Catriona T. Prendergast

Cutting Edge: Th1 Cells Facilitate the Entry of Th17 Cells to the Central Nervous System during Experimental Autoimmune Encephalomyelitis

It has recently been proposed that experimental autoimmune encephalomyelitis, once considered the classical Th1 disease, is predominantly Th17 driven. In this study we show that myelin-reactive Th1 preparations devoid of contaminating IL-17+ cells are highly pathogenic. In contrast, Th17 preparations lacking IFN-γ+ cells do not cause disease. Our key observation is that only Th1 cells can access the noninflamed CNS. Once Th1 cells establish the experimental autoimmune encephalomyelitis lesion, Th17 cells appear in the CNS. These data shed important new light on the ability of Th1 vs Th17 cells to access inflamed vs normal tissue. Because the IL-17-triggered release of chemokines by stromal cells could attract many other immune cells, allowing Th17 cells to access the tissues only under conditions of inflammation may be a key process limiting (auto)immune pathology. This has major implications for the design of therapeutic interventions, many of which are now aiming at Th17 rather than Th1 cells.

Immune cell entry to central nervous system--current understanding and prospective therapeutic targets.

Under normal physiological conditions there is minimal entry of immune cells into the central nervous system (CNS) for the purpose of immune surveillance. During inflammation, however, extensive infiltration of immune cells can lead to the induction of CNS autoimmune disease, for example multiple sclerosis (MS) and its animal model experimental autoimmune encephalomyelitis (EAE). The barriers that regulate cellular entry are the blood-brain barrier (BBB) within the CNS parenchyma, and the blood-cerebrospinal fluid (blood-CSF) barrier within the choroid plexus. Understanding how these barriers function to allow the passage of leukocytes from the periphery into the CNS for normal immune surveillance, and under inflammatory conditions, is vital for the development of novel therapeutics targeting immune cell migration in CNS diseases. Contributions from selectins, chemokines, integrins and matrix metalloproteinases allow the migration of leukocytes across the BBB and into the CNS parenchyma. In EAE and MS, the strict maintenance of this process is lost and a large influx of cells is seen. This review focuses on the role of these homing molecules, chemokines and enzymes in the entry of leukocytes into the CNS during inflammatory conditions. It concludes with a model of immune cell entry of the CNS, summarising the current knowledge in this area. Targeting specific molecules to prevent infiltration of inflammatory cells into the CNS could allow disease inhibition without compromising beneficial immune surveillance.

IL-10 Production in Macrophages Is Regulated by a TLR-Driven CREB-Mediated Mechanism That Is Linked to Genes Involved in Cell Metabolism

IL-10 is produced by macrophages in diverse immune settings and is critical in limiting immune-mediated pathology. In helminth infections, macrophages are an important source of IL-10; however, the molecular mechanism underpinning production of IL-10 by these cells is poorly characterized. In this study, bone marrow–derived macrophages exposed to excretory/secretory products released by Schistosoma mansoni cercariae rapidly produce IL-10 as a result of MyD88-mediated activation of MEK/ERK/RSK and p38. The phosphorylation of these kinases was triggered by TLR2 and TLR4 and converged on activation of the transcription factor CREB. Following phosphorylation, CREB is recruited to a novel regulatory element in the Il10 promoter and is also responsible for regulating a network of genes involved in metabolic processes, such as glycolysis, the tricarboxylic acid cycle, and oxidative phosphorylation. Moreover, skin-resident tissue macrophages, which encounter S. mansoni excretory/secretory products during infection, are the first monocytes to produce IL-10 in vivo early postinfection with S. mansoni cercariae. The early and rapid release of IL-10 by these cells has the potential to condition the dermal microenvironment encountered by immune cells recruited to this infection site, and we propose a mechanism by which CREB regulates the production of IL-10 by macrophages in the skin, but also has a major effect on their metabolic state.

Epigenetic modification of the PD-1 (Pdcd1) promoter in effector CD4+ T cells tolerized by peptide immunotherapy

Clinically effective antigen-based immunotherapy must silence antigen-experienced effector T cells (Teff) driving ongoing immune pathology. Using CD4+ autoimmune Teff cells, we demonstrate that peptide immunotherapy (PIT) is strictly dependent upon sustained T cell expression of the co-inhibitory molecule PD-1. We found high levels of 5-hydroxymethylcytosine (5hmC) at the PD-1 (Pdcd1) promoter of non-tolerant T cells. 5hmC was lost in response to PIT, with DNA hypomethylation of the promoter. We identified dynamic changes in expression of the genes encoding the Ten-Eleven-Translocation (TET) proteins that are associated with the oxidative conversion 5-methylcytosine and 5hmC, during cytosine demethylation. We describe a model whereby promoter demethylation requires the co-incident expression of permissive histone modifications at the Pdcd1 promoter together with TET availability. This combination was only seen in tolerant Teff cells following PIT, but not in Teff that transiently express PD-1. Epigenetic changes at the Pdcd1 locus therefore determine the tolerizing potential of TCR-ligation. DOI: http://dx.doi.org/10.7554/eLife.03416.001

