The nonpolymorphic soluble HLA-G1 (sHLA-G1) isoform has been reported to be secreted by trophoblast cells at the materno-fetal interface, suggesting that it may act as immunomodulator during pregnancy. In this paper, we report that affinity-purified beta2-microglobulin-associated sHLA-G1 triggered apoptosis in activated, but not resting CD8+ peripheral blood cells. We demonstrate by Western blotting that sHLA-G1 enhanced CD95 ligand expression in activated CD8+ cells. Cytotoxicity was inhibited by preincubation of the cells with a CD95 antagonist mAb (ZB4) or a soluble recombinant CD95-Fc, indicating that apoptosis is mediated through the CD95/CD95 ligand pathway. Finally, we show that such sHLA-G1-induced apoptosis depends on the interaction with CD8 molecules, with cell death being blocked by various CD8 mAbs.

No abstract

Highlights d IPH5201 and IPH5301 block cell-borne and soluble CD39 and CD73, respectively d IPH5201 maintains immunogenic extracellular ATP d When used in combination with chemotherapy, IPH5201 promotes antitumor immunity d Targeting CD39 and CD73 synergistically promotes cancer patient T cell activation

Herein we describe the molecular characterization of the human leukocyte activation antigen CD100 and identify it as the first semaphorin, to our knowledge, in the immune system. Semaphorins have recently been described as neuronal chemorepellants that direct pioneering neurons during nervous system development. In this study we demonstrate that CD100 induces B cells to aggregate and improves their viability in vitro. We show that CD100 modifies CD40-CD40L B-cell signaling by augmenting B-cell aggregation and survival and down-regulating CD23 expression. Thus, these results suggest that semaphorins as exemplified by CD100 also play a functional role in the immune system.

Singapore IL-10-producing regulatory B cells have been identified in mice and shown to downregulate inflammation, making them potentially important for maintenance of tolerance. In this study, we isolated B cells from human blood and spleen, and showed that after a short period of ex vivo stimulation a number of these cells produced IL-10. The IL-10-producing B cells did not fall within a single clearly defined subpopulation, but were enriched in both the memory (CD27 CD25À T-cell proliferation in vitro by a partially IL-10-dependent mechanism.These findings imply that manipulating IL-10 production by human B cells could be a useful therapeutic strategy for modulating immune responses in humans. 2686 Frontline Results and discussionIdentification of IL-10-producing B cells in human blood and spleen B cells in mice express IL-10 following stimulation with LPS or CpG, which bind TLR4 and TLR9, respectively [10,15]. Human B cells do not express TLR4 but do express TLR9. CpG-B 2006 is a ligand for human B-cell TLR-9 that induces IL-10 production [16][17][18]. We tested the ability of human B cells to produce IL-10 after a short stimulation, which decreased the likelihood of selective B-cell subset's expansion or surface molecule changes, which would not reflect the physiological condition. After stimulating purified B cells with CpG-B, PMA and ionomycin for 5 h, the frequency of IL-10-producing B cells from blood was 1.8% (n 5 10) and from spleen 1.1% (n 5 3) (po10 À6 and po0.05, compared with isotype control, respectively) (Fig. 1A). The combination of CpG-B with PMA and ionomycin was the most potent stimulator of IL-10 production, when compared with the other stimuli tested (anti-Ig, preplated CD40L transfected L cells or LPS, data not shown). The observed frequency of IL-10-producing B cells in blood differs from that in other studies [19][20][21][22], most likely because of the short-time stimulation system and the use of an intracellular cytokine assay instead of a cytokine secretion assay. In addition, our strategy employed a perfectly matched isotype control (same Fc fragment for the IL-10 and the control Ab from Miltenyi Biotec) to increase the specificity of IL-10 1 and IL-10 À B-cell discrimination, as opposed to the use of nonactivated B cells as a negative control which is prone to false positives. For the following phenotypic and functional experiments, only blood B cells were used due to the difficulty in obtaining sufficient spleen samples. CpG-B1anti-Ig stimulation triggers maximal IL-10 production by human blood B cells in vitroWe assessed the ability of different stimuli to induce maximal IL-10 production from purified B cells. Our cell selection method resulted in a 98% pure B-cell population (Fig. 1C, left Fig. 2B). This implies that other factors such as cell to cell contact may play a role in the effect observed, for example CD80 or CD86 interactions as suggested recently [23]. Using directly purified B cells activated with CpG-B 1anti-Ig, we also observed a high level of B-cell proliferati...

