Extended clinical and immunological phenotype and transplant outcome in CD27 and CD70 deficiency.

Biallelic mutations in the genes encoding CD27 or its ligand CD70 underlie inborn errors of immunity (IEIs) characterized predominantly by Epstein-Barr virus (EBV)-associated immune dysregulation, such as chronic viremia, severe infectious mononucleosis, hemophagocytic lymphohistiocytosis (HLH), lymphoproliferation, and malignancy. A comprehensive understanding of the natural history, immune characteristics, and transplant outcomes has remained elusive. Here, in a multi-institutional global collaboration, we collected the clinical information of 49 patients from 29 families (CD27, n = 33; CD70, n = 16), including 24 previously unreported individuals and identified a total of 16 distinct mutations in CD27, and 8 in CD70, respectively. The majority of patients (90%) were EBV+ at diagnosis, but only ∼30% presented with infectious mononucleosis. Lymphoproliferation and lymphoma were the main clinical manifestations (70% and 43%, respectively), and 9 of the CD27-deficient patients developed HLH. Twenty-one patients (43%) developed autoinflammatory features including uveitis, arthritis, and periodic fever. Detailed immunological characterization revealed aberrant generation of memory B and T cells, including a paucity of EBV-specific T cells, and impaired effector function of CD8+ T cells, thereby providing mechanistic insight into cellular defects underpinning the clinical features of disrupted CD27/CD70 signaling. Nineteen patients underwent allogeneic hematopoietic stem cell transplantation (HSCT) prior to adulthood predominantly because of lymphoma, with 95% survival without disease recurrence. Our data highlight the marked predisposition to lymphoma of both CD27- and CD70-deficient patients. The excellent outcome after HSCT supports the timely implementation of this treatment modality particularly in patients presenting with malignant transformation to lymphoma.

The TNFRSF members CD27 and OX40 coordinately limit TH17 differentiation in regulatory T cells.

Regulatory T cells (Tregs) are closely related to TH17 cells and use aspects of the TH17-differentiation program for optimal immune regulation. In several chronic inflammatory human diseases, Tregs express IL-17A, suggesting that dysregulation of TH17-associated pathways in Tregs may result in either loss of suppressive function and/or conversion into pathogenic cells. The pathways that regulate the TH17 program in Tregs are poorly understood. We have identified two TNF receptor superfamily (TNFRSF) members, CD27 and OX40, that are preferentially expressed by skin-resident Tregs Both CD27 and OX40 signaling suppressed the expression of TH17-associated genes from Tregs in a cell-intrinsic manner in vitro and in vivo. However, only OX40 played a nonredundant role in promoting Treg accumulation. Tregs that lacked both CD27 and OX40 were defective in controlling skin inflammation and expressed high levels of IL-17A, as well as the master TH17 transcription factor, RORγt. Last, we found that CD27 expression was inversely correlated with Treg IL-17 production in skin of patients with psoriasis and hidradenitis suppurativa. Together, our results suggest that TNFRSF members play both redundant and distinct roles in regulating Treg plasticity in tissues.

The role of CD27-CD70-mediated T cell co-stimulation in vasculogenesis, arteriogenesis and angiogenesis.

BACKGROUND: T cells have a distinctive role in neovascularization, which consists of arteriogenesis and angiogenesis under pathological conditions and vasculogenesis under physiological conditions. However, the role of co-stimulation in T cell activation in neovascularization has yet to be established. The aim of this study was to investigate the role T cell co-stimulation and inhibition in angiogenesis, arteriogenesis and vasculogenesis. METHODS AND RESULTS: Hind limb ischemia was induced by double ligation of the left femoral artery in mice and blood flow recovery was measured with Laser Doppler Perfusion Imaging in control, CD70-/-, CD80/86-/-, CD70/80/86-/- and CTLA4+/- mice. Blood flow recovery was significantly impaired in mice lacking CD70 compared to control mice, but was similar in CD80/86-/-, CTLA4+/- and control mice. Mice lacking CD70 showed impaired vasculogenesis, since the number of pre-existing collaterals was reduced as observed in the pia mater compared to control mice. In vitro an impaired capability of vascular smooth muscle cells (VSMC) to activate T cells was observed in VSMC lacking CD70. Furthermore, CD70-/-, CD80/86-/- and CD70/80/86-/- mice showed reduced angiogenesis in the soleus muscle 10 days after ligation. Arteriogenesis was also decreased in CD70-/- compared to control mice 10 and 28 days after surgery. CONCLUSIONS: The present study is the first to describe an important role for T cell activation via co-stimulation in angiogenesis, arteriogenesis and vasculogenesis, where the CD27-CD70 T cell co-stimulation pathway appears to be the most important co-stimulation pathway in pre-existing collateral formation and post-ischemic blood flow recovery, by arteriogenesis and angiogenesis.

