mRNA vaccine against malaria tailored for liver-resident memory T cells.

Malaria is caused by Plasmodium species transmitted by Anopheles mosquitoes. Following a mosquito bite, Plasmodium sporozoites migrate from skin to liver, where extensive replication occurs, emerging later as merozoites that can infect red blood cells and cause symptoms of disease. As liver tissue-resident memory T cells (Trm cells) have recently been shown to control liver-stage infections, we embarked on a messenger RNA (mRNA)-based vaccine strategy to induce liver Trm cells to prevent malaria. Although a standard mRNA vaccine was unable to generate liver Trm or protect against challenge with Plasmodium berghei sporozoites in mice, addition of an agonist that recruits T cell help from type I natural killer T cells under mRNA-vaccination conditions resulted in significant generation of liver Trm cells and effective protection. Moreover, whereas previous exposure of mice to blood-stage infection impaired traditional vaccines based on attenuated sporozoites, mRNA vaccination was unaffected, underlining the potential for such a rational mRNA-based strategy in malaria-endemic regions.

Memory T cell subsets, migration patterns, and tissue residence.

Tissues such as the skin and mucosae are frequently exposed to microbial pathogens. Infectious agents must be quickly and efficiently controlled by our immune system, but the low frequency of naive T cells specific for any one pathogen means dependence on primary responses initiated in draining lymph nodes, often allowing time for serious infection to develop. These responses imprint effectors with the capacity to home to infected tissues; this process, combined with inflammatory signals, ensures the effective targeting of primary immunity. Upon vaccination or previous pathogen exposure, increased pathogen-specific T cell numbers together with altered migratory patterns of memory T cells can greatly improve immune efficacy, ensuring infections are prevented or at least remain subclinical. Until recently, memory T cell populations were considered to comprise central memory T cells (TCM), which are restricted to the secondary lymphoid tissues and blood, and effector memory T cells (TEM), which broadly migrate between peripheral tissues, the blood, and the spleen. Here we review evidence for these two memory populations, highlight a relatively new player, the tissue-resident memory T cell (TRM), and emphasize the potential differences between the migratory patterns of CD4(+) and CD8(+) T cells. This new understanding raises important considerations for vaccine design and for the measurement of immune parameters critical to the control of infectious disease, autoimmunity, and cancer.

Discrete tissue microenvironments instruct diversity in resident memory T cell function and plasticity.

Tissue-resident memory T (TRM) cells are non-recirculating cells that exist throughout the body. Although TRM cells in various organs rely on common transcriptional networks to establish tissue residency, location-specific factors adapt these cells to their tissue of lodgment. Here we analyze TRM cell heterogeneity between organs and find that the different environments in which these cells differentiate dictate TRM cell function, durability and malleability. We find that unequal responsiveness to TGFß is a major driver of this diversity. Notably, dampened TGFß signaling results in CD103- TRM cells with increased proliferative potential, enhanced function and reduced longevity compared with their TGFß-responsive CD103+ TRM counterparts. Furthermore, whereas CD103- TRM cells readily modified their phenotype upon relocation, CD103+ TRM cells were comparatively resistant to transdifferentiation. Thus, despite common requirements for TRM cell development, tissue adaptation of these cells confers discrete functional properties such that TRM cells exist along a spectrum of differentiation potential that is governed by their local tissue microenvironment.

The NK cell granule protein NKG7 regulates cytotoxic granule exocytosis and inflammation.

Immune-modulating therapies have revolutionized the treatment of chronic diseases, particularly cancer. However, their success is restricted and there is a need to identify new therapeutic targets. Here, we show that natural killer cell granule protein 7 (NKG7) is a regulator of lymphocyte granule exocytosis and downstream inflammation in a broad range of diseases. NKG7 expressed by CD4+ and CD8+ T cells played key roles in promoting inflammation during visceral leishmaniasis and malaria-two important parasitic diseases. Additionally, NKG7 expressed by natural killer cells was critical for controlling cancer initiation, growth and metastasis. NKG7 function in natural killer and CD8+ T cells was linked with their ability to regulate the translocation of CD107a to the cell surface and kill cellular targets, while NKG7 also had a major impact on CD4+ T cell activation following infection. Thus, we report a novel therapeutic target expressed on a range of immune cells with functions in different immune responses.

