ASSESSING EFFECTS OF CUCURBITURILS ON MONOCYTES AND NK-CELLS IN HEALTHY VOLUNTEERS

Nanotechnology in immunology is a prospectively developing area in fundamental and practical medicine. Cucurbiturils are macrocyclic cavitands with a definite amount of glycoluril fragments (n) that determine the size of the cavity of these compounds. Nowadays, there are six synthesized homologues: 5, 6, 7, 8, 10 and 14. They differ from each other in the portal size and the size of the cavities. They are characterized by special physicochemical and biological properties, such as biocompatibility, stability, high ability to encapsulate chemical compounds. It is known that cucurbiturils encapsulate molecules by forming guest-host complexes, which allow the substance to be released from the complex and increase the solubility of the compounds. These advantages allow using cucurbiturils as drug delivery systems. Immunomodulatory activity of cucurbiturils depends on its specific nanoscale characteristics: functional groups, shape, size, surface, solubility in various media. Each nanoparticle depending on these properties has different effects on cells. The effects of cucurbiturils can be different even for one subpopulation of cells, depending on the homologue or dosage. The interaction of innate immune cells with cucurbiturils are not yet sufficiently characterized.The aim of this study was to assess the effects of cucurbit[n]urils (n = 6, n = 7, n = 8) on innate immune cells – monocytes, NK-cells, NKT-cells.The immunological recearch included the isolation of peripheral blood mononuclear cells from healthy donors (n = 8) on the density gradient of ficoll-urografin and flow cytometry with the determination of the amount of immunocompetent cells according to the classic markers of differentiation of these cells – CD3- CD16+CD56+ for NK-cells, CD3+CD16+CD56+ for NKT-cells and CD3- CD14+ for monocytes. Monocyte activation was determined by the expression of surface HLA-DR.The cells were cultured for 72 hours with the addition of cucurbiturils CB[6], CB[7] at concentrations of 0.1 mM, 0.3 mM, 0.5 mM and CB[8] at concentration of 0.01 mM, due to its poor solubility.There were a significant decrease in the quantity of NK-cells (p < 0.01 for the test concentrations of CB[7]), an increase in the quantity of NKT-cells (p < 0.04 and p < 0.02 respectively for the concentrations of CB[6] and CB[7]). There was a tendency to increase the expression of HLA-DR on monocytes (p = 0.06 for CB[6]).Considering a variative effects of cucurbiturils, in the future it is possible to consider a possibility of using cucurbiturils as an immunomodulators, antitumor agents, in autoimmune diseases.

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17β-Estradiol Promotes Proinflammatory and Procoagulatory Phenotype of Innate Immune Cells in the Presence of Antiphospholipid Antibodies

Biomedicines ◽

10.3390/biomedicines8060162 ◽

2020 ◽

Vol 8 (6) ◽

pp. 162

Author(s):

Gayane Manukyan ◽

Anush Martirosyan ◽

Ludek Slavik ◽

Jana Ulehlova ◽

Martin Dihel ◽

...

Keyword(s):

Immune Cells ◽

Antiphospholipid Antibodies ◽

Mononuclear Cells ◽

Immune Cell ◽

Innate Immune ◽

Innate Immune Cells ◽

Cytokine Release ◽

Peripheral Blood Mononuclear ◽

17Β Estradiol ◽

Acquired Thrombophilia

Antiphospholipid syndrome (APS) is the most common cause of acquired thrombophilia and recurrent spontaneous miscarriages associated with extended persistence of antiphospholipid antibodies (aPL). How circulating aPL and high-17β-estradiol (E2) environment contribute to the pregnancy complications in APS is poorly defined. Therefore, we aimed to analyse whether E2 could be responsible for the immune cell hyperactivation in aPL- positive (lupus anticoagulant, anti-cardiolipin, anti-β2-glycoprotein) in women. For this, peripheral blood mononuclear cells (PBMCs) from 14 aPL- positive and 13 aPL- negative women were cultured in the presence or absence of E2, LPS or E2+LPS and cell immunophenotype and cytokine release were analysed. In the aPL+ group, E2 presence markedly increased the percentage of NK cells positive for CD69 (p < 0.05), monocytes positive for tissue factor (TF, CD142) (p < 0.05), and B cells expressing PD-L1 (p < 0.05), as well as the elevated production of IL-1β comparing to aPL- women (p < 0.01). Regardless of aPL positivity, E2 augmented the procoagulatory response elicited by LPS in monocytes. Our findings show the ability of E2 to promote proinflammatory and procoagulatory phenotype of innate immune cells in individuals with aPL positivity. Our data highlights the significant impact of female hormones on the activation of immune cells in the presence of aPL.

