Tumor-induced apoptosis of T lymphocytes: elucidation of intracellular apoptotic events

Blood ◽  
2000 ◽  
Vol 95 (6) ◽  
pp. 2015-2023 ◽  
Author(s):  
Brian R. Gastman ◽  
Daniel E. Johnson ◽  
Theresa L. Whiteside ◽  
Hannah Rabinowich
Keyword(s):  
T Cells ◽  
Cell Death ◽  
T Cell ◽  
T Lymphocytes ◽  
Tumor Cells ◽  
Caspase 3 ◽  
Jurkat Cells ◽  
Caspase 8 ◽  
Induced Apoptosis ◽  
Fluoromethyl Ketone

Abstract Our recent studies suggest that human squamous cell carcinoma of the head and neck (SCCHN) is capable of activating an intrinsic mechanism of programmed-cell death in interacting lymphocytes in situ and in vitro. The current study used Jurkat T-cell line as a model to investigate intracellular apoptotic events in T cells interacting with SCCHN. Apoptosis induced in T lymphocytes by tumor cells was in part Fas-mediated, since it was partially, but significantly, inhibited in the presence of anti-Fas ligand Ab or in Fas-resistant Jurkat cells. The synthetic caspase inhibitors, N-benzyloxycarbonyl-Val-Ala-Asp-fluoromethyl ketone (Z-VAD-FMK) and N-benzyloxycarbonyl-Asp-glu-Val-Asp-fluoromethyl ketone (Z-DEVD-FMK), effectively blocked apoptosis of Jurkat cells co-incubated with SCCHN cell lines, suggesting the involvement of caspases in tumor-induced apoptosis of lymphocytes. Overexpression of CrmA, an inhibitor of caspase-1 and caspase-8, partially inhibited tumor-induced T-cell death. Caspase-8 and caspase-3 were identified as effector molecules in the execution of tumor-induced T-cell death, since the proform enzymes were processed into active subunits during co-incubation of T cells with tumor cells. Furthermore, co-incubation with tumor cells resulted in cleavage of poly(ADP-ribose) polymerase (PARP), a common caspase-3 substrate, and in cleavage of TcR-ζ chain, shown by us to be a T-cell specific caspase-3 substrate. Overexpression of Bcl-2 did not provide protection of T cells from SCCHN-induced DNA degradation. Instead, the Bcl-2 protein was cleaved in the target T cells during their co-incubation with tumor cells. These findings demonstrate that tumor cells can trigger in T lymphocytes caspase-dependent apoptotic cascades, which are not effectively protected by Bcl-2.

SMAC-Mimetic BV-6 Sensitizes Therapeutic Agents-Induced Apoptosis In AML Cells

Blood ◽  
2010 ◽  
Vol 116 (21) ◽  
pp. 2177-2177
Author(s):  
Duncan H Mak ◽  
Christa Manton ◽  
Michael Andreeff ◽  
Bing Z Carter
Keyword(s):  
Cell Death ◽  
Vector Control ◽  
Caspase 3 ◽  
Jurkat Cells ◽  
Leukemic Cells ◽  
Caspase 8 ◽  
Caspase 9 ◽  
Smac Mimetic ◽  
P53 Activation ◽  
Induced Apoptosis

