scholarly journals Impact of Secondary Structure of Toll-Like Receptor 9 Agonists on Interferon Alpha Induction

2008 ◽  
Vol 52 (12) ◽  
pp. 4320-4325 ◽  
Author(s):  
Dong Yu ◽  
Mallikarjuna R. Putta ◽  
Lakshmi Bhagat ◽  
Meiru Dai ◽  
Daqing Wang ◽  
...  
Keyword(s):  
Immune Responses ◽  
Interferon Alpha ◽  
Nonhuman Primates ◽  
Human Peripheral Blood ◽  
Plasmacytoid Dendritic Cell ◽  
Subcutaneous Administration ◽  
Melting Transition ◽  
Toll Like Receptor ◽  
Peripheral Blood Mononuclear

ABSTRACT Oligodeoxynucleotides containing a CpG motif and double- or multistranded structure-forming sequences act as agonists of Toll-like receptor 9 (TLR9) and induce high levels of interferon alpha (IFN-α) in addition to other Th1-type cytokines. In the present study, we evaluated three highly effective IFN-α-inducing agonists of TLR9 to determine the type of duplex structures formed and the agonist's ability to induce immune responses, including IFN-α induction, in human cell-based assays and in vivo in mice and nonhuman primates. Thermal melting studies showed that two of the agonists evaluated had a single melting transition with similar hyperchromicity in both heating and cooling cycles, suggesting the formation of intermolecular duplexes. A third agonist showed a biphasic melting transition in the heating cycle and a monophasic melting transition with lower hyperchromicity during the cooling cycle, suggesting the formation of both intra- and intermolecular duplexes. All three agonists induced the production of Th1-type cytokines and chemokines, including high levels of IFN-α, in human peripheral blood mononuclear cell and plasmacytoid dendritic cell cultures. Subcutaneous administration of the two intermolecular duplex-forming agonists, but not the intramolecular duplex-forming agonist, induced cytokine secretion in mice. In nonhuman primates, the two agonists that formed intermolecular duplexes induced IFN-α and IP-10 secretion. On the contrary, the agonist that formed an intramolecular duplex induced only low levels of cytokines in nonhuman primates, suggesting that this type of structure formation is less immunostimulatory in vivo than the other structure. Taken together, the present results suggest that oligonucleotide-based agonists of TLR9 that form intermolecular duplexes induce potent immune responses in vivo.

scholarly journals Neospora caninum: a new class of biopharmaceuticals in the therapeutic arsenal against cancer

2020 ◽  
Vol 8 (2) ◽  
pp. e001242
Author(s):  
Louis Lantier ◽  
Agathe Poupée-Beaugé ◽  
Anne di Tommaso ◽  
Céline Ducournau ◽  
Mathieu Epardaud ◽  
...  
Keyword(s):  
Immune Responses ◽  
Mononuclear Cells ◽  
Antitumor Agents ◽  
Neospora Caninum ◽  
Human Peripheral Blood ◽  
Severe Combined Immunodeficiency ◽  
Peripheral Blood Mononuclear ◽  
Ifn Γ

BackgroundMicroorganisms that can be used for their lytic activity against tumor cells as well as inducing or reactivating antitumor immune responses are a relevant part of the available immunotherapy strategies. Viruses, bacteria and even protozoa have been largely explored with success as effective human antitumor agents. To date, only one oncolytic virus—T-VEC—has been approved by the US Food and Drug Administration for use in biological cancer therapy in clinical trials. The goal of our study is to evaluate the potential of a livestock pathogen, the protozoan Neospora caninum, non-pathogenic in humans, as an effective and safe antitumorous agent.Methods/ResultsWe demonstrated that the treatment of murine thymoma EG7 by subcutaneous injection of N. caninum tachyzoites either in or remotely from the tumor strongly inhibits tumor development, and often causes their complete eradication. Analysis of immune responses showed that N. caninum had the ability to 1) lyze infected cancer cells, 2) reactivate the immunosuppressed immune cells and 3) activate the systemic immune system by generating a protective antitumor response dependent on natural killer cells, CD8-T cells and associated with a strong interferon (IFN)-γ secretion in the tumor microenvironment. Most importantly, we observed a total clearance of the injected agent in the treated animals: N. caninum exhibited strong anticancer effects without persisting in the organism of treated mice. We also established in vitro and an in vivo non-obese diabetic/severe combined immunodeficiency mouse model that N. caninum infected and induced a strong regression of human Merkel cell carcinoma. Finally, we engineered a N. caninum strain to secrete human interleukin (IL)-15, associated with the alpha-subunit of the IL-15 receptor thus strengthening the immuno-stimulatory properties of N. caninum. Indeed, this NC1-IL15hRec strain induced both proliferation of and IFN-γ secretion by human peripheral blood mononuclear cells, as well as improved efficacy in vivo in the EG7 tumor model.ConclusionThese results highlight N. caninum as a potential, extremely effective and non-toxic anticancer agent, capable of being engineered to either express at its surface or to secrete biodrugs. Our work has identified the broad clinical possibilities of using N. caninum as an oncolytic protozoan in human medicine.