Helminth Infection and Commensal Microbiota Drive Early IL-10 Production in the Skin by CD4 + T Cells That Are Functionally Suppressive

The skin provides an important first line of defence and immunological barrier to invasive pathogens, but immune responses must also be regulated to maintain barrier function and ensure tolerance of skin surface commensal organisms. In schistosomiasis-endemic regions, populations can experience repeated percutaneous exposure to schistosome larvae, however little is known about how repeated exposure to pathogens affects immune regulation in the skin. Here, using a murine model of repeated infection with Schistosoma mansoni larvae, we show that the skin infection site becomes rich in regulatory IL-10, whilst in its absence, inflammation, neutrophil recruitment, and local lymphocyte proliferation is increased. Whilst CD4+ T cells are the primary cellular source of regulatory IL-10, they expressed none of the markers conventionally associated with T regulatory (Treg) cells (i.e. FoxP3, Helios, Nrp1, CD223, or CD49b). Nevertheless, these IL-10+ CD4+ T cells in the skin from repeatedly infected mice are functionally suppressive as they reduced proliferation of responsive CD4+ T cells from the skin draining lymph node. Moreover, the skin of infected Rag-/- mice had impaired IL-10 production and increased neutrophil recruitment. Finally, we show that the mechanism behind IL-10 production by CD4+ T cells in the skin is due to a combination of an initial (day 1) response specific to skin commensal bacteria, and then over the following days schistosome-specific CD4+ T cell responses, which together contribute towards limiting inflammation and tissue damage following schistosome infection. We propose CD4+ T cells in the skin that do not express markers of conventional T regulatory cell populations have a significant role in immune regulation after repeated pathogen exposure and speculate that these cells may also help to maintain skin barrier function in the context of repeated percutaneous insult by other skin pathogens.

Epidermal keratinocytes initiate wound healing and pro-inflammatory immune responses following percutaneous schistosome infection.

Keratinocytes constitute the majority of cells in the skins epidermis, the first line of defence against percutaneous pathogens. Schistosome larvae (cercariae) actively penetrate the epidermis to establish infection, however the response of keratinocytes to invading cercariae has not been investigated. Here we address the hypothesis that cercariae activate epidermal keratinocytes to promote the development of a pro-inflammatory immune response in the skin. C57BL/6 mice were exposed to Schistosoma mansoni cercariae via each pinna and non-haematopoietic cells isolated from epidermal tissue were characterised for the presence of different keratinocyte sub-sets at 6, 24 and 96 h p.i. We identified an expansion of epidermal keratinocyte precursors (CD45(-), CD326(-), CD34(+)) within 24 h of infection relative to naïve animals. Following infection, cells within the precursor population displayed a more differentiated phenotype (α6integrin(-)) than in uninfected skin. Parallel immunohistochemical analysis of pinnae cryosections showed that this expansion corresponded to an increase in the intensity of CD34 staining, specifically in the basal bulge region of hair follicles of infected mice, and a higher frequency of keratinocyte Ki67(+) nuclei in both the hair follicle and interfollicular epidermis. Expression of pro-inflammatory cytokine and stress-associated keratin 6b genes was also transiently upregulated in the epidermal tissue of infected mice. In vitro exposure of keratinocyte precursors isolated from neonatal mouse skin to excretory/secretory antigens released by penetrating cercariae elicited IL-1α and IL-1β production, supporting a role for keratinocyte precursors in initiating cutaneous inflammatory immune responses. Together, these observations indicate that S.mansoni cercariae and their excretory/secretory products act directly upon epidermal keratinocytes, which respond by initiating barrier repair and pro-inflammatory mechanisms similar to those observed in epidermal wound healing.

CD4+ T Cell Hyporesponsiveness after Repeated Exposure to Schistosoma mansoni Larvae Is Dependent upon Interleukin-10