Interplay between liver and blood stages of Plasmodium infection dictates malaria severity via γδ T cells and IL-17-promoted stress erythropoiesis.

Plasmodium replicates within the liver prior to reaching the bloodstream and infecting red blood cells. Because clinical manifestations of malaria only arise during the blood stage of infection, a perception exists that liver infection does not impact disease pathology. By developing a murine model where the liver and blood stages of infection are uncoupled, we showed that the integration of signals from both stages dictated mortality outcomes. This dichotomy relied on liver stage-dependent activation of Vγ4+ γδ T cells. Subsequent blood stage parasite loads dictated their cytokine profiles, where low parasite loads preferentially expanded IL-17-producing γδ T cells. IL-17 drove extra-medullary erythropoiesis and concomitant reticulocytosis, which protected mice from lethal experimental cerebral malaria (ECM). Adoptive transfer of erythroid precursors could rescue mice from ECM. Modeling of γδ T cell dynamics suggests that this protective mechanism may be key for the establishment of naturally acquired malaria immunity among frequently exposed individuals.

Local proliferation maintains a stable pool of tissue-resident memory T cells after antiviral recall responses.

Although tissue-resident memory T cells (TRM cells) are critical in fighting infection, their fate after local pathogen re-encounter is unknown. Here we found that skin TRM cells engaged virus-infected cells, proliferated in situ in response to local antigen encounter and did not migrate out of the epidermis, where they exclusively reside. As a consequence, secondary TRM cells formed from pre-existing TRM cells, as well as from precursors recruited from the circulation. Newly recruited antigen-specific or bystander TRM cells were generated in the skin without displacement of the pre-existing TRM cell pool. Thus, pre-existing skin TRM cell populations are not displaced after subsequent infections, which enables multiple TRM cell specificities to be stably maintained within the tissue.

Adrenergic regulation of the vasculature impairs leukocyte interstitial migration and suppresses immune responses.

The sympathetic nervous system (SNS) controls various physiological functions via the neurotransmitter noradrenaline. Activation of the SNS in response to psychological or physical stress is frequently associated with weakened immunity. Here, we investigated how adrenoceptor signaling influences leukocyte behavior. Intravital two-photon imaging after injection of noradrenaline revealed transient inhibition of CD8+ and CD4+ T cell locomotion in tissues. Expression of ß-adrenergic receptor in hematopoietic cells was not required for NA-mediated inhibition of motility. Rather, chemogenetic activation of the SNS or treatment with adrenergic receptor agonists induced vasoconstriction and decreased local blood flow, resulting in abrupt hypoxia that triggered rapid calcium signaling in leukocytes and halted cell motility. Oxygen supplementation reversed these effects. Treatment with adrenergic receptor agonists impaired T cell responses induced in response to viral and parasitic infections, as well as anti-tumor responses. Thus, stimulation of the SNS impairs leukocyte mobility, providing a mechanistic understanding of the link between adrenergic receptors and compromised immunity.

A Liver Capsular Network of Monocyte-Derived Macrophages Restricts Hepatic Dissemination of Intraperitoneal Bacteria by Neutrophil Recruitment.

The liver is positioned at the interface between two routes traversed by pathogens in disseminating infection. Whereas blood-borne pathogens are efficiently cleared in hepatic sinusoids by Kupffer cells (KCs), it is unknown how the liver prevents dissemination of peritoneal pathogens accessing its outer membrane. We report here that the hepatic capsule harbors a contiguous cellular network of liver-resident macrophages phenotypically distinct from KCs. These liver capsular macrophages (LCMs) were replenished in the steady state from blood monocytes, unlike KCs that are embryonically derived and self-renewing. LCM numbers increased after weaning in a microbiota-dependent process. LCMs sensed peritoneal bacteria and promoted neutrophil recruitment to the capsule, and their specific ablation resulted in decreased neutrophil recruitment and increased intrahepatic bacterial burden. Thus, the liver contains two separate and non-overlapping niches occupied by distinct resident macrophage populations mediating immunosurveillance at these two pathogen entry points to the liver.

The skin-resident and migratory immune system in steady state and memory: innate lymphocytes, dendritic cells and T cells.

The skin is a highly complex organ interspersed with a variety of smaller organ-like structures and a plethora of cell types that together perform essential functions such as physical sensing, temperature control, barrier maintenance and immunity. In this Review, we outline many of the innate and adaptive immune cell types associated with the skin, focusing on the steady state in mice and men, and include a broad update of dendritic cell function and T cell surveillance.