Aiolos Overexpression in Systemic Lupus Erythematosus B Cell Subtypes and BAFF-Induced Memory B Cell Differentiation Are Reduced by CC-220 Modulation of Cereblon Activity.

BAFF is a B cell survival and maturation factor implicated in the pathogenesis of systemic lupus erythematosus (SLE). In this in vitro study, we describe that soluble BAFF in combination with IL-2 and IL-21 is a T cell contact-independent inducer of human B cell proliferation, plasmablast differentiation, and IgG secretion from circulating CD27+ memory and memory-like CD27-IgD- double-negative (DN) B cells, but not CD27-IgD+ naive B cells. In contrast, soluble CD40L in combination with IL-2 and IL-21 induces these activities in both memory and naive B cells. Blood from healthy donors and SLE patients have similar circulating levels of IL-2, whereas SLE patients exhibit elevated BAFF and DN B cells and reduced IL-21. B cell differentiation transcription factors in memory, DN, and naive B cells in SLE show elevated levels of Aiolos, whereas Ikaros levels are unchanged. Treatment with CC-220, a modulator of the cullin ring ligase 4-cereblon E3 ubiquitin ligase complex, reduces Aiolos and Ikaros protein levels and BAFF- and CD40L-induced proliferation, plasmablast differentiation, and IgG secretion. The observation that the soluble factors BAFF, IL-2, and IL-21 induce memory and DN B cell activation and differentiation has implications for extrafollicular plasmablast development within inflamed tissue. Inhibition of B cell plasmablast differentiation by reduction of Aiolos and Ikaros may have utility in the treatment of SLE, where elevated levels of BAFF and Aiolos may prime CD27+ memory and DN memory-like B cells to become Ab-producing plasmablasts in the presence of BAFF and proinflammatory cytokines.

CD70 reverse signaling enhances NK cell function and immunosurveillance in CD27-expressing B-cell malignancies.

The interaction of the tumor necrosis factor receptor (TNFR) CD27 with its ligand CD70 is an emerging target to treat cancer. CD27 signaling provides costimulatory signals to cytotoxic T cells but also increases the frequency of regulatory T cells. Similar to other TNFR ligands, CD70 has been shown to initiate intracellular signaling pathways (CD70 reverse signaling). CD27 is expressed on a majority of B-cell non-Hodgkin lymphoma, but its role in the immune control of lymphoma and leukemia is unknown. We therefore generated a cytoplasmic deletion mutant of CD27 (CD27-trunc) to study the role of CD70 reverse signaling in the immunosurveillance of B-cell malignancies in vivo. Expression of CD27-trunc on malignant cells increased the number of tumor-infiltrating interferon γ-producing natural killer (NK) cells. In contrast, the antitumoral T-cell response remained largely unchanged. CD70 reverse signaling in NK cells was mediated via the AKT signaling pathway and increased NK cell survival and effector function. The improved immune control by activated NK cells prolonged survival of CD27-trunc-expressing lymphoma-bearing mice. Finally, CD70 reverse signaling enhanced survival and effector function of human NK cells in a B-cell acute lymphoblastic leukemia xenotransplants model. Therefore, CD70 reverse signaling in NK cells contributes to the immune control of CD27-expressing B-cell lymphoma and leukemia.

Donor-specific alloreactive T cells can be quantified from whole blood, and may predict cellular rejection after renal transplantation.

Preformed cellular alloreactivity can exist prior to transplantation and may contribute to rejection. Here, we used a rapid flow-cytometric whole-blood assay to characterize the extent of alloreactive T cells among 1491 stimulatory reactions from 61 renal transplant candidates and 75 controls. The role of preformed donor-specific alloreactive T cells in cellular rejection was prospectively analyzed in 21 renal transplant recipients. Alloreactive CD8+ T cells were more frequent than respective CD4+ T cells, and these levels were stable over time. CD8+ T cells were effector-memory T cells largely negative for expression of CD27, CD62L, and CCR7, and were susceptible to steroid and calcineurin inhibitor inhibition. Alloreactivity was more frequent in samples with higher number of HLA mismatches. Moreover, the percentage of individuals with alloreactive T cells was higher in transplant candidates than in controls. Among transplant candidates, 5/61 exhibited alloreactive CD8+ T cells against most stimulators, 23/61 toward a limited number of stimulators, and 33/61 did not show any alloreactivity. Among 21 renal transplant recipients followed prospectively, one had donor-specific preformed T-cell alloreactivity. She was the only patient who developed cellular rejection posttransplantation. In conclusion, donor-specific alloreactive T cells may be rapidly quantified from whole blood, and may predict cellular rejection after transplantation.

