James M. Ertelt

Immunosuppressive CD71+ erythroid cells compromise neonatal host defence against infection

Newborn infants are highly susceptible to infection. This defect in host defence has generally been ascribed to the immaturity of neonatal immune cells; however, the degree of hyporesponsiveness is highly variable and depends on the stimulation conditions. These discordant responses illustrate the need for a more unified explanation for why immunity is compromised in neonates. Here we show that physiologically enriched CD71+ erythroid cells in neonatal mice and human cord blood have distinctive immunosuppressive properties. The production of innate immune protective cytokines by adult cells is diminished after transfer to neonatal mice or after co-culture with neonatal splenocytes. Neonatal CD71+ cells express the enzyme arginase-2, and arginase activity is essential for the immunosuppressive properties of these cells because molecular inhibition of this enzyme or supplementation with l-arginine overrides immunosuppression. In addition, the ablation of CD71+ cells in neonatal mice, or the decline in number of these cells as postnatal development progresses parallels the loss of suppression, and restored resistance to the perinatal pathogens Listeria monocytogenes and Escherichia coli. However, CD71+ cell-mediated susceptibility to infection is counterbalanced by CD71+ cell-mediated protection against aberrant immune cell activation in the intestine, where colonization with commensal microorganisms occurs swiftly after parturition. Conversely, circumventing such colonization by using antimicrobials or gnotobiotic germ-free mice overrides these protective benefits. Thus, CD71+ cells quench the excessive inflammation induced by abrupt colonization with commensal microorganisms after parturition. This finding challenges the idea that the susceptibility of neonates to infection reflects immune-cell-intrinsic defects and instead highlights processes that are developmentally more essential and inadvertently mitigate innate immune protection. We anticipate that these results will spark renewed investigation into the need for immunosuppression in neonates, as well as improved strategies for augmenting host defence in this vulnerable population.

Pathogen-specific Treg cells expand early during mycobacterium tuberculosis infection but are later eliminated in response to Interleukin-12.

Thymically derived Foxp3⁺ regulatory T (Treg) cells have a propensity to recognize self-peptide:MHC complexes, but their ability to respond to epitope-defined foreign antigens during infectious challenge has not been demonstrated. Here we show that pulmonary infection with Mycobacterium tuberculosis (Mtb), but not Listeria monocytogenes (Lm), induced robust lymph node expansion of a highly activated population of pathogen-specific Treg cells from the pre-existing pool of thymically derived Treg cells. These antigen-specific Treg cells peaked in numbers 3 weeks after infection but subsequently underwent selective elimination driven, in part, by interleukin-12-induced intrinsic expression of the Th1-cell-promoting transcription factor T-bet. Thus, the initial Mtb-induced inflammatory response promotes pathogen-specific Treg cell proliferation, but these cells are actively culled later, probably to prevent suppression during later stages of infection. These findings have important implications for the prevention and treatment of tuberculosis and other chronic diseases in which antigen-specific Treg cells restrict immunity.

Proteolytic elimination of N-myristoyl modifications by the Shigella virulence factor IpaJ

Protein N-myristoylation is a 14-carbon fatty-acid modification that is conserved across eukaryotic species and occurs on nearly 1% of the cellular proteome. The ability of the myristoyl group to facilitate dynamic protein–protein and protein–membrane interactions (known as the myristoyl switch) makes it an essential feature of many signal transduction systems. Thus pathogenic strategies that facilitate protein demyristoylation would markedly alter the signalling landscape of infected host cells. Here we describe an irreversible mechanism of protein demyristoylation catalysed by invasion plasmid antigen J (IpaJ), a previously uncharacterized Shigella flexneri type III effector protein with cysteine protease activity. A yeast genetic screen for IpaJ substrates identified ADP-ribosylation factor (ARF)1p and ARF2p, small molecular mass GTPases that regulate cargo transport through the Golgi apparatus. Mass spectrometry showed that IpaJ cleaved the peptide bond between N-myristoylated glycine-2 and asparagine-3 of human ARF1, thereby providing a new mechanism for host secretory inhibition by a bacterial pathogen. We further demonstrate that IpaJ cleaves an array of N-myristoylated proteins involved in cellular growth, signal transduction, autophagasome maturation and organelle function. Taken together, these findings show a previously unrecognized pathogenic mechanism for the site-specific elimination of N-myristoyl protein modification.