The developmental pathway for CD103(+)CD8+ tissue-resident memory T cells of skin.

Tissue-resident memory T cells (T(RM) cells) provide superior protection against infection in extralymphoid tissues. Here we found that CD103(+)CD8(+) T(RM) cells developed in the skin from epithelium-infiltrating precursor cells that lacked expression of the effector-cell marker KLRG1. A combination of entry into the epithelium plus local signaling by interleukin 15 (IL-15) and transforming growth factor-ß (TGF-ß) was required for the formation of these long-lived memory cells. Notably, differentiation into T(RM) cells resulted in the progressive acquisition of a unique transcriptional profile that differed from that of circulating memory cells and other types of T cells that permanently reside in skin epithelium. We provide a comprehensive molecular framework for the local differentiation of a distinct peripheral population of memory cells that forms a first-line immunological defense system in barrier tissues.

Liver-Resident Memory CD8+ T Cells Form a Front-Line Defense against Malaria Liver-Stage Infection.

In recent years, various intervention strategies have reduced malaria morbidity and mortality, but further improvements probably depend upon development of a broadly protective vaccine. To better understand immune requirement for protection, we examined liver-stage immunity after vaccination with irradiated sporozoites, an effective though logistically difficult vaccine. We identified a population of memory CD8+ T cells that expressed the gene signature of tissue-resident memory T (Trm) cells and remained permanently within the liver, where they patrolled the sinusoids. Exploring the requirements for liver Trm cell induction, we showed that by combining dendritic cell-targeted priming with liver inflammation and antigen recognition on hepatocytes, high frequencies of Trm cells could be induced and these cells were essential for protection against malaria sporozoite challenge. Our study highlights the immune potential of liver Trm cells and provides approaches for their selective transfer, expansion, or depletion, which may be harnessed to control liver infections or autoimmunity.

Hair follicles: gatekeepers to the epidermis.

Substantial depletion of Langerhans cells leads to their replenishment by bone marrow­derived precursors that access the epidermis through hair follicles, a site of crucial chemokine production.

NLRC4 inflammasomes in dendritic cells regulate noncognate effector function by memory CD8⁺ T cells.

Memory T cells exert antigen-independent effector functions, but how these responses are regulated is unclear. We discovered an in vivo link between flagellin-induced NLRC4 inflammasome activation in splenic dendritic cells (DCs) and host protective interferon-γ (IFN-γ) secretion by noncognate memory CD8(+) T cells, which could be activated by Salmonella enterica serovar Typhimurium, Yersinia pseudotuberculosis and Pseudomonas aeruginosa. We show that CD8α(+) DCs were particularly efficient at sensing bacterial flagellin through NLRC4 inflammasomes. Although this activation released interleukin 18 (IL-18) and IL-1ß, only IL-18 was required for IFN-γ production by memory CD8(+) T cells. Conversely, only the release of IL-1ß, but not IL-18, depended on priming signals mediated by Toll-like receptors. These findings provide a comprehensive mechanistic framework for the regulation of noncognate memory T cell responses during bacterial immunity.

Krüppel-ling of IRF4-Dependent DCs into Two Functionally Distinct DC Subsets.

IRF4-dependent DCs have been associated with induction of both Th1 and Th17 cells. In this issue of Immunity, Tussiwand et al. (2015) demonstrate that a dependence on KLF4 identifies a subset of IRF4-dependent DC that preferentially promotes Th2 cell differentiation.

Spatiotemporally Distinct Interactions with Dendritic Cell Subsets Facilitates CD4+ and CD8+ T Cell Activation to Localized Viral Infection.

The dynamics of when and where CD4(+) T cells provide help for CD8(+) T cell priming and which dendritic cells (DCs) activate CD4(+) T cells in vivo after localized infection are poorly understood. By using a cutaneous herpes simplex virus infection model combined with intravital 2-photon imaging of the draining lymph node (LN) to concurrently visualize pathogen-specific CD4(+) and CD8(+) T cells, we found that early priming of CD4(+) T cells involved clustering with migratory skin DCs. CD8(+) T cells did not interact with migratory DCs and their activation was delayed, requiring later clustering interactions with LN-resident XCR1(+) DCs. CD4(+) T cells interacted with these late CD8(+) T cell clusters on resident XCR1(+) DCs. Together, these data reveal asynchronous T cell activation by distinct DC subsets and highlight the key role of XCR1(+) DCs as the central platform for cytotoxic T lymphocyte activation and the delivery of CD4(+) T cell help.