HIV-1 infection is characterized by a chronic activation of the immune system and suppressed function of T lymphocytes. Regulatory CD4+ CD25high FoxP3+CD127low T cells (Treg) play a key role in both conditions. Here, we show that HIV-1 positive patients have a significant increase of Treg-associated expression of CD39/ENTPD1, an ectoenzyme which in concert with CD73 generates adenosine. We show in vitro that the CD39/adenosine axis is involved in Treg suppression in HIV infection. Treg inhibitory effects are relieved by CD39 down modulation and are reproduced by an adenosine-agonist in accordance with a higher expression of the adenosine A2A receptor on patients' T cells. Notably, the expansion of the Treg CD39+ correlates with the level of immune activation and lower CD4+ counts in HIV-1 infected patients. Finally, in a genetic association study performed in three different cohorts, we identified a CD39 gene polymorphism that was associated with down-modulated CD39 expression and a slower progression to AIDS.

Toxic epidermal necrolysis is an extremely severe drug reaction, manifesting itself by widespread apoptosis of keratinocytes, generally considered to result from Fas/CD95-FasLigand interaction, but of unknown primary mechanism. We looked at the role of cells present in the skin blisters as probable effectors of this immune reaction. In a patient suffering from cotrimoxazole-induced toxic epidermal necrolysis, blister fluid cells were phenotyped by FACS and tested without prior restimulation for cytotoxicity on autologous and allogeneic cells in the presence of the drug. Blister fluid lymphocytes were predominantly CD8+, DR+, CLA+, CD56+ T lymphocytes, perforin positive and expressing preferentially two Vbeta chains of the T cell receptor repertoire. These lymphocytes were cytotoxic only in the presence of the drug towards autologous EBV transformed lymphocytes and towards allogeneic cells sharing HLA-Cw4. Cytotoxicity occurred in the presence of either cotrimoxazole, sulfamethoxazole, or the nitroso metabolite of sulfamethoxazole, but not with the hydroxylamine metabolite of sulfamethoxazole. The lysis was blocked by an anti-MHC class I monoclonal antibody. It was abolished by EGTA and CMA, but neither by anti-fas, brefeldin A, nor by anti-TRAIL receptor monoclonal antibodies, strongly suggesting perforin/granzyme-mediated cytotoxicity, without implication of Fas or TRAIL at this stage. This is direct evidence that T lymphocytes present within the lesions of toxic epidermal necrolysis may exhibit, without any re-stimulation, a drug-specific cytotoxicity against autologous cells. Harboring the markers of classical CTL and MHC class I restriction these lymphocytes reacted against the parent drug and one of its reactive metabolites. These results challenge several current concepts and could support new therapeutic approaches.

In wound healing and many pathologic conditions, keratinocytes become activated: they turn into migratory, hyperproliferative cells that produce and secrete extracellular matrix components and signaling polypeptides. At the same time, their cytoskeleton is also altered by the production of specific keratin proteins. These changes are orchestrated by growth factors, chemokines, and cytokines produced by keratinocytes and other cutaneous cell types. The responding intracellular signaling pathways activate transcription factors that regulate expression of keratin genes. Analysis of these processes led us to propose the existence of a keratinocyte activation cycle, in which the cells first become activated by the release of IL-1. Subsequently, they maintain the activated state by autocrine production of proinflammatory and proliferative signals. Keratins K6 and K16 are markers of the active state. Signals from the lymphocytes, in the form of Interferon-gamma, induce the expression of K17 and make keratinocytes contractile. This enables the keratinocytes to shrink the provisional fibronectin-rich basement membrane. Signals from the fibroblasts, in the form of TGF-beta, induce the expression of K5 and K14, revert the keratinocytes to the healthy basal phenotype, and thus complete the activation cycle.