scholarly journals Dialyzable Leukocyte Extracts Activate TLR-2 on Monocytes

2014 ◽  
Vol 9 (6) ◽  
pp. 1934578X1400900 ◽  
Author(s):  
Uriel García-Hernández ◽  
Frank H. Robledo-Ávila ◽  
Violeta D. Álvarez-Jiménez ◽  
Octavio Rodríguez-Cortés ◽  
Isabel Wong-Baeza ◽  
...  
Keyword(s):  
Infectious Diseases ◽  
Immune Responses ◽  
Mononuclear Cells ◽  
Specific Cell ◽  
Toll Like Receptor ◽  
Peripheral Blood Mononuclear ◽  
Tnf Α ◽  
Activated Monocytes

Dialyzable leukocyte extracts (DLE) transfer specific cell-mediated immune responses from sensitized donors to non-immune recipients. In addition, DLE have several immunomodulatory effects and are used for the treatment of several infectious and non-infectious diseases. Previous studies showed that human DLE obtained from virus-infected leukocytes and bovine DLE decrease the production of the pro-inflammatory cytokine TNF-α in response to bacterial lipopolysaccharide, in vitro and in vivo. In the present work, we inquire as to whether DLE from uninfected human leukocytes have the ability to regulate cytokine production in peripheral blood mononuclear cells (PBMC) in vitro. We observed that PBMC from healthy individuals were able to produce TNF-α, IL-12 and IL-10 after stimulation with DLE. Moreover, we identified monocytes as the main cell population that produced TNF-α after DLE stimulation. Interestingly, we found that DLE contain unidentified ligands that activate Toll-like receptor (TLR)-2. Finally, we observed that DLE directly activated monocytes through TLR-2. These results reveal a new biological activity of DLE, and suggest that part of the immunomodulatory properties of DLE could be attributed to TLR-2 activation on monocytes and to the induction of a pro-inflammatory environment that is crucial for control of infectious diseases.

scholarly journals Cholera Toxin Suppresses Interleukin (IL)-12 Production and IL-12 Receptor β1 and β2 Chain Expression

1999 ◽  
Vol 189 (3) ◽  
pp. 541-552 ◽  
Author(s):  
Michael C. Braun ◽  
Jianping He ◽  
Chang-You Wu ◽  
Brian L. Kelsall
Keyword(s):  
Immune Responses ◽  
Cholera Toxin ◽  
Transforming Growth Factor ◽  
Human Peripheral Blood ◽  
Serum Levels ◽  
Mucosal Vaccine ◽  
Dependent Manner ◽  
Peripheral Blood Mononuclear ◽  
Tgf Β1

Cholera toxin (CT) is a potent mucosal vaccine adjuvant, which has been shown to induce T helper cell type 2 (Th2) responses in systemic and mucosal tissues. We report that CT inhibits the production of interleukin (IL)-12, a major Th2 counterregulatory cytokine. IL-12 p70 production by stimulated human monocytes was inhibited by CT in a dose-dependent manner. This suppression occurred at the level of gene transcription, was maximal at low concentrations of CT, and was dependent on the A subunit of the toxin, since purified CT B subunit had minimal effect. CT also inhibited the production of IL-12 p70 by monocyte-derived dendritic cells, as well as the production of tumor necrosis factor α, but not IL-10, IL-6, or transforming growth factor (TGF)-β1, by stimulated monocytes. The effects of CT were not due to autocrine production of IL-10, TGF-β1, or prostaglandin E2. CT inhibited the production of IFN-γ by anti-CD3-stimulated human peripheral blood mononuclear cell, due in part to suppression of IL-12 production, but also to the inhibition of expression of the β1 and β2 chains of the IL-12 receptor on T cells. In vivo, mice given CT before systemic challenge with lipopolysaccharide had markedly reduced serum levels of IL-12 p40 and interferon γ. These data demonstrate two novel mechanisms by which CT can inhibit Th1 immune responses, and help explain the ability of mucosally administered CT to enhance Th2-dependent immune responses.