Abstract Abstract 2177 The antiapoptotic function of the inhibitors of apoptosis family of proteins (IAPs) is antagonized by mitochondria-released SMAC protein. The IAP-member XIAP suppresses apoptosis by directly binding and inhibiting caspase-9 and caspase-3, while cIAP1, a component of the cytoplasmic signaling complex containing TNF receptor associated factors, suppresses apoptosis via the caspase-8-mediated pathway. BV-6 (Genentech) is a bivalent SMAC-mimetic and has been shown to promote cell death by inducing cIAP autoubiquitination, NF-κB activation, and TNFα-dependent apoptosis. We examined its effect on leukemic cells and found that BV-6 only moderately induced apoptosis. The EC50 was found to be 15.3±5.1 μM at 48 hours in OCI-AML3 cells which are relatively sensitive. We then determined whether BV-6 sensitizes leukemic cells to the HDM2-inhibitor nutlin-3a and to Ara-C. p53 modulates the expression and activity of Bcl-2 family proteins and promotes the mitochondrial-mediated apoptosis. We showed previously that activation of p53 by nutlin-3a sensitizes AML cells to XIAP inhibition induced-death in part by promoting the release of SMAC from mitochondrion (Carter BZ et al., Blood 2010). We treated OCI-AML3 cells with BV-6, nutlin-3a or Ara-C, and BV-6+nutlin-3a or BV-6+Ara-C and found that the combination of BV-6 and nutlin-3a or BV-6 and Ara-C synergistically induced cell death in OCI-AML3 cells with a combination index (CI) of 0.27±0.11 and 0.22±0.05 (48 hours), respectively. To demonstrate that p53 activation is essential for the synergism of BV-6+nutlin-3a combination, we treated OCI-AML3 vector control and p53 knockdown cells with these two agents and found that the combination synergistically promoted cell death in the vector control (CI=0.47±0.15) but not in the p53 knockdown cells, as expected, while BV6+Ara-C was synergistic in both vector control and p53 knockdown cells (CI=0.15±0.03 and 0.08±0.03, respectively, 48 hours). BV-6 induced activation of caspase-8, caspase-9, and caspase-3 and decreased XIAP levels, but did not cause rapid cIAP1 degradation, as reported by others. To assess the contribution of death receptor-mediated apoptosis in BV-6-induced cell death, we treated Jurkat and caspase-8 mutated Jurkat cells (JurkatI9.2) with BV-6 and found that BV-6 induced cell death and significantly potentiated TRAIL-induced apoptosis in Jurkat cells (CI=0.14±0.08, 48 hours). Caspase-8 mutated JurkatI9.2 cells were significantly less sensitive to BV-6 than Jurkat cells and as expected, JurkatI9.2 was completely resistant to TRAIL. Collectively, we showed that the bivalent SMAC-mimetic BV-6 potentiates p53 activation-, chemotherapy-, and TRAIL-induced cell death, but has only minimal activity by itself in leukemic cells. SMAC-mimetics could be useful in enhancing the efficacy of different classes of therapeutic agents used in AML therapy. Disclosures: No relevant conflicts of interest to declare.

scholarly journals In Vitro Effects of Ciprofloxacin and Roxithromycin on Apoptosis of Jurkat T Lymphocytes

2003 ◽  
Vol 47 (3) ◽  
pp. 1161-1164 ◽  
Author(s):  
Yong-Taek Jun ◽  
Hee-Jung Kim ◽  
Min-Jin Song ◽  
Ji-Hyang Lim ◽  
Dong-Gun Lee ◽  
...  
Keyword(s):  
T Cells ◽  
Cell Death ◽  
Caspase 3 ◽  
Caspase 8 ◽  
Jurkat T Cells ◽  
Concentration Dependency ◽  
Enhanced Expression ◽  
In Vitro Effects ◽  
Induced Apoptosis

ABSTRACT Ciprofloxacin (CPFX) and roxithromycin (RXM) induced apoptosis of activated Jurkat T cells in vitro. CPFX showed concentration-dependent acceleration of apoptosis of activated Jurkat T cells by enhancing the expression of FasL and activities of caspase-3 and -8. RXM accelerated cell death, enhanced expression of FasL and caspase-3 but not caspase-8, and did not show the concentration dependency.

scholarly journals Butyric Acid-Induced T-Cell Apoptosis Is Mediated by Caspase-8 and -9 Activation in a Fas-Independent Manner