High-dimensional immune profiling of total and rotavirus VP6-specific intestinal and circulating B cells by mass cytometry.

In-depth phenotyping of human intestinal antibody secreting cells (ASCs) and their precursors is important for developing improved mucosal vaccines. We used single-cell mass cytometry to simultaneously analyze 34 differentiation and trafficking markers on intestinal and circulating B cells. In addition, we labeled rotavirus (RV) double-layered particles with a metal isotope and characterized B cells specific to the RV VP6 major structural protein. We describe the heterogeneity of the intestinal B-cell compartment, dominated by ASCs with some phenotypic and transcriptional characteristics of long-lived plasma cells. Using principal component analysis, we visualized the phenotypic relationships between major B-cell subsets in the intestine and blood, and revealed that IgM(+) memory B cells (MBCs) and naive B cells were phenotypically related as were CD27(-) MBCs and switched MBCs. ASCs in the intestine and blood were highly clonally related, but associated with distinct trajectories of phenotypic development. VP6-specific B cells were present among diverse B-cell subsets in immune donors, including naive B cells, with phenotypes representative of the overall B-cell pool. These data provide a high dimensional view of intestinal B cells and the determinants regulating humoral memory to a ubiquitous, mucosal pathogen at steady-state.

Novel mutations in TNFRSF7/CD27: Clinical, immunologic, and genetic characterization of human CD27 deficiency.

BACKGROUND: The clinical and immunologic features of CD27 deficiency remain obscure because only a few patients have been identified to date. OBJECTIVE: We sought to identify novel mutations in TNFRSF7/CD27 and to provide an overview of clinical, immunologic, and laboratory phenotypes in patients with CD27 deficiency. METHODS: Review of the medical records and molecular, genetic, and flow cytometric analyses of the patients and family members were performed. Treatment outcomes of previously described patients were followed up. RESULTS: In addition to the previously reported homozygous mutations c.G24A/p.W8X (n = 2) and c.G158A/p.C53Y (n = 8), 4 novel mutations were identified: homozygous missense c.G287A/p.C96Y (n = 4), homozygous missense c.C232T/p.R78W (n = 1), heterozygous nonsense c.C30A/p.C10X (n = 1), and compound heterozygous c.C319T/p.R107C-c.G24A/p.W8X (n = 1). EBV-associated lymphoproliferative disease/hemophagocytic lymphohistiocytosis, Hodgkin lymphoma, uveitis, and recurrent infections were the predominant clinical features. Expression of cell-surface and soluble CD27 was significantly reduced in patients and heterozygous family members. Immunoglobulin substitution therapy was administered in 5 of the newly diagnosed cases. CONCLUSION: CD27 deficiency is potentially fatal and should be excluded in all cases of severe EBV infections to minimize diagnostic delay. Flow cytometric immunophenotyping offers a reliable initial test for CD27 deficiency. Determining the precise role of CD27 in immunity against EBV might provide a framework for new therapeutic concepts.

CD27low natural killer cells prolong allograft survival in mice by controlling alloreactive CD8+ T cells in a T-bet-dependent manner.

