the way for the transcription factor and its dependent pathways as new diagnostic, prognostic, and therapeutic targets in patients with liver fibrosis.