2001 ◽  
Vol 8 (2) ◽  
pp. 325-332 ◽  
Author(s):  
Tomoko Kurita-Ochiai ◽  
Kuniyasu Ochiai ◽  
Kazuo Fukushima
Keyword(s):  
T Cells ◽  
T Cell ◽  
Butyric Acid ◽  
Cell Apoptosis ◽  
Acid Treatment ◽  
Jurkat Cells ◽  
Receptor Interaction ◽  
Caspase 8 ◽  
T Cell Apoptosis ◽  
Induced Apoptosis

ABSTRACT Our previous study demonstrated that butyric acid, an extracellular metabolite of periodontopathic bacteria, induced apoptosis in murine thymocytes, splenic T cells, and human Jurkat cells. In this study, we examined whether CD95 ligand-receptor interaction is involved in butyric acid-induced T-cell apoptosis. Flow cytometry analysis indicated that expression of Fas in Jurkat and T cells from peripheral blood mononuclear cells was not affected by butyric acid treatment. Furthermore, the expression of Fas and FasL protein in Western blotting was not affected by butyric acid treatment. Coincubation with blocking anti-Fas antibodies prevented Fas-induced apoptosis but not butyric acid-induced apoptosis. Anti-FasL antibodies also did not prevent butyric acid-induced apoptosis at any dose examined. Although cytotoxic anti-Fas antibody affected butyric acid-induced apoptosis, a synergistic effect was not seen. Time-dependent activation of caspase-8 and -9 was recognized in butyric acid- as well as Fas-mediated apoptosis. IETD-CHO and LEHD-CHO, specific inhibitors of caspase-8 and -9, respectively, completely blocked Fas-mediated apoptosis and partially prevented butyric acid-induced apoptosis. These results suggest that the Fas-FasL interaction is not involved in butyric acid-induced apoptosis and that caspase-8 and -9-dependent apoptosis plays an important role in butyric acid-induced apoptosis, as well as Fas-induced apoptosis.

Functional CD95 ligand and CD95 death-inducing signaling complex in activation-induced cell death and doxorubicin-induced apoptosis in leukemic T cells

Blood ◽  
2000 ◽  
Vol 95 (1) ◽  
pp. 301-308 ◽  
Author(s):  
Simone Fulda ◽  
Gudrun Strauss ◽  
Eric Meyer ◽  
Klaus-Michael Debatin
Keyword(s):  
T Cells ◽  
Cell Death ◽  
Leukemia Cell Line ◽  
Receptor Interaction ◽  
Caspase 8 ◽  
Drug Induced ◽  
Signaling Complex ◽  
Cd95 Ligand ◽  
Activation Induced Cell Death ◽  
Induced Apoptosis

Abstract Activation-induced cell death (AICD) in T cells is mediated by CD95 ligand (CD95L)/receptor interaction, which has also been implicated in apoptosis induction by some anticancer agents. In this article we show that both anti-CD3-triggering (AICD) and doxorubicin treatment led to the production of a functionally active CD95L in the CD3+/T-cell receptor-positive (TCR+) T leukemia cell line H9. CD95L-expressing H9 cells killed CD95-sensitive J16 or CEM target cells, but not CD95-resistant CEM or J16 cells overexpressing dominant negative FADD (J16/FADD-DN). By immunoprecipitation, CD95L was physically bound to CD95, suggesting that AICD and doxorubicin-induced apoptosis involve CD95L-mediated CD95 aggregation, thereby triggering the CD95 death pathway. CD95 aggregation was associated with the recruitment of FADD and caspase-8 to the CD95 receptor to form the CD95 death-inducing signaling complex (DISC), resulting in caspase-8 activation and cleavage of the effector caspase-3 and PARP. Blocking of the CD95L/receptor interaction by antagonistic antibodies to CD95 or to CD95L also blocked AICD and inhibited the early phase of doxorubicin-induced apoptosis, though cell death induced by doxorubicin eventually proceeded in a CD95-independent manner. These findings may explain some conflicting data on the role of death receptor systems in drug-induced apoptosis. Thus, in cells with an inducible CD95 receptor/ligand system, drug-induced apoptosis may be mediated by CD95L-initiated DISC formation and activation of downstream effector programs similar to AICD in T cells. (Blood. 2000;95:301-308)

scholarly journals The Caspase 8 Inhibitor c-FLIPL Modulates T-Cell Receptor-Induced Proliferation but Not Activation-Induced Cell Death of Lymphocytes