scholarly journals Toll-like receptor-9 stimulated plasmacytoid dendritic cell precursors suppress autoimmune neuroinflammation in a murine model of multiple sclerosis

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Hélène Letscher ◽  
Viviane A. Agbogan ◽  
Sarantis Korniotis ◽  
Pauline Gastineau ◽  
Emmanuel Tejerina ◽  
...  
Keyword(s):  
Dendritic Cell ◽  
Plasmacytoid Dendritic Cell ◽  
Toll Like Receptor ◽  
Hematopoietic Progenitors ◽  
Autoimmune Encephalomyelitis ◽  
Hematopoietic Precursor ◽  
Therapeutic Properties ◽  
Experimental Autoimmune

AbstractEarly innate education of hematopoietic progenitors within the bone marrow (BM) stably primes them for either trained immunity or instead immunoregulatory functions. We herein demonstrate that in vivo or in vitro activation within the BM via Toll-like receptor-9 generates a population of plasmacytoid dendritic cell (pDC) precursors (CpG-pre-pDCs) that, unlike pDC precursors isolated from PBS-incubated BM (PBS-pre-pDCs), are endowed with the capacity to halt progression of ongoing experimental autoimmune encephalomyelitis. CpG activation enhances the selective migration of pDC precursors to the inflamed spinal cord, induces their immediate production of TGF-β, and after migration, of enhanced levels of IL-27. CpG-pre-pDC derived TGF-β and IL-27 ensure protection at early and late phases of the disease, respectively. Spinal cords of CpG-pre-pDC-protected recipient mice display enhanced percentages of host-derived pDCs expressing TGF-β as well as an accumulation of IL-10 producing B cells and of CD11c+ CD11b+ dendritic cells. These results reveal that pDC precursors are conferred stable therapeutic properties by early innate activation within the BM. They further extend to the pDC lineage promising perspectives for cell therapy of autoimmune diseases with innate activated hematopoietic precursor cells.

scholarly journals Monocyte tissue factor induction by lipopolysaccharide (LPS): dependence on LPS-binding protein and CD14, and inhibition by a recombinant fragment of bactericidal/permeability-increasing protein

Blood ◽  
1994 ◽  
Vol 83 (9) ◽  
pp. 2516-2525 ◽  
Author(s):  
K Meszaros ◽  
S Aberle ◽  
R Dedrick ◽  
R Machovich ◽  
A Horwitz ◽  
...  
Keyword(s):  
Tissue Factor ◽  
Mononuclear Cells ◽  
Human Peripheral Blood ◽  
Binding Protein ◽  
Mononuclear Phagocytes ◽  
Factor Vii ◽  
Peripheral Blood Mononuclear ◽  
Recombinant Fragment

Abstract Mononuclear phagocytes, stimulated by bacterial lipopolysaccharide (LPS), have been implicated in the activation of coagulation in sepsis and endotoxemia. In monocytes LPS induces the synthesis of tissue factor (TF) which, assembled with factor VII, initiates the blood coagulation cascades. In this study we investigated the mechanism of LPS recognition by monocytes, and the consequent expression of TF mRNA and TF activity. We also studied the inhibition of these effects of LPS by rBPI23, a 23-kD recombinant fragment of bactericidal/permeability increasing protein, which has been shown to antagonize LPS in vitro and in vivo. Human peripheral blood mononuclear cells, or monocytes isolated by adherence, were stimulated with Escherichia coli O113 LPS at physiologically relevant concentrations (> or = 10 pg/mL). The effect of LPS was dependent on the presence of the serum protein LBP (lipopolysaccharide-binding protein), as shown by the potentiating effect of human recombinant LBP or serum. Furthermore, recognition of low amounts of LPS by monocytes was also dependent on CD14 receptors, because monoclonal antibodies against CD14 greatly reduced the LPS sensitivity of monocytes in the presence of serum or rLBP. Induction of TF activity and mRNA expression by LPS were inhibited by rBPI23. The expression of tumor necrosis factor showed qualitatively similar changes. Considering the involvement of LPS-induced TF in the potentially lethal intravascular coagulation in sepsis, inhibition of TF induction by rBPI23 may be of therapeutic benefit.