BACKGROUND: Natural killer (NK) cells play a dichotomous role in alloimmune responses because they are known to promote both allograft survival and rejection. The aim of this study was to investigate the role of functionally distinct NK cell subsets in alloimmunity with the hypothesis that this dichotomy is explained by the functional heterogeneity of distinct NK cell subsets. METHODS: Because T-bet controls thematuration of NK cells from CD27high to terminally differentiated CD27low NK cells, we used Rag−/−T-bet−/− mice that lackmature CD27low NK cells to study the distinct roles of CD27low versus CD27high NK cells in a model of Tcell­mediated skin transplant rejection under costimulatory blockade conditions. RESULTS: We found that T cell­reconstituted Rag1−/− recipients (possessing CD27low NK cells) show significantly prolonged allograft survival on costimulatory blockade when compared to Rag1−/−T-bet−/− mice (lacking CD27low NK cells), indicating that CD27low but not CD27high NK cells enhance allograft survival. Critically, Rag1−/−T-bet−/− recipients showed strikingly increased alloreactive memory CD8+ Tcell responses, as indicated by increased CD8+ Tcell proliferation and interferon-γ production. Therefore, we speculated that CD27low NK cells directly regulate alloreactive CD8+ Tcell responses under costimulatory blockade conditions. To test this, we adoptively transferred CD27low NK cells into Rag1−/−T-bet−/− skin transplant recipients and found that the CD27low NK cells restore better allograft survival by inhibiting the proliferation of alloreactive interferon-γ+CD8+ T cells. CONCLUSIONS: In summary, mature CD27low NK cells promote allograft survival under costimulatory blockade conditions by regulating alloreactive memory CD8+ T-cell responses.

Murine CD27(-) Vγ6(+) γδ T cells producing IL-17A promote ovarian cancer growth via mobilization of protumor small peritoneal macrophages.

Cancer-associated inflammation mobilizes a variety of leukocyte populations that can inhibit or enhance tumor cell growth in situ. These subsets include γδ T cells, which can infiltrate tumors and typically provide large amounts of antitumor cytokines, such as IFN-γ. By contrast, we report here that in a well-established transplantable (ID8 cell line) model of peritoneal/ovarian cancer, γδ T cells promote tumor cell growth. γδ T cells accumulated in the peritoneal cavity in response to tumor challenge and could be visualized within solid tumor foci. Functional characterization of tumor-associated γδ T cells revealed preferential production of interleukin-17A (IL-17), rather than IFN-γ. Consistent with this finding, both T cell receptor (TCR)δ-deficient and IL-17-deficient mice displayed reduced ID8 tumor growth compared with wild-type animals. IL-17 production by γδ T cells in the tumor environment was essentially restricted to a highly proliferative CD27((-)) subset that expressed Vγ6 instead of the more common Vγ1 and Vγ4 TCR chains. The preferential expansion of IL-17-secreting CD27((-)) Vγ6((+)) γδ T cells associated with the selective mobilization of unconventional small peritoneal macrophages (SPMs) that, in comparison with large peritoneal macrophages, were enriched for IL-17 receptor A, and for protumor and proangiogenic molecular mediators, which were up-regulated by IL-17. Importantly, SPMs were uniquely and directly capable of promoting ovarian cancer cell proliferation. Collectively, this work identifies an IL-17-dependent lymphoid/myeloid cross-talk involving γδ T cells and SPMs that promotes tumor cell growth and thus counteracts cancer immunosurveillance.

The majority of human memory B cells recognizing RhD and tetanus resides in IgM+ B cells.

B cell memory to T cell-dependent (TD) Ags are considered to largely reside in class-switched CD27(+) cells. However, we previously observed that anti-RhD (D) Igs cloned from two donors, hyperimmunized with D(+) erythrocytes, were predominantly of the IgM isotype. We therefore analyzed in this study the phenotype and frequency of D- and tetanus toxoid-specific B cells by culturing B cells in limiting dilution upon irradiated CD40L-expressing EL4.B5 cells and testing the culture supernatant. Most Ag-specific B cells for both TD Ags were found to reside in the IgM-expressing B cells, including CD27(-) B cells, in both hyperimmunized donors and nonhyperimmunized volunteers. Only shortly after immunization a sharp increase in Ag-specific CD27(+)IgG(+) B cells was observed. Next, B cells were enriched with D(+) erythrocyte ghosts and sorted as single cells. Sequencing of IGHV, IGLV, IGKV, and BCL6 genes from these D-specific B cell clones demonstrated that both CD27(-)IgM(+) and CD27(+)IgM(+) B cells harbored somatic mutations, documenting their Ag-selected nature. Furthermore, sequencing revealed a clonal relationship between the CD27(-)IgM(+), CD27(+)IgM(+), and CD27(+)IgG(+) B cell subsets. These data strongly support the recently described multiple layers of memory B cells to TD Ags in mice, where IgM(+) B cells represent a memory reservoir which can re-enter the germinal center and ensure replenishment of class-switched memory CD27(+) B cells from Ag-experienced precursors.