CD4+ T Cell Tolerance to Tissue-Restricted Self Antigens Is Mediated by Antigen-Specific Regulatory T Cells Rather Than Deletion

Deletion of self-antigen-specific T cells during thymic development provides protection from autoimmunity. However, it is unclear how efficiently this occurs for tissue-restricted self antigens, or how immune tolerance is maintained for self-antigen-specific T cells that routinely escape deletion. Here we show that endogenous CD4+ T cells with specificity for a set of tissue-restricted self antigens were not deleted at all. For pancreatic self antigen, this resulted in an absence of steady-state tolerance, while for the lung and intestine, tolerance was maintained by the enhanced presence of thymically-derived antigen-specific Foxp3+ regulatory T (Treg) cells. Unlike deletional tolerance, Treg cell-mediated tolerance was broken by successive antigen challenges. These findings reveal that for some tissue-restricted self antigens, tolerance relies entirely on nondeletional mechanisms that are less durable than T cell deletion. This might explain why autoimmunity is often tissue-specific, and it offers a rationale for cancer vaccine strategies targeting tissue-restricted tumor antigens.

Cross-Generational Reproductive Fitness Enforced by Microchimeric Maternal Cells

Exposure to maternal tissue during in utero development imprints tolerance to immunologically foreign non-inherited maternal antigens (NIMA) that persists into adulthood. The biological advantage of this tolerance, conserved across mammalian species, remains unclear. Here, we show maternal cells that establish microchimerism in female offspring during development promote systemic accumulation of immune suppressive regulatory T cells (Tregs) with NIMA specificity. NIMA-specific Tregs expand during pregnancies sired by males expressing alloantigens with overlapping NIMA specificity, thereby averting fetal wastage triggered by prenatal infection and non-infectious disruptions of fetal tolerance. Therefore, exposure to NIMA selectively enhances reproductive success in second-generation females carrying embryos with overlapping paternally inherited antigens. These findings demonstrate that genetic fitness, canonically thought to be restricted to Mendelian inheritance, is enhanced in female placental mammals through vertically transferred maternal cells that promote conservation of NIMA and enforce cross-generational reproductive benefits.

CXCR3 blockade protects against Listeria monocytogenes infection–induced fetal wastage

Mammalian pregnancy requires protection against immunological rejection of the developing fetus bearing discordant paternal antigens. Immune evasion in this developmental context entails silenced expression of chemoattractant proteins (chemokines), thereby preventing harmful immune cells from penetrating the maternal-fetal interface. Here, we demonstrate that fetal wastage triggered by prenatal Listeria monocytogenes infection is driven by placental recruitment of CXCL9-producing inflammatory neutrophils and macrophages that promote infiltration of fetal-specific T cells into the decidua. Maternal CD8+ T cells with fetal specificity upregulated expression of the chemokine receptor CXCR3 and, together with neutrophils and macrophages, were essential for L. monocytogenes-induced fetal resorption. Conversely, decidual accumulation of maternal T cells with fetal specificity and fetal wastage were extinguished by CXCR3 blockade or in CXCR3-deficient mice. Remarkably, protection against fetal wastage and in utero L. monocytogenes invasion was maintained even when CXCR3 neutralization was initiated after infection, and this protective effect extended to fetal resorption triggered by partial ablation of immune-suppressive maternal Tregs, which expand during pregnancy to sustain fetal tolerance. Together, our results indicate that functionally overriding chemokine silencing at the maternal-fetal interface promotes the pathogenesis of prenatal infection and suggest that therapeutically reinforcing this pathway represents a universal approach for mitigating immune-mediated pregnancy complications.

Regulatory T Cells: New Keys for Further Unlocking the Enigma of Fetal Tolerance and Pregnancy Complications

The immunological alterations required for successful pregnancy in eutherian placental mammals have remained a scientific enigma since the discovery of MHC haplotype diversity and unique immune signatures among individuals. Within the past 10 years, accumulating data suggest that immune-suppressive regulatory T cells (Tregs) confer essential protective benefits in sustaining tolerance to the semiallogeneic fetus during pregnancy, along with their more established roles in maintaining tolerance to self and “extended self” commensal Ags that averts autoimmunity. Reciprocally, many human pregnancy complications stemming from inadequacies in fetal tolerance have been associated with defects in maternal Tregs. Thus, further elucidating the immunological shifts during pregnancy not only have direct translational implications for improving perinatal health, they have enormous potential for unveiling new clues about how Tregs work in other biological contexts. In this article, epidemiological data in human pregnancy and complementary animal studies implicating a pivotal protective role for maternal Tregs are summarized.