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CFTR-mediated monocyte-macrophage dysfunction revealed by cystic fibrosis proband-parent comparisons

10.1101/2021.06.30.21259182 ◽

2021 ◽

Author(s):

Xi Zhang ◽

Camille Moore ◽

Laura Harmacek ◽

Joanne Domenico ◽

Vittobai Rangaraj ◽

...

Keyword(s):

Cystic Fibrosis ◽

Immune Cells ◽

Molecular Mechanisms ◽

Mononuclear Cells ◽

Mononuclear Phagocyte ◽

Innate Immune ◽

Innate Immune Cells ◽

Peripheral Blood Mononuclear ◽

Increased Risk ◽

Biallelic Mutations

Cystic fibrosis (CF) is an inherited disorder caused by biallelic mutations of the cystic fibrosis transmembrane conductance regulator gene (CFTR). Converging lines of evidence suggest that CF carriers with only one defective CFTR copy are at increased risk for CF-related conditions and pulmonary infections, but the molecular mechanisms underpinning this effect remain unknown. Here, we performed transcriptomic profiling of peripheral blood mononuclear cells (PBMCs) of CF child-parent trios (proband, father, and mother) and healthy control PBMCs or THP-1 cells incubated with the plasma of these subjects. Transcriptomic analyses revealed suppression of cytokine-enriched immune-related genes (IL-1, CXCL8, CREM) implicating lipopolysaccharide tolerance in innate immune cells (monocytes) of CF probands and their parents and in the control innate immune cells incubated with proband or parent plasma. These data suggest that not only a homozygous but also a heterozygous CFTR mutation can modulate the immune/inflammatory system. This conclusion is further supported by the findings of lower numbers of circulating monocytes in CF probands and their parents compared to healthy controls, the abundance of mononuclear phagocyte subsets (macrophages, monocytes, and activated dendritic cells) which correlated with Pseudomonas aeruginosa infection, lung disease severity, and CF progression in the probands. This study provides insight into demonstrated CFTR-related innate immune dysfunction in individuals with CF and carriers of a CFTR mutation that may serve as a target for personalized therapy.

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Adaptive and Innate Immune Cells in Fetal Human Cytomegalovirus-Infected Brains

Microorganisms ◽

10.3390/microorganisms8020176 ◽

2020 ◽

Vol 8 (2) ◽

pp. 176 ◽

Cited By ~ 1

Author(s):

Yann Sellier ◽

Florence Marliot ◽

Bettina Bessières ◽

Julien Stirnemann ◽

Ferechte Encha-Razavi ◽

...

Keyword(s):

T Cells ◽

Nk Cells ◽

Immune Cells ◽

Plasma Cells ◽

Ex Vivo ◽

Fetal Brain ◽

Brain Lesions ◽

Innate Immune ◽

Innate Immune Cells ◽

Infected Cells

Background: The understanding of the pathogenesis of cytomegalovirus (CMV)-induced fetal brain lesions is limited. We aimed to quantify adaptive and innate immune cells and CMV-infected cells in fetal brains with various degrees of brain damage. Methods: In total, 26 archived embedded fetal brains were studied, of which 21 were CMV-infected and classified in severely affected (n = 13) and moderately affected (n = 8), and 5 were uninfected controls. The respective magnitude of infected cells, immune cells (CD8+, B cells, plasma cells, NK cells, and macrophages), and expression of immune checkpoint receptors (PD-1/PD-L1 and LAG-3) were measured by immunochemistry and quantified by quantitative imaging analysis. Results: Quantities of CD8+, plasma cells, NK cells, macrophages, and HCMV+ cells and expression of PD-1/PD-L1 and LAG-3 were significantly higher in severely affected than in moderately affected brains (all p values < 0.05). A strong link between higher number of stained cells for HCMV/CD8 and PD-1 and severity of brain lesions was found by component analysis. Conclusions: The higher expression of CD8, PD-1, and LAG-3 in severely affected brains could reflect immune exhaustion of cerebral T cells. These exhausted T cells could be ineffective in controlling viral multiplication itself, leading to more severe brain lesions. The study of the functionality of brain leucocytes ex vivo is needed to confirm this hypothesis.