The unexpected contribution of conventional type 1 dendritic cells in driving antibody responses.

Antibodies are hallmarks of most effective vaccines. For successful T-dependent antibody responses, conventional dendritic cells (cDC) have been largely attributed the role of priming T cells. By contrast, follicular dendritic cells and macrophages have been seen as responsible for B cell activation, due to their strategic location within secondary lymphoid tissues and capacity to present native antigen to B cells. This review summarizes the mounting evidence that cDC can also present native antigen to B cells. cDC2 have been the main subset linked to humoral responses, based largely on their favorable location, capacity to prime CD4+ T cells, and ability to present native antigen to B cells. However, studies using strategies to deliver antigen to receptors on cDC1, reveal this subset can also contribute to naïve B cell activation, as well as T cell priming. cDC1 location within lymphoid tissues reveals their juxtaposition to B cell follicles, with ready access to B cells for presentation of native antigen. These findings support the view that both cDC1 and cDC2 are capable of initiating humoral responses provided antigen is captured by relevant surface receptors attuned to this process. Such understanding is fundamental for the development of innovative humoral vaccination approaches.

Extrafollicular CD4 T cell-derived IL-10 functions rapidly and transiently to support anti-Plasmodium humoral immunity.

Immunity against malaria depends on germinal center (GC)-derived antibody responses that are orchestrated by T follicular helper (TFH) cells. Emerging data show that the regulatory cytokine IL-10 plays an essential role in promoting GC B cell responses during both experimental malaria and virus infections. Here we investigated the cellular source and temporal role of IL-10, and whether IL-10 additionally signals to CD4 T-cells to support anti-Plasmodium humoral immunity. Distinct from reports of virus infection, we found that IL-10 was expressed by conventional, Foxp3-negative effector CD4 T cells and functioned in a B cell-intrinsic manner only during the first 96 hours of Plasmodium infection to support humoral immunity. The critical functions of IL-10 manifested only before the orchestration of GC responses and were primarily localized outside of B cell follicles. Mechanistically, our studies showed that the rapid and transient provision of IL-10 promoted B cell expression of anti-apoptotic factors, MHC class II, CD83, and cell-cell adhesion proteins that are essential for B cell survival and interaction with CD4 T cells. Together, our data reveal temporal features and mechanisms by which IL-10 critically supports humoral immunity during blood-stage Plasmodium infection, information that may be useful for developing new strategies designed to lessen the burden of malaria.

Type I interferon production elicits differential CD4+ T-cell responses in mice infected with Plasmodium berghei ANKA and P. chabaudi.

Plasmodium parasites that infect humans are highly polymorphic, and induce various infections ranging from an asymptomatic state to life-threatening diseases. However, how the differences between the parasites affect host immune responses during blood-stage infection remains largely unknown. We investigated the CD4+ T-cell immune responses in mice infected with P. berghei ANKA (PbA) or P. chabaudi chabaudi AS (Pcc) using PbT-II cells, which recognize a common epitope of these parasites. In the acute phase of infection, CD4+ T-cell responses in PbA-infected mice showed a lower involvement of Th1 cells and a lower proportion of Ly6Clo effector CD4+ T cells than those in Pcc-infected mice. Transcriptome analysis of PbT-II cells indicated that type I interferon (IFN)-regulated genes were expressed at higher levels in both Th1- and Tfh-type PbT-II cells from PbA-infected mice than those from Pcc-infected mice. Moreover, IFN-α levels were considerably higher in PbA-infected mice than in Pcc-infected mice. Inhibition of type I IFN signaling increased PbT-II and partially reversed the Th1 over Tfh bias of the PbT-II cells in both PbA- and Pcc-infected mice. In the memory phase, PbT-II cells in PbA-primed mice maintained higher numbers and exhibited a better recall response to the antigen. However, recall responses were not significantly different between the infection groups after re-challenge with PbA, suggesting the effect of the inflammatory environment by the infection. These observations suggest that the differences in Plasmodium-specific CD4+ T-cell responses between PbA- and Pcc-infected mice were associated with the difference in type I IFN production during the early phase of the infection.