scholarly journals Essential role of IRAK-4 protein and its kinase activity in Toll-like receptor–mediated immune responses but not in TCR signaling

2007 ◽  
Vol 204 (5) ◽  
pp. 1013-1024 ◽  
Author(s):  
Tatsukata Kawagoe ◽  
Shintaro Sato ◽  
Andreas Jung ◽  
Masahiro Yamamoto ◽  
Kosuke Matsui ◽  
...  
Keyword(s):  
Immune Responses ◽  
Kinase Activity ◽  
Interleukin 1 ◽  
Cell Receptor ◽  
Nuclear Factor Κb ◽  
Toll Like Receptor ◽  
Tcr Signaling ◽  
Mitogen Activated Protein

Interleukin-1 receptor–associated kinase 4 (IRAK-4) was reported to be essential for the Toll-like receptor (TLR)– and T cell receptor (TCR)–mediated signaling leading to the activation of nuclear factor κB (NF-κB). However, the importance of kinase activity of IRAK family members is unclear. In this study, we investigated the functional role of IRAK-4 activity in vivo by generating mice carrying a knockin mutation (KK213AA) that abrogates its kinase activity. IRAK-4KN/KN mice were highly resistant to TLR-induced shock response. The cytokine production in response to TLR ligands was severely impaired in IRAK-4KN/KN as well as IRAK-4−/− macrophages. The IRAK-4 activity was essential for the activation of signaling pathways leading to mitogen-activated protein kinases. TLR-induced IRAK-4/IRAK-1–dependent and –independent pathways were involved in early induction of NF-κB–regulated genes in response to TLR ligands such as tumor necrosis factor α and IκBζ. In contrast to a previous paper (Suzuki, N., S. Suzuki, D.G. Millar, M. Unno, H. Hara, T. Calzascia, S. Yamasaki, T. Yokosuka, N.J. Chen, A.R. Elford, et al. 2006. Science. 311:1927–1932), the TCR signaling was not impaired in IRAK-4−/− and IRAK-4KN/KN mice. Thus, the kinase activity of IRAK-4 is essential for the regulation of TLR-mediated innate immune responses.

scholarly journals Characteristics of a Pathogenic Molecular Clone of an End-Stage Serum-Derived Variant of Simian Immunodeficiency Virus (SIVF359)

2001 ◽  
Vol 75 (19) ◽  
pp. 9328-9338 ◽  
Author(s):  
Lennart Holterman ◽  
Rob Dubbes ◽  
James Mullins ◽  
Gerald Learn ◽  
Henk Niphuis ◽  
...  
Keyword(s):  
Simian Immunodeficiency Virus ◽  
Peripheral Blood ◽  
Mononuclear Cells ◽  
Human Peripheral Blood ◽  
Peripheral Blood Lymphocytes ◽  
Peripheral Blood Mononuclear ◽  
Molecular Clone ◽  
Immunodeficiency Virus ◽  
End Stage

ABSTRACT End-stage simian immunodeficiency virus (SIV) isolates are suggested to be the most fit of the evolved virulent variants that precipitate the progression to AIDS. To determine if there were common characteristics of end-stage variants which emerge from accelerated cases of AIDS, a molecular clone was derived directly from serum following in vivo selection of a highly virulent SIV isolate obtained by serial end-stage passage in rhesus monkeys (Macaca mulatta). This dominant variant caused a marked cytopathic effect and replicated to very high levels in activated but not resting peripheral blood lymphocytes. Furthermore, although this clone infected but did not replicate to detectable levels in rhesus monocyte-derived macrophages, these cells were able to transmit infection to autologous T cells upon contact. Interestingly, although at low doses this end-stage variant did not use any of the known coreceptors except CCR5, it was able to infect and replicate in human peripheral blood mononuclear cells homozygous for the Δ32 deletion of CCR5, suggesting the use of a novel coreceptor. It represents the first pathogenic molecular clone of SIV derived from viral RNA in serum and provides evidence that not only the genetic but also the biological characteristics acquired by highly fit late-stage disease variants may be distinct in different hosts.

scholarly journals RNA-Seq Transcriptome Analysis Reveals Long Terminal Repeat Retrotransposon Modulation in Human Peripheral Blood Mononuclear Cells after In Vivo Lipopolysaccharide Injection