A glycosylation-dependent CD45RB epitope defines previously unacknowledged CD27⁻IgM(high) B cell subpopulations enriched in young children and after hematopoietic stem cell transplantation.

The immune system is dysfunctional for years after hematopoietic stem cell transplantation (HSCT). A potential cause is an intrinsic B cell deficiency. In a cohort of pediatric HSCT patients few CD27(+) B cells formed after transplantation with the number of CD27(+)IgM(high) cells more affected than class-switched ones. A previously unacknowledged population of CD27(-)IgM(high) cells made up the majority of B cells and this population was also enlarged in healthy children compared to adults. Only a minority of these CD27(-)IgM(high) B cells expressed markers typical for transitional B cells, and the non-transitional CD27(-)IgM(high) cells could be further divided into subpopulations based on their ability to extrude the dye Rhodamine 123 and their expression of CD45RB(MEM55), a glycosylation-dependent epitope. Thus, we define several novel human CD27(-)IgM(high) B cell subpopulations in blood, all of which are present in higher frequencies and numbers in young children and after HSCT than in adults.

Combined immunodeficiency with life-threatening EBV-associated lymphoproliferative disorder in patients lacking functional CD27.

CD27, a tumor necrosis factor receptor family member, interacts with CD70 and influences T-, B- and NK-cell functions. Disturbance of this axis impairs immunity and memory generation against viruses including Epstein Barr virus (EBV), influenza, and others. CD27 is commonly used as marker of memory B cells for the classification of B-cell deficiencies including common variable immune deficiency. Flow cytometric immunophenotyping including expression analysis of CD27 on lymphoid cells was followed by capillary sequencing of CD27 in index patients, their parents, and non-affected siblings. More comprehensive genetic analysis employed single nucleotide polymorphism-based homozygosity mapping and whole exome sequencing. Analysis of exome sequencing data was performed at two centers using slightly different data analysis pipelines, each based on the Genome Analysis ToolKit Best Practice version 3 recommendations. A comprehensive clinical characterization was correlated to genotype. We report the simultaneous confirmation of human CD27 deficiency in 3 independent families (8 patients) due to a homozygous mutation (p. Cys53Tyr) revealed by whole exome sequencing, leading to disruption of an evolutionarily conserved cystein knot motif of the transmembrane receptor. Phenotypes varied from asymptomatic memory B-cell deficiency (n=3) to EBV-associated hemophagocytosis and lymphoproliferative disorder (LPD; n=3) and malignant lymphoma (n=2; +1 after LPD). Following EBV infection, hypogammaglobulinemia developed in at least 3 of the affected individuals, while specific anti-viral and anti-polysaccharide antibodies and EBV-specific T-cell responses were detectable. In severely affected patients, numbers of iNKT cells and NK-cell function were reduced. Two of 8 patients died, 2 others underwent allogeneic hematopoietic stem cell transplantation successfully, and one received anti-CD20 (rituximab) therapy repeatedly. Since homozygosity mapping and exome sequencing did not reveal additional modifying factors, our findings suggest that lack of functional CD27 predisposes towards a combined immunodeficiency associated with potentially fatal EBV-driven hemo-phagocytosis, lymphoproliferation, and lymphoma development.

Transient lymphocyte decrease due to adhesion and migration following catumaxomab (anti-EpCAM x anti-CD3) treatment in vivo

INTRODUCTION: In patients, a transient decrease in peripheral blood lymphocyte counts was observed following intraperitoneal administration of the trifunctional monoclonal antibody catumaxomab (anti-human EpCAM x anti-human CD3). The aim of this study was to clarify the observed effect in a preclinical mouse model and to analyse the related mechanism of action in vitro. MATERIALS AND METHODS: A related antibody, BiLu (antihuman EpCAM x anti-mouse CD3), was administered to mice and blood leukocytes were analysed. In vitro studies measured activation and cytokine secretion from human peripheral blood mononuclear cells (PBMC). For the analysis of T cell adhesion, PBMC were preincubated with catumaxomab and then co-cultured with human endothelial cells (HUVEC); T cell adhesion was assessed in the presence or absence of endothelial cell preactivation by TNFα. Adherent T cells were determined by flow cytometry. RESULTS: Treatment of mice with BiLu resulted in a dosedependent transient decrease in CD3+ T cells (both CD4+ and CD8+) that returned to the normal range within 48 h. Catumaxomab physiologically activated T cells in vitro (increased CD69 expression) and induced cytokine release (TNFα, IFNγ). TNFα increased expression of adhesion molecules CD54 and CD62E on endothelial cells. Furthermore, catumaxomab dose-dependently enhanced adhesion of T cells to endothelial cells. Adhesion was further increased when endothelial cells were preactivated with TNFα. CONCLUSIONS: Catumaxomab increases adhesion of T cells to endothelial cells due to antibody-mediated activation of T cells and production of T cell cytokines that up-regulate endothelial cell adhesion molecules. These results provide a mechanistic rationale for the transient, reversible decrease in lymphocyte counts observed following catumaxomab administration in patients, which is likely to be due to redistribution of lymphocytes (AU)