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Natural Killer Dendritic Cells Enhance Immune Responses Elicited byα-Galactosylceramide-Stimulated Natural Killer T Cells

BioMed Research International ◽

10.1155/2013/460706 ◽

2013 ◽

Vol 2013 ◽

pp. 1-18 ◽

Cited By ~ 3

Author(s):

Sung Won Lee ◽

Hyun Jung Park ◽

Nayoung Kim ◽

Seokmann Hong

Keyword(s):

T Cells ◽

Dendritic Cells ◽

Immune Responses ◽

Natural Killer ◽

Immune Cells ◽

Tumor Antigens ◽

Nkt Cells ◽

Innate Immune ◽

Innate Immune Cells ◽

Natural Killer T

Natural killer dendritic cells (NKDCs) possess potent anti-tumor activity, but the cellular effect of NKDC interactions with other innate immune cells is unclear. In this study, we demonstrate that the interaction of NKDCs and natural killer T (NKT) cells is required for the anti-tumor immune responses that are elicited byα-galactosylceramide (α-GC) in mice. The rapid and strong expression of interferon-γby NKDCs afterα-GC stimulation was dependent on NKT cells. Various NK and DC molecular markers and cytotoxic molecules were up-regulated followingα-GC administration. This up-regulation could improve NKDC presentation of tumor antigens and increase cytotoxicity against tumor cells. NKDCs were required for the stimulation of DCs, NK cells, and NKT cells. The strong anti-tumor immune responses elicited byα-GC may be due to the down-regulation of regulatory T cells. Furthermore, the depletion of NKDCs dampened the tumor clearance mediated byα-GC-stimulated NKT cellsin vivo. Taken together, these results indicate that complex interactions of innate immune cells might be required to achieve optimal anti-tumor immune responses during the early stages of tumorigenesis.

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TLR agonists enhance responsiveness of inflammatory innate immune cells in HLA-B*57-positive HIV patients

Journal of Molecular Medicine ◽

10.1007/s00109-020-01996-7 ◽

2020 ◽

Vol 99 (1) ◽

pp. 147-158

Author(s):

L. Dold ◽

L. Zimmer ◽

C. Schwarze-Zander ◽

C. Boesecke ◽

R. Mohr ◽

...

Keyword(s):

Hiv Infection ◽

Nk Cells ◽

Immune Cells ◽

Nk Cell ◽

Innate Immune ◽

Innate Immune Cells ◽

Double Positive ◽

Hiv Patients ◽

Ifn Gamma ◽

Tlr Agonists

Abstract HLA-B*57 affects the course of HIV infection. Under antiretroviral therapy, its effects cannot be explained by outstandingly efficient T cell responses alone but may also involve cells of innate immunity. Studying in vitro stimulation with Pam3CSK4, E. coli LPS-B5 and CpG-ODN-2216, we observed greater induction of IL-6/IL-1beta double-positive CD14+CD16++ monocytes as well as IFN-gamma-positive cytotoxic CD56highCD16neg NK cells in HLA-B*57- versus HLA-B*44-positive HIV patients, while TNF-alpha induction remained unchanged. Differences were not seen in the other monocyte and NK cell subsets or in HLA-matched healthy controls. Our findings show that, in virally suppressed HIV infection, HLA-B*57 is associated with enhanced responsiveness of inflammatory innate immune cells to TLR ligands, possibly contributing to increased vulnerability in sepsis. Key messages • HLA-B*57 is a host factor affecting clinical outcomes of HIV infection. • HLA-B*57 modifies inflammatory subsets of NK cells and monocytes in HIV infection. • In HLA-B*57-positive HIV patients TLR agonists induce enhanced IL-6/IL-1beta in monocytes. • NK cells from HLA-B*57 HIV patients release more IFN-gamma upon TLR costimulation. • HLA-B*57 is linked to enhanced inflammatory responsiveness to TLR ligands.