2020 ◽  
Vol 94 (19) ◽  
Author(s):  
Maria Paola Pisano ◽  
Olivier Tabone ◽  
Maxime Bodinier ◽  
Nicole Grandi ◽  
Julien Textoris ◽  
...  
Keyword(s):  
Peripheral Blood Mononuclear Cells ◽  
Peripheral Blood ◽  
Mononuclear Cells ◽  
Human Peripheral Blood ◽  
Ltr Retrotransposon ◽  
Ltr Retrotransposons ◽  
Rna Seq ◽  
Peripheral Blood Mononuclear ◽  
Blood Mononuclear Cells

ABSTRACT Human endogenous retroviruses (HERVs) and mammalian apparent long terminal repeat (LTR) retrotransposons (MaLRs) are retroviral sequences that integrated into germ line cells millions of years ago. Transcripts of these LTR retrotransposons are present in several tissues, and their expression is modulated in pathological conditions, although their function remains often far from being understood. Here, we focused on the HERV/MaLR expression and modulation in a scenario of immune system activation. We used a public data set of human peripheral blood mononuclear cells (PBMCs) RNA-Seq from 15 healthy participants to a clinical trial before and after exposure to lipopolysaccharide (LPS), for which we established an RNA-Seq workflow for the identification of expressed and modulated cellular genes and LTR retrotransposon elements. IMPORTANCE We described the HERV and MaLR transcriptome in PBMCs, finding that about 8.4% of the LTR retrotransposon loci were expressed and identifying the betaretrovirus-like HERVs as those with the highest percentage of expressed loci. We found 4,607 HERV and MaLR loci that were modulated as a result of in vivo stimulation with LPS. The HERV-H group showed the highest number of differentially expressed most intact proviruses. We characterized the HERV and MaLR loci as differentially expressed, checking their genomic context of insertion and observing a general colocalization with genes that are involved and modulated in the immune response, as a consequence of LPS stimulation. The analyses of HERV and MaLR expression and modulation show that these LTR retrotransposons are expressed in PBMCs and regulated in inflammatory settings. The similar regulation of HERVs/MaLRs and genes after LPS stimulation suggests possible interactions of LTR retrotransposons and the immune host response.

scholarly journals Chloroquine and Rapamycin Augment Interleukin-37 Expression via the LC3, ERK, and AP-1 Axis in the Presence of Lipopolysaccharides

2020 ◽  
Vol 2020 ◽  
pp. 1-14
Author(s):  
Xiaoyi Shi ◽  
Chunhui Lai ◽  
Lianyu Zhao ◽  
Mingying Zhang ◽  
Xi Liu ◽  
...  
Keyword(s):  
Mononuclear Cells ◽  
Therapeutic Potential ◽  
Inflammatory Diseases ◽  
Human Peripheral Blood ◽  
Signal Pathways ◽  
U937 Cells ◽  
Healthy Human ◽  
Peripheral Blood Mononuclear ◽  
Phosphorylated Proteins

IL-37 is a cytokine that plays critical protective roles in many metabolic inflammatory diseases, and its therapeutic potential has been confirmed by exogenous IL-37 administration. However, its regulatory mechanisms remain unclear. U937 cells were treated with autophagy-modifying reagents (3-MA, chloroquine, and rapamycin) with or without LPS stimulation. Thereafter, IL-37 expression and autophagic markers (Beclin1, P62/SQSTM1, and LC3) were determined. For regulatory signal pathways, phosphorylated proteins of NF-κB (p65 and IκBα), AP-1 (c-Fos/c-Jun), and MAPK signal pathways (Erk1/2 and p38 MAPK) were quantified, and the agonists and antagonists of MAPK and NF-κB pathways were also used. Healthy human peripheral blood mononuclear cells were treated similarly to confirm our results. Four rhesus monkeys were also administered chloroquine to evaluate IL-37 induction in vivo and its bioactivity on CD4 proliferation and activation. IL-37 was upregulated by rapamycin and chloroquine in both U937 cells and human PBMCs in the presence of LPS. IL-37 was preferentially induced in autophagic cells associated with LC3 conversion. AP-1 and p65 binding motifs could be deduced in the sequence of the IL-37 promoter. Inductive IL-37 expression was accompanied with increased phosphorylated Erk1/2 and AP-1 and could be completely abolished by an Erk1/2 inhibitor or augmented by Erk1/2 agonists. In monkeys, chloroquine increased IL-37 expression, which was inversely correlated with CD4 proliferation and phosphorylated STAT3. IL-37 levels were induced by rapamycin and chloroquine through the LC3, Erk1/2, and NF-κB/AP-1 pathways. Functional IL-37 could also be induced in vivo.

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