CD27-deficient mice show normal NK-cell differentiation but impaired function upon stimulation.

Natural killer (NK) cells are part of the first line defense against tumors, parasites and virus-infected cells. Therefore, factors that control NK-cell numbers and their function are important. CD27 is constitutively expressed on NK cells and its expression correlates with sequential phases in NK-cell development, discriminating phenotypically and functionally different subsets within the NK-cell population. Although CD27 has been described to have an important regulatory role in effector and memory T and B lymphocytes, its role in NK-cell biology remains to be addressed. In this study, we used CD27(-/-) mice to investigate the role of CD27 in NK-cell development and function, both during the resting state and upon stimulation. The results show that NK-cell numbers are not impaired in CD27(-/-) mice. Moreover, CD27(-/-) NK cells reach full phenotypic maturity, evidenced by normal expression of CD49b, CD43 and CD11b. Expression of activating receptors is unaltered, whereas expression of several inhibitory receptors is increased. Cytotoxicity and interferon-γ production by NK cells from CD27(-/-) mice in the resting state are normal. However, upon in vivo anti-CD40- or poly-I:C-mediated activation, or in vitro interleukin-15 priming plus anti-NKp46 stimulation, the absence of CD27 results in decreased cytolytic activity and cytokine production by spleen and liver NK cells. In conclusion, this study demonstrates that CD27 is dispensable for the development of functional NK cells. However, upon stimulation of NK cells, CD27 displays an important role in their activation and functionality.

p53 target Siva regulates apoptosis in ischemic kidneys.

The role of p53 in inducing apoptosis following acute kidney injury is well-established; however, the molecular mechanisms remain largely unknown. We report here that the p53 proapoptotic target Siva and its receptor CD27, a member of the tumor necrosis factor receptor family, are upregulated following renal ischemia-reperfusion injury (IRI). Inhibition of Siva using antisense oligonucleotides conferred functional and morphological protection, and it prevented apoptosis postrenal IRI in mice. Renal IRI in CD27-deficient mice displayed functional protection and partial inhibition of apoptosis, suggesting an incomplete role for CD27 in Siva-mediated apoptosis. To further elucidate mechanisms by which Siva elicits apoptosis, in vitro studies were performed. In Siva-transfected LLC-PK(1)cells, Siva is persistently expressed in the nucleus at 3 h onwards and its translocation to mitochondria and the plasma membrane occurred at 6 h. Moreover, Siva overexpression induced mitochondrial permeability, cytochrome c release, caspase-8 and -9 activation, translocation of apoptosis-inducing factor (AIF) to the nucleus, and apoptosis. Inhibition of Siva in ischemic kidneys prevented mitochondrial release of cytochrome c and AIF. These data indicate that Siva function is pivotal in regulating apoptosis in the pathology of renal IRI. Targeting Siva may offer a potential therapeutic strategy for renal IRI.

The TNFR family members OX40 and CD27 link viral virulence to protective T cell vaccines in mice.

Induction of CD8+ T cell immunity is a key characteristic of an effective vaccine. For safety reasons, human vaccination strategies largely use attenuated nonreplicating or weakly replicating poxvirus-based vectors, but these often elicit poor CD8+ T cell immunity and might not result in optimal protection. Recent studies have suggested that virulence is directly linked to immunogenicity, but the molecular mechanisms underlying optimal CD8+ T cell responses remain to be defined. Here, using natural and recombinant vaccinia virus (VACV) strains, we have shown in mice that VACV strains of differing virulence induce distinct levels of T cell memory because of the differential use of TNF receptor (TNFR) family costimulatory receptors. With strongly replicating (i.e., virulent) VACV, the TNFR family costimulatory receptors OX40 (also known as CD134) and CD27 were engaged and promoted the generation of high numbers of memory CD8+ T cells, which protected against a lethal virus challenge in the absence of other mechanisms, including antibody and help from CD4+ T cells. In contrast, weakly replicating (i.e., low-virulence) VACV strains were poor at eliciting protective CD8+ T cell memory, as only the Ig family costimulatory receptor CD28 was engaged, and not OX40 or CD27. Our results suggest that the virulence of a virus dictates costimulatory receptor usage to determine the level of protective CD8+ T cell immunity.