2002 ◽  
Vol 22 (15) ◽  
pp. 5419-5433 ◽  
Author(s):  
Susanne M. A. Lens ◽  
Takao Kataoka ◽  
Karen A. Fortner ◽  
Antoine Tinel ◽  
Isabel Ferrero ◽  
...  
Keyword(s):  
T Cells ◽  
Cell Death ◽  
T Cell ◽  
T Cell Receptor ◽  
Death Receptor ◽  
Cell Receptor ◽  
Caspase 8 ◽  
Activation Induced Cell Death

ABSTRACT The caspase 8 inhibitor c-FLIPL can act in vitro as a molecular switch between cell death and growth signals transmitted by the death receptor Fas (CD95). To elucidate its function in vivo, transgenic mice were generated that overexpress c-FLIPL in the T-cell compartment (c-FLIPL Tg mice). As anticipated, FasL-induced apoptosis was inhibited in T cells from the c-FLIPL Tg mice. In contrast, activation-induced cell death of T cells in c-FLIPL Tg mice was unaffected, suggesting that this deletion process can proceed in the absence of active caspase 8. Accordingly, c-FLIPL Tg mice differed from Fas-deficient mice by showing no accumulation of B220+ CD4− CD8− T cells. However, stimulation of T lymphocytes with suboptimal doses of anti-CD3 or antigen revealed increased proliferative responses in T cells from c-FLIPL Tg mice. Thus, a major role of c-FLIPL in vivo is the modulation of T-cell proliferation by decreasing the T-cell receptor signaling threshold.

IL15 Enhances Proliferation and Effector Function of Antigen-Specific Cytotoxic T Lymphocytes (CTLs) and Mitigates the Suppressive Action of Regulatory T Cells (Tregs).

Blood ◽  
2009 ◽  
Vol 114 (22) ◽  
pp. 4088-4088
Author(s):  
Serena Kimi Perna ◽  
Biagio De Angelis ◽  
Daria Pagliara ◽  
Lan Zhan ◽  
Cliona M Rooney ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
T Lymphocytes ◽  
Tumor Cells ◽  
Residual Tumor ◽  
Facs Analysis ◽  
Activated T Cells ◽  
Specific Ctls ◽  
Infected Cells ◽  
And Function