Cutting edge: committed Th1 CD4+ T cell differentiation blocks pregnancy-induced Foxp3 expression with antigen-specific fetal loss.

Pregnancy stimulates induced Foxp3 expression among maternal CD4+ T cells with fetal specificity. Although sustained maternal regulatory CD4+ T cell (Treg) expansion is essential for maintaining fetal tolerance during pregnancy, the necessity for Foxp3+ cells with fetal specificity remains undefined. In this study, we demonstrate that mitigating Treg differentiation among maternal CD4+ T cells with a single surrogate fetal specificity elicits Ag-specific fetal loss. Using recombinant Listeria monocytogenes to prime stably differentiated Th1 CD4+ T cells with fetal I-Ab:2W1S55–68 specificity refractory to pregnancy-induced Foxp3 expression, we show that Ag delivery by cytoplasmic L. monocytogenes causes selective loss of 2W1S+ offspring through CD4 cell– and IFN-γ–dependent pathways. In contrast, CD4+ T cells primed by L. monocytogenes restricted from the cell cytoplasm are markedly more plastic for induced Foxp3 expression, with normal pregnancy outcomes. Thus, committed Th1 polarization blocks pregnancy induced Treg differentiation among maternal CD4+ T cells with fetal specificity and triggers Ag-specific fetal loss.

B7-1/B7-2 blockade overrides the activation of protective CD8 T cells stimulated in the absence of Foxp3 regulatory T cells

Although T cell activation has been classically described to require distinct, positive stimulation signals that include B7‐1 (CD80) and B7‐2 (CD86) costimulation, overriding suppression signals that avert immune‐mediated host injury are equally important. How these opposing stimulation and suppression signals work together remains incompletely defined. Our recent studies demonstrate that CD8 Teff activation in response to cognate peptide stimulation is actively suppressed by the Foxp3+ subset of CD4 cells, called Tregs. Here, we show that the elimination of Treg suppression does not bypass the requirement for positive B7‐1/B7‐2 costimulation. The expansion, IFN‐γ cytokine production, cytolytic, and protective features of antigen‐specific CD8 T cells stimulated with purified cognate peptide in Treg‐ablated mice were each neutralized effectively by CTLA‐4‐Ig that blocks B7‐1/B7‐2. In turn, given the efficiency whereby CTLA‐4‐Ig overrides the effects of Treg ablation, the role of Foxp3+ cell‐intrinsic CTLA‐4 in mitigating CD8 Teff activation was also investigated. With the use of mixed chimera mice that contain CTLA‐4‐deficient Tregs exclusively after the ablation of WT Foxp3+ cells, a critical role for Treg CTLA‐4 in suppressing the expansion, cytokine production, cytotoxicity, and protective features of peptide‐stimulated CD8 T cells is revealed. Thus, the activation of protective CD8 T cells requires positive B7‐1/B7‐2 costimulation even when suppression by Tregs and in particular, Treg‐intrinsic CTLA‐4 is circumvented.

Tolerance to noninherited maternal antigens, reproductive microchimerism and regulatory T cell memory: 60 years after ‘Evidence for actively acquired tolerance to Rh antigens’

ABSTRACT Compulsory exposure to genetically foreign maternal tissue imprints in offspring sustained tolerance to noninherited maternal antigens (NIMA). Immunological tolerance to NIMA was first described by Dr. Ray D. Owen for women genetically negative for erythrocyte rhesus (Rh) antigen with reduced sensitization from developmental Rh exposure by their mothers. Extending this analysis to HLA haplotypes has uncovered the exciting potential for therapeutically exploiting NIMA-specific tolerance naturally engrained in mammalian reproduction for improved clinical outcomes after allogeneic transplantation. Herein, we summarize emerging scientific concepts stemming from tolerance to NIMA that includes postnatal maintenance of microchimeric maternal origin cells in offspring, expanded accumulation of immune suppressive regulatory T cells with NIMA-specificity, along with teleological benefits and immunological consequences of NIMA-specific tolerance conserved across mammalian species.