Katrin Hebel

Plasma cell differentiation in T-independent type 2 immune responses is independent of CD11chigh dendritic cells

Dendritic cells (DC) play an important role as antigen‐presenting cells in T cell stimulation. Interestingly, a number of recent studies also imply DC as critical accessory cells in B cell activation, isotype switching and plasma blast maintenance. Here we use the conditional in vivo ablation of CD11chigh DC to investigate the role of these cells in T‐independent type 2 immune responses. We show that CD11chigh DC are dispensable for the initiation and maintenance of a primary immune response against the T‐independent type 2 antigen (4‐hydroxy‐3‐nirophenyl)acetyl‐Ficoll. Our results suggest that support for plasma cell formation in T cell‐independent immune responses can be provided by non‐DC such as stromal cells, or is independent of external signals. Interestingly, we found plasma blasts to express CD11c and to be diphtheria toxin‐sensitive in CD11c‐diphtheria toxin receptor‐transgenic mice, providing a unique tool for future analysis of in vivo aspects of plasma cell biology.

IL-1β and TGF-β act antagonistically in induction and differentially in propagation of human proinflammatory precursor CD4+ T cells.

Cytokines are critical messengers that control the differentiation of Th cells. To evaluate their impact on the fate of human naive CD4+ T cells from cord and adult blood, early T cell differentiation was monitored after T cell activation in the presence of pro- and anti-inflammatory cytokines. Interestingly, the analysis of Th cell lineage-specific molecules revealed that IL-1β on its own mediates differentiation of Th cells that secrete a wide range of proinflammatory cytokines and stably express CD69, STAT1, IFN-γ, and IL-17. Notably, our data suggest that IL-1β induces Th17 cells independent of RORC upregulation. In contrast, TGF-β that triggers RORC prevents Th17 cell development. This suppressive function of TGF-β is characterized by inhibition of STAT1, STAT3, and CD69. However, after repeated anti-CD3 and anti-CD28 stimulation, we observe that TGF-β provokes an increase in Th17 cells that presumably relies on reactivation of a default pathway by preferential inhibition of IFN-γ. Hence, our data extend the view that the principal cytokines for determining Th cell fate are IL-12 for the Th1 lineage, IL-4 for the Th2 lineage, and TGF-β in conjunction with IL-6 for the Th17 lineage. We propose that IL-1β induces a general proinflammatory Th cell precursor that, in the presence of the lineage-specifying cytokines, further differentiates into one of the specific Th cell subpopulations.

CD4+ T Cells from Human Neonates and Infants Are Poised Spontaneously To Run a Nonclassical IL-4 Program

Senescence or biological aging impacts a vast variety of molecular and cellular processes. To date, it is unknown whether CD4+ Th cells display an age-dependent bias for development into specific subpopulations. In this study, we show the appearance of a distinct CD4+ T cell subset expressing IL-4 at an early stage of development in infant adenoids and cord blood that is lost during aging. We identified by flow cytometric, fluorescent microscopic, immunoblot, and mass spectrometric analysis a population of CD4+ T cells that expressed an unglycosylated isoform of IL-4. This T cell subpopulation was found in neonatal but not in adult CD4+ T cells. Furthermore, we show that the mRNA of the Th2 master transcription factor GATA3 is preferentially expressed in neonatal CD4+ T cells. The Th2 phenotype of the IL-4+CD4+ T cells could be reinforced in the presence of TGF-β. Although the IL-4+CD4+ T cells most likely originate from CD31+CD4+ T recent thymic emigrants, CD31 was downregulated prior to secretion of IL-4. Notably, the secretion of IL-4 requires a so far unidentified trigger in neonatal T cells. This emphasizes that cytokine expression and secretion are differentially regulated processes. Our data support the hypothesis of an endogenously poised cytokine profile in neonates and suggest a link between cytokine production and the developmental stage of an organism. The determination of the IL-4 isoform–expressing cells in humans might allow the identification of Th2 precursor cells, which could provide novel intervention strategies directed against Th2-driven immunopathologies such as allergies.

Blockade of CTLA-4 Decreases the Generation of Multifunctional Memory CD4+ T Cells In Vivo

CTLA-4 is known as a central inhibitor of T cell responses. It terminates T cell activation and proliferation and induces resistance against activation induced cell death. However, its impact on memory formation of adaptive immune responses is still unknown. In this study, we demonstrate that although anti–CTLA-4 mAb treatment during primary immunization of mice initially enhances the number of IFN-γ–producing CD4+ T cells, it does not affect the size of the memory pool. Interestingly, we find that the CTLA-4 blockade modulates the quality of the memory pool: it decreases the amount of specialized “multifunctional” memory CD4+ T cells coproducing IFN-γ, TNF-α, and IL-2 in response to Ag. The reduction of these cells causes an immense decrease of IFN-γ–producing T cells after in vivo antigenic rechallenge. Chimeric mice expressing CTLA-4–competent and –deficient cells unmask, which these CTLA-4–driven mechanisms are mediated CD4+ T cell nonautonomously. In addition, the depletion of CD25+ T cells prior to the generation of Ag-specific memory cells reveals that the constitutively CTLA-4–expressing natural regulatory T cells determine the quality of memory CD4+ T cells. Taken together, these results indicate that although the inhibitory molecule CTLA-4 damps the primary immune response, its engagement positively regulates the formation of a high-quality memory pool equipped with multifunctional CD4+ T cells capable of mounting a robust response to Ag rechallenge.