Abstract Abstract 4088 Poster Board III-1023 Although adoptive transfer of antigen-specific CTLs is generally safe and can be clinically effective for the treatment of several malignancies, the administration of stimulatory cytokines may be required to sustain their long-term growth and persistence in vivo. IL2, a γ-chain T-cell growth cytokine, has been used clinically, but is associated with significant toxicities. In addition, IL2 supports the expansion and function of Tregs, counterbalancing its stimulatory effects on CTLs and favoring the establishment of an immune-protected microenvironment for cancer. IL15, like IL2, is a γ-chain cytokine capable of sustaining the expansion and function of effector T cells. We have explored whether this cytokine also shares with IL2 an unwanted stimulatory effect on Tregs. Naturally occurring Tregs (CD4+CD25bright) were isolated from buffy coat preparations from healthy volunteers (mean of Treg recovery: 0.7% ± 0.05% of the starting population of mononuclear cells). The suppressive function of isolated Tregs was confirmed by their ability to inhibit the proliferation of activated T lymphocytes labeled with carboxyfluorescin diacetate succinimidyl ester (CFSE) using FACS analysis to measure CFSE dilution after 5-6 days of culture (activated T cell:Treg ratio 1:1). The proliferation of activated T cells in the presence of Tregs was significantly reduced (28%±5%) as compared to activated T cells cultured in the presence of control CD4+CD25– T cells (59%±5%) (p<0.05). Following addition of IL15 (2.5 ng/mL), however, proliferation of activated T cells continued even in the presence of Tregs (83%±5% plus IL15 without Tregs vs. 80%±5% plus IL15 and Tregs) (p=0.9), suggesting that this cytokine mitigates the immunosuppressive effects of Tregs. We then analyzed whether Tregs affected the anti-tumor activity of antigen-specific CTLs. We used our Epstein-Barr-Virus-(EBV)-specific CTLs as tumor model. EBV-CTLs were co-cultured with EBV-infected cells (LCLs) (CTL:LCL ratio 1:2). Residual tumor cells were enumerated by FACS analysis after 5-7 days of culture. In the absence of exogenous IL-15, EBV-CTLs failed to eliminate EBV-infected cells (residual LCLs: 37%±8%), while the addition of IL15 (2.5 ng/mL) increased the anti-tumor effect of CTLs, so that only 4%±1% tumor cells were detectable at the end of the culture. We then explored the effects of adding Treg to the cultures (Treg:CTL ratio 1:1). The percentage of tumor cells increased rather than decreased by day 5-7 when CTLs were cultured with Tregs in the absence of IL15 (residual tumor cells from 37%±8% in the absence of Tregs to 53%±9% in the presence of Tregs) (p<0.05). When IL15 was added, Treg were more limited in their ability to inhibit T effector cells, so that residual tumor cells were 4%±1% and 11%±3% % in the absence or in the presence of Tregs, respectively. To discover if IL15 has a direct effect on Tregs, we analyzed STAT5 phosphorylation after exposing Tregs to the cytokine. We found that this molecule was phosphorylated in 47%±18% of Tregs 15 minutes after exposure to IL15 (2.5 ng/mL). This effect was mediated by the specific interaction of the cytokine with its own receptor, as no phosphorylation occurred when Treg cells were pre-incubated with an IL-15Rα blocking antibody. This action on Tregs notwithstanding, IL15 stimulation did not modulate Treg inhibitory function, since these cells, even after exposure to IL15 (2.5 ng/mL) for 3-5 days, continued to significantly inhibit the proliferation of T lymphocytes activated in the absence of IL-15 (74%±17% inhibition). Hence, IL15 enhance the proliferative and anti-tumor effects of antigen-specific CTLs, and these effects are not impaired by the presence of Tregs. Administration of IL15 may therefore benefit patients receiving adoptive T cell therapies. Disclosures: No relevant conflicts of interest to declare.

Functional CD95 ligand and CD95 death-inducing signaling complex in activation-induced cell death and doxorubicin-induced apoptosis in leukemic T cells

Blood ◽  
2000 ◽  
Vol 95 (1) ◽  
pp. 301-308 ◽  
Author(s):  
Simone Fulda ◽  
Gudrun Strauss ◽  
Eric Meyer ◽  
Klaus-Michael Debatin
Keyword(s):  
T Cells ◽  
Cell Death ◽  
Leukemia Cell Line ◽  
Receptor Interaction ◽  
Caspase 8 ◽  
Drug Induced ◽  
Signaling Complex ◽  
Cd95 Ligand ◽  
Activation Induced Cell Death ◽  
Induced Apoptosis