The Pim kinase pathway contributes to survival signaling in primed CD8+ T cells upon CD27 costimulation.

Stimulation of the costimulatory receptor CD27 by its ligand CD70 has proved important for the generation of primary and memory CD8(+) T cell responses in various models of antigenic challenge. CD27/CD70-mediated costimulation promotes the survival of primed T cells and thereby increases the size of effector and memory populations. In this paper, we reveal molecular mechanisms underlying the prosurvival effect of CD27. CD27 signaling upregulated expression of the antiapoptotic Bcl-2 family member Bcl-x(L). However, genetic reconstitution of Cd27(-/-) CD8(+) T cells with Bcl-x(L) alone or in combination with the related protein Mcl-1 did not compensate for CD27 deficiency in the response to influenza virus infection. This suggested that CD27 supports generation of the CD8(+) effector T cell pool not only by counteracting apoptosis via Bcl-2 family members. Genome-wide mRNA expression profiling indicated that CD27 directs expression of the Pim1 gene. Pim-1 is a serine/threonine kinase that sustains survival of rapidly proliferating cells by antiapoptotic and prometabolic effects that are independent of the mammalian target of rapamycin (mTOR) pathway. In TCR-primed CD8(+) T cells, CD27 could increment Pim-1 protein expression and promote cell survival throughout clonal expansion independent of the mTOR and IL-2R pathways. In addition, introduction of the Pim1 gene in Cd27(-/-) CD8(+) T cells partially corrected their defect in clonal expansion and formation of an effector pool. We conclude that CD27 may contribute to the survival of primed CD8(+) T cells by the upregulation of antiapoptotic Bcl-2 family members but also calls the Pim-1 kinase survival pathway into action.

Cutting edge: adaptive versus innate receptor signals selectively control the pool sizes of murine IFN-γ- or IL-17-producing γδ T cells upon infection.

γδ T lymphocytes are commonly viewed as embracing properties of both adaptive and innate immunity. Contributing to this is their responsiveness to pathogen products, either with or without the involvement of the TCR and its coreceptors. This study clarifies this paradoxical behavior by showing that these two modes of responsiveness are the properties of two discrete sets of murine lymphoid γδ T cells. Thus, MyD88 deficiency severely impaired the response to malaria infection of CD27((-)), IL-17A-producing γδ T cells, but not of IFN-γ-producing γδ cells. Instead, the latter compartment was severely contracted by ablating CD27, which synergizes with TCRγδ in the induction of antiapoptotic mediators and cell cycle-promoting genes in CD27((+)), IFN-γ-secreting γδ T cells. Hence, innate versus adaptive receptors differentially control the peripheral pool sizes of discrete proinflammatory γδ T cell subsets during immune responses to infection.

Reduced CD27 expression on antigen-specific CD4+ T cells correlates with persistent active tuberculosis.

OBJECTIVE: CD27, a member of the tumor necrosis factor receptor family, has important role in generation of T cell immunity. In this study, association of CD27 expression on mycobacterial antigen-specific CD4+ T cells with pulmonary tuberculosis (TB) was investigated. METHODS: Mycobacterial antigen-specific CD4+ T cells were identified based on CD154 expression and CD27 expression on antigen-specific CD4 T cells was analyzed by flow cytometry. RESULTS: Compared with tuberculin-positive controls, patients with bacterial culture-positive pulmonary TB had significantly reduced CD27 expression on antigen-specific CD4 T cells. The persistent active TB patients had much lower percentages of CD27+ antigen-specific CD4 T cells than culture-positive new TB patients (P=0.008) and healthy controls (P=0.005). Logistic regression analysis on frequencies of CD27-expressing antigen-specific CD4 T cells and TB patients' clinical characteristics indicated that low percentage of CD27+ antigen-specific CD4 T cells correlated significantly with persistent active tuberculosis (P=0.002, odds ratio=19.6). CONCLUSION: It is concluded that frequency of CD27+ antigen-specific CD4 T cells could be used as an immunological marker for persistent active TB.