Pro-inflammatory effector Th cells transmigrate through anti-inflammatory environments into the murine fetus

The presence of maternal DNA or even maternal cells within the offspring (microchimerism) has been reported for many fetal tissues, including the liver, heart, and spleen. Microchimerism is believed to be involved in the pathogenesis of autoimmune diseases; however, the cellular origin of this phenomenon remains unknown. Here, we determined whether differentiated T lymphocytes could transmigrate through the immunosuppressive environment of the placenta to reach the fetus. In vitro-differentiated effector/memory Th1 and Th17 cells from OVA₃₂₃₋₃₃₉-specific TCR(tg) T cells of OT-II mice were adoptively transferred (i.v.) into the tail veins of pregnant Ly5.1 mice at d15 and d19 of gestation. Mice were then sacrificed 40 h after adoptive cell transfer. Using radioactive labeling of T cells with sodium chromate [Cr⁵¹] prior to adoptive transfer, we observed that homing of pro-inflammatory Th cells was equally efficient in both pregnant and non-pregnant mice. Transmigration of Th1- and Th17-like cells through the highly immunosuppressive environment of the placenta into the fetus was significantly enhanced in experimental mice compared to control mice (P < 0.0001). In addition, a substantial amount of effector Th cells accumulated in the placenta. Finally, we found that treatment with Pertussis Toxin resulted in a 3-fold increase in the transmigration of effector Th17 cells into the fetus (P < 0.0001). When pro-inflammatory Th1-or Th17-like cells were injected into syngeneic mothers, almost all of the fetuses analyzed exhibited radioactivity, suggesting that transmigration of effector T cells occurs frequently. Our results suggest the possibility of novel roles for these maternal effector cells in the pathogenesis or reduction of disease.

Surface CD152 (CTLA-4) Expression and Signaling Dictates Longevity of CD28null T Cells

CD28null T cells are a highly enriched subset of proinflammatory T cells in patients with autoimmune diseases that are oligoclonal and autoreactive. In this study, we analyzed the role of CD152 signaling on the longevity of human CD28null T cells. Using a sensitive staining method for CD152, we show that human CD4+CD28null and CD8+CD28null T cells rapidly express surface CD152. Serological inactivation of CD152 using specific Fab or blockade of CD152 ligands using CTLA-4Ig in CD4+CD28null and CD8+CD28null T cells enhances apoptosis in a Fas/FasL-dependent manner. CD152 cross-linking on activated CD28null cells prevents activation-induced cell death as a result of reduced caspase activity. Apoptosis protection conferred by CD152 is mediated by phosphatidylinositol 3′-kinase-dependent activation of the kinase Akt, resulting in enhanced phosphorylation and thereby inhibition of the proapoptotic molecule Bad. We show that signals triggered by CD152 act directly on activated CD28null T lymphocytes and, due to its exclusive expression as a receptor for CD80/CD86 on CD28null T cells, prevention of CD152-mediated signaling is likely a target mechanism taking place during therapy with CTLA-4Ig. Our data imply strongly that antagonistic approaches using CD152 signals for chronic immune responses might be beneficial.

Clinically relevant doses of FLT3-kinase inhibitors quizartinib and midostaurin do not impair T-cell reactivity and function.

The vast majority of acute myeloid leukemia (AML) patients harboring an FLT3-ITD mutation experience relapse within a short period of time after discontinuation of chemotherapy.[1][1] Treatment options include experimental trials using FLT3-tyrosine kinase inhibitors (TKI) or allogeneic stem cell

Tetanus Toxoid-Pulsed Monocyte Vaccination for Augmentation of Collateral Vessel Growth