Activation-induced cell death (AICD) in T cells is mediated by CD95 ligand (CD95L)/receptor interaction, which has also been implicated in apoptosis induction by some anticancer agents. In this article we show that both anti-CD3-triggering (AICD) and doxorubicin treatment led to the production of a functionally active CD95L in the CD3+/T-cell receptor-positive (TCR+) T leukemia cell line H9. CD95L-expressing H9 cells killed CD95-sensitive J16 or CEM target cells, but not CD95-resistant CEM or J16 cells overexpressing dominant negative FADD (J16/FADD-DN). By immunoprecipitation, CD95L was physically bound to CD95, suggesting that AICD and doxorubicin-induced apoptosis involve CD95L-mediated CD95 aggregation, thereby triggering the CD95 death pathway. CD95 aggregation was associated with the recruitment of FADD and caspase-8 to the CD95 receptor to form the CD95 death-inducing signaling complex (DISC), resulting in caspase-8 activation and cleavage of the effector caspase-3 and PARP. Blocking of the CD95L/receptor interaction by antagonistic antibodies to CD95 or to CD95L also blocked AICD and inhibited the early phase of doxorubicin-induced apoptosis, though cell death induced by doxorubicin eventually proceeded in a CD95-independent manner. These findings may explain some conflicting data on the role of death receptor systems in drug-induced apoptosis. Thus, in cells with an inducible CD95 receptor/ligand system, drug-induced apoptosis may be mediated by CD95L-initiated DISC formation and activation of downstream effector programs similar to AICD in T cells. (Blood. 2000;95:301-308)

An emerging player in the adaptive immune response: microRNA-146a is a modulator of IL-2 expression and activation-induced cell death in T lymphocytes

Blood ◽  
2010 ◽  
Vol 115 (2) ◽  
pp. 265-273 ◽  
Author(s):  
Graziella Curtale ◽  
Franca Citarella ◽  
Claudia Carissimi ◽  
Marina Goldoni ◽  
Nicoletta Carucci ◽  
...  
Keyword(s):  
Immune Response ◽  
T Cells ◽  
Cell Death ◽  
T Cell ◽  
T Lymphocytes ◽  
T Cell Activation ◽  
Cell Activation ◽  
Interleukin 2 ◽  
Activation Induced Cell Death

Abstract Activation of the T cell–mediated immune response has been associated with changes in the expression of specific microRNAs (miRNAs). However, the role of miRNAs in the development of an effective immune response is just beginning to be explored. This study focuses on the functional role of miR-146a in T lymphocyte–mediated immune response and provides interesting clues on the transcriptional regulation of miR-146a during T-cell activation. We show that miR-146a is low in human naive T cells and is abundantly expressed in human memory T cells; consistently, miR-146a is induced in human primary T lymphocytes upon T-cell receptor (TCR) stimulation. Moreover, we identified NF-kB and c-ETS binding sites as required for the induction of miR-146a transcription upon TCR engagement. Our results demonstrate that several signaling pathways, other than inflammation, are influenced by miR-146a. In particular, we provide experimental evidence that miR-146a modulates activation-induced cell death (AICD), acting as an antiapoptotic factor, and that Fas-associated death domain (FADD) is a target of miR-146a. Furthermore, miR-146a enforced expression impairs both activator protein 1 (AP-1) activity and interleukin-2 (IL-2) production induced by TCR engagement, thus suggesting a role of this miRNA in the modulation of adaptive immunity.

scholarly journals T Cell Bispecific Antibodies: An Antibody-Based Delivery System for Inducing Antitumor Immunity

2021 ◽  
Vol 14 (11) ◽  
pp. 1172
Author(s):  
Daisuke Kamakura ◽  
Ryutaro Asano ◽  
Masahiro Yasunaga
Keyword(s):  
T Cells ◽  
Cell Death ◽  
T Cell ◽  
Tumor Cell ◽  
Tumor Cells ◽  
T Cell Activation ◽  
Antitumor Immunity ◽  
Bispecific Antibodies ◽  
Tumor Site ◽  
Tumor Cell Death