ABSTRACT The effect that multiple percutaneous exposures to Schistosoma larvae has on the development of early CD4+ lymphocyte reactivity is unclear, yet it is important in the context of humans living in areas where schistosomiasis is endemic. In a murine model of multiple infections, we show that exposure of mice to repeated doses (4×) of Schistosoma mansoni cercariae, compared to a single dose (1×), results in CD4+ T cell hyporesponsiveness within the skin-draining lymph nodes (sdLN), manifested as reduced CD4+ cell proliferation and cytokine production. FoxP3+ CD4+ regulatory T cells were present in similar numbers in the sdLN of 4× and 1× mice and thus are unlikely to have a role in effecting hyporesponsiveness. Moreover, anergy of the CD4+ cell population from 4× mice was slight, as proliferation was only partly circumvented through the in vitro addition of exogenous interleukin-2 (IL-2), and the in vivo blockade of the regulatory molecule PD1 had a minimal effect on restoring responsiveness. In contrast, IL-10 was observed to be critical in mediating hyporesponsiveness, as CD4+ cells from the sdLN of 4× mice deficient for IL-10 were readily able to proliferate, unlike those from 4× wild-type cohorts. CD4+ cells from the sdLN of 4× mice exhibited higher levels of apoptosis and cell death, but in the absence of IL-10, there was significantly less cell death. Combined, our data show that IL-10 is a key factor in the development of CD4+ T cell hyporesponsiveness after repeated parasite exposure involving CD4+ cell apoptosis.

CD4 T-cell hyporesponsiveness induced by schistosome larvae is not dependent upon eosinophils but may involve connective tissue mast cells

In areas endemic for schistosomiasis, people can often be in contact with contaminated water resulting in repeated exposures to infective Schistosoma mansoni cercariae. Using a murine model, repeated infections result in IL‐10‐dependent CD4+ T‐cell hyporesponsiveness in the skin‐draining lymph nodes (sdLN), which could be caused by an abundance of eosinophils and connective tissue mast cells at the skin infection site. Here, we show that whilst the absence of eosinophils did not have a significant effect on cytokine production, MHC‐II+ cells were more numerous in the dermal cell exudate population. Nevertheless, the absence of dermal eosinophils did not lead to an increase in the responsiveness of CD4+ T cells in the sdLN, revealing that eosinophils in repeatedly exposed skin did not impact on the development of CD4+ T‐cell hyporesponsiveness. On the other hand, the absence of connective tissue mast cells led to a reduction in dermal IL‐10 and to an increase in the number of MHC‐II+ cells infiltrating the skin. There was also a small but significant alleviation of hyporesponsiveness in the sdLN, suggesting that mast cells may have a role in regulating immune responses after repeated exposures of the skin to S. mansoni cercariae.

The thymus and rheumatology: should we care?

Purpose of reviewThe purpose of this review is to discuss the mechanisms of central and peripheral tolerance in relation to T-cell mediated autoimmunity in rheumatoid arthritis (RA). Recent findingsThe well established association between major histocompatibility complex class II and RA has led us to understand that T cells, and the adaptive immune response, are important in the pathogenesis of disease. In order for autoimmune disease to develop, there is a breach of tolerance to self antigen and the mechanisms of both central and peripheral tolerance aim to prevent this. Here, we review evidence from mouse models indicating that alterations in T-cell receptor signalling thresholds during thymic selection may be linked to the escape of T cells that mediate autoimmune arthritis. In addition, we summarize the role of dendritic cells and Foxp3+ regulatory T cells in both peripheral and thymic tolerance, and highlight their relevance to what we know about the aetiology of RA. SummaryMechanisms of central tolerance in the thymus and peripheral tolerance are in place to control autoreactive T cells and to prevent the development of autoimmune disease. We anticipate that a better understanding of these mechanisms will lead to the development of better, antigen-specific therapeutics to restore tolerance.

Visualising the interaction of CD4 T cells and DCs in the evolution of inflammatory arthritis

Objectives Successful early intervention in rheumatoid arthritis (RA) with the aim of resetting immunological tolerance requires a clearer understanding of how specificity, cellular kinetics and spatial behaviour shape the evolution of articular T cell responses. We aimed to define initial seeding of articular CD4+ T cell responses in early experimental arthritis, evaluating their dynamic behaviour and interactions with dendritic cells (DCs) in the inflamed articular environment. Methods Antigen-induced arthritis was used to model articular inflammation. Flow cytometry and PCR of T cell receptor (TCR) diversity genes allowed phenotypic analysis of infiltrating T cells. The dynamic interactions of T cells with joint residing DCs were visualised using intravital multiphoton microscopy. Results Initial recruitment of antigen-specific T cells into the joint was paralleled by accumulation of CD4+ T cells with diverse antigen-receptor expression and ability to produce tumour necrosis factor alpha (TNFα) and interferon gamma (IFNγ) on mitogenic restimulation. A proportion of this infiltrate demonstrated slower motility speeds and engaged for longer periods with articular DCs in vivo. Abatacept treatment did not disrupt these interactions but did reduce T cell expression of inducible costimulatory (ICOS) molecule. We also demonstrated that non-specific CD4+ T cells could be recruited during these early articular events. Conclusions We demonstrate that CD4+ T cells engage with articular DCs supporting antigen specific T cell reactivation. This cellular dialogue can be targeted therapeutically to reduce local T cell activation.