Background The pathogenesis of collateral growth (arteriogenesis) has been linked to both the innate and adaptive immune systems. While therapeutic approaches for the augmentation of arteriogenesis have focused on innate immunity, exploiting both innate and adaptive immune responses has not been examined. We hypothesized that tetanus toxoid (tt) immunization of mice followed by transplantation of monocytes (Mo) exposed ex vivo to tt augments arteriogenesis after ligation of the hind limb. Methods and Results Mo were generated from nonimmunized BALB/c mice, exposed ex vivo to tt for 24 hours and intravenously injected (ttMo, 2.5×106) into the tail veins of tt‐immunized syngeneic mice whose hind limbs had been ligated 24 hours prior to transplantation. Laser Doppler perfusion imaging was applied, and a perfusion index (PI) was calculated (ratio ligated/unligated). Twenty‐one days after ligation, the arteriogenesis of untreated BALB/c mice was limited (PI=0.49±0.09). Hind limb function was impaired in 80% of animals. Injection of non‐engineered Mo insignificantly increased the PI to 0.56±0.07. However, ttMo transplantation resulted in a strong increase of the PI to 0.82±0.08 (n=7; P<0.001), with no (0%) detectable functional impairment. ttMo injected into nonimmunized mice had no effect. The strong arteriogenic response of ttMo transplantation into immunized mice was prevented when mice had been depleted of T‐helper cells by CD4‐antibody pretreatment (PI=0.50±0.08; n=17; P<0.001), supporting the hypothesis that transplanted cells interact with recipient lymphocytes. Conclusions Transplantation of ttMo into pre‐immunized mice strongly promotes arteriogenesis. This therapeutic approach is feasible and highly attractive for the alleviation of morbidity associated with vascular occlusive disease.

The accumulation of multifunctional T cells in hypertrophied adenoids is dependent on age and related to clinical manifestation

The quality of T cell responses differs dramatically during infancy and has been implicated in the susceptibility to infections, but the underlying cellular pathways are not well known. Here, we determined the effector phenotype of T cells in adenoids of 103 children with adenoid hypertrophy that underwent adenoidectomy. We compared children with sole upper airway obstruction with those suffering from concomitant otitis media with effusion or recurrent infections of the upper airways. Multifunctionality of T cells was determined as co-production of IFNγ, IL2, IL17 and TNFα using flow cytometry. Boolean gating was performed in FlowJo and analyzed using SPICE software. Statistical analysis was done by one-way-ANOVA.

Das neonatale Immunsystem: Modulation durch regulatorische T-Zellen und CTLA-4 (CD152)

Zusammenfassung Fehlregulationen von CTLA-4 (CD152), einem auf der Oberfläche von Lymphozyten exprimiertem Glykoprotein, können zu chronischen Entzündungsreaktionen führen. Aufgrund neuer Erkenntnisse wird deutlich, dass CTLA-4 die Effektorfunktionen von T-Lymphozyten abschaltet und damit die Effektorphase von T-Lymphozyten beendet. Interessanterweise sind die CTLA-4 exprimierenden T-Lymphozyten resistent gegen Apoptose (programmierter Zelltod) und migrieren verstärkt in Lymphknoten und Gewebe. Weitere Studien zeigen, dass regulatorische T-Zellen, die unerwünschte Immunantworten abschalten, in vivo nur inhibieren können, wenn Sie über ein intaktes CTLA-4-Gen verfügen. Darüber hinaus hat sich bestätigt, dass CTLA-4 nicht nur-wie angenommen-auf T-Lymphozyten exprimiert wird, sondern auch auf B-Lymphozyten. So zeigen Mäuse mit genetischer Inaktivierung von CTLA-4 in B-Lymphozyten eine verstärkte Produktion von IgM nach Immunisierung. Interessanterweise exprimieren insbesondere B-Lymphozyten und T-Lymphozyten von Neugeborenen und Kleinkindern stark CTLA-4, was auf eine zentrale immunregulatorische Rolle bei frühkindlichen Immunantworten hindeutet. Moleküle, die wie CTLA-4 die Differenzierung von Lymphozyten regulieren, könnten einen neuen Ansatzpunkt bieten, um bereits im Kindesalter die Weichen für ein vor Autoimmunität und Allergie geschütztes Immunsystem zu stellen. Abstract The dysregulation of CTLA-4 (CD152), a glycoprotein expressed on the surface of lymphocytes, may lead to chronic inflammation. Based on the recent scientific findings, it has become clear that CTLA-4 inhibits the effector function and, thus, shuts down the effector phase of the T-lymphocytes. Interestingly, the CTLA-4-expressing cells become resistant to apoptosis (programmed cell death) and increasingly migrate to the lymph nodes and tissues. Studies have shown that regulatory T cells (Tregs), which switch off unwanted immune responses can inhibit in vivo only if they express an intact CTLA-4 gene. Moreover, it was confirmed that CTLA-4 is not only expressed on the T-lymphocytes but also on the B-lymphocytes. The mice with genetic inactivation of CTLA-4 in the B-lymphocytes show an increased production of the IgM antibodies after immunization. Interestingly, in particular, the B- and T-lymphocytes from the newborns and infants show a strongly increased CTLA-4 expression, suggesting a key immunoregulatory role in the neonatal immune responses. The molecules such as CTLA-4, which regulate the differentiation of the lymphocytes, could provide therapeutic targets during the early childhood to set the course for protection against autoimmunity and allergy.