As a breakthrough immunotherapy, T cell bispecific antibodies (T-BsAbs) are a promising antibody therapy for various kinds of cancer. In general, T-BsAbs have dual-binding specificity to a tumor-associated antigen and a CD3 subunit forming a complex with the TCR. This enables T-BsAbs to crosslink tumor cells and T cells, inducing T cell activation and subsequent tumor cell death. Unlike immune checkpoint inhibitors, which release the brake of the immune system, T-BsAbs serve as an accelerator of T cells by stimulating their immune response via CD3 engagement. Therefore, they can actively redirect host immunity toward tumors, including T cell recruitment from the periphery to the tumor site and immunological synapse formation between tumor cells and T cells. Although the low immunogenicity of solid tumors increases the challenge of cancer immunotherapy, T-BsAbs capable of immune redirection can greatly benefit patients with such tumors. To investigate the detailed relationship between T-BsAbs delivery and their T cell redirection activity, it is necessary to determine how T-BsAbs deliver antitumor immunity to the tumor site and bring about tumor cell death. This review article discusses T-BsAb properties, specifically their pharmacokinetics, redirection of anticancer immunity, and local mechanism of action within tumor tissues, and discuss further challenges to expediting T-BsAb development.

scholarly journals Streptococcal Pyrogenic Exotoxin A-Stimulated Monocytes Mediate Regulatory T-Cell Accumulation through PD-L1 and Kynurenine

2019 ◽  
Vol 20 (16) ◽  
pp. 3933 ◽  
Author(s):  
Katharina Giesbrecht ◽  
Sandra Förmer ◽  
Aline Sähr ◽  
Klaus Heeg ◽  
Dagmar Hildebrand
Keyword(s):  
T Cells ◽  
Cell Death ◽  
T Cell ◽  
T Lymphocytes ◽  
Mhc Class Ii ◽  
Inflammatory Cytokines ◽  
Class Ii ◽  
Cd4 T Cell ◽  
Exotoxin A ◽  
T Effector Cells

Bacterial superantigens (SAgs) are exotoxins that promote a fulminant activation of the immune system. The subsequent intense release of inflammatory cytokines often results in hypotension, shock, and organ failure with high mortality rates. In the current paradigm, the direct and simultaneous binding of SAgs with T-cell receptor (TCR)-bearing Vβ regions and conserved structures on major histocompatibility complex class II (MHC class II) on antigen-presenting cells (APCs) induces the activation of both cell types. However, by crosslinking MHC class II molecules, APCs can be activated by SAgs independently of T lymphocytes. Recently, we showed that streptococcal pyrogenic exotoxin A (SPEA) of Streptococcus pyogenes stimulates an immunogenic APC phenotype with upregulated costimulatory molecules and inflammatory cytokines. Additionally, we revealed that SPEA triggers immunosuppressive programs in monocytes that facilitate the accumulation of regulatory T cells (Tregs) in in vitro monocyte/CD4+ T-cell cocultures. Immunosuppressive factors include anti-inflammatory interleukin 10 (IL-10), co-inhibitory surface molecule programmed cell death 1 ligand 1 (PD-L1), and the inhibitory indoleamine 2,3-dioxygenase (IDO)/kynurenine effector system. In the present study, we investigated the underlying mechanism of SPEA-stimulated monocyte-mediated accumulation of Tregs. Blood-derived monocytes from healthy donors were stimulated with SPEA for 48 h (SPEA-monocytes). For the evaluation of SPEA-monocyte-mediated modulation of CD4+ T lymphocytes, SPEA was removed from the culture through extensive washing of cells before adding allogeneic CD3/CD28-activated T cells. Results: In coculture with allogeneic CD4+ T cells, SPEA-monocytes mediate apoptosis of CD4+Foxp3− lymphocytes and accumulation of CD4+Foxp3+ Tregs. PD-L1 and kynurenine are critically involved in the mediated cell death because blocking both factors diminished apoptosis and decreased the proportion of the CD25+/Foxp3+ Treg subpopulation significantly. Upregulation of PD-L1 and kynurenine as well as SPEA-monocyte-mediated effects on T cells depend on inflammatory IL-1β. Our study shows that monocytes activated by SPEA mediate apoptosis of CD4+Foxp3− T effector cells through PD-L1 and kynurenine. CD4+Foxp3+ T cells are resistant to apoptosis and accumulate in SPEA-monocyte/CD4+ T-cell coculture.

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