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The role of IL-17 producing NK cells and other innate immune cells in poly(I:C) triggered exacerbation of experimental asthma

10.1183/13993003.congress-2016.pa1102 ◽

2016 ◽

Cited By ~ 1

Author(s):

Lars Lunding ◽

Christina Vock ◽

Sina Webering ◽

Jochen Behrends ◽

Christoph Hölscher ◽

...

Keyword(s):

Nk Cells ◽

Immune Cells ◽

Innate Immune ◽

Poly I:C ◽

Innate Immune Cells

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Function of ex vivo stimulated Natural killer T cells from patients with chronic HIV-1 infection

10.1101/831834 ◽

2019 ◽

Author(s):

Prakash Thapa ◽

Pramod Nehete ◽

Hong He ◽

Bharti Nehete ◽

Stephanie J. Buchl ◽

...

Keyword(s):

Natural Killer ◽

Peripheral Blood Mononuclear Cells ◽

Immune Cells ◽

Peripheral Blood ◽

Mononuclear Cells ◽

Ex Vivo ◽

Nkt Cells ◽

Peripheral Blood Mononuclear ◽

Blood Mononuclear Cells ◽

Hiv 1

AbstractNatural killer T (NKT) cells are innate immune cells that are responsible for the first line of antiviral defense, through crosstalk with downstream antigen-presenting cells, natural killer cells, and adaptive immune cells. Previous studies have indicated that NKT cell function is severely impaired in patients with chronic HIV-1 infection. It was reported that alpha-galactosylceramide, a potent agonist antigen for NKT cells, failed to trigger the expansion of NKT cells, or the production of anti-viral cytokines by NKT cells from HIV-1 infected patients in an in vitro assay, in which peripheral blood mononuclear cells (PBMCs) were cultured in the presence of alpha-galactosylceramide. In this study, we stimulated banked peripheral blood mononuclear cells from HIV-1-infected patients with dendritic cells (DC) generated ex vivo and loaded with alpha-galactosylceramide. The results showed that NKT cells were expanded in HIV infected subjects except in patients with advanced AIDS. Expanded NKT cells were capable of producing antiviral cytokines. Our results indicate that NKT cells in HIV infected individuals are potential targets for therapeutic intervention.

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Understanding the Role of Innate Immune Cells and Identifying Genes in Breast Cancer Microenvironment

Cancers ◽

10.3390/cancers12082226 ◽

2020 ◽

Vol 12 (8) ◽

pp. 2226

Author(s):

Israa Shihab ◽

Bariaa A. Khalil ◽

Noha Mousaad Elemam ◽

Ibrahim Y. Hachim ◽

Mahmood Yaseen Hachim ◽

...

Keyword(s):

Breast Cancer ◽

Immune System ◽

Tumor Microenvironment ◽

Nk Cells ◽

Immune Cells ◽

Innate Immune System ◽

Adaptive Immune System ◽

Innate Immune ◽

Innate Immune Cells ◽

Cancer Microenvironment

The innate immune system is the first line of defense against invading pathogens and has a major role in clearing transformed cells, besides its essential role in activating the adaptive immune system. Macrophages, dendritic cells, NK cells, and granulocytes are part of the innate immune system that accumulate in the tumor microenvironment such as breast cancer. These cells induce inflammation in situ by secreting cytokines and chemokines that promote tumor growth and progression, in addition to orchestrating the activities of other immune cells. In breast cancer microenvironment, innate immune cells are skewed towards immunosuppression that may lead to tumor evasion. However, the mechanisms by which immune cells could interact with breast cancer cells are complex and not fully understood. Therefore, the importance of the mammary tumor microenvironment in the development, growth, and progression of cancer is widely recognized. With the advances of using bioinformatics and analyzing data from gene banks, several genes involved in NK cells of breast cancer individuals have been identified. In this review, we discuss the activities of certain genes involved in the cross-talk among NK cells and breast cancer. Consequently, altering tumor immune microenvironment can make breast tumors more responsive to immunotherapy.

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Natural Killer Cells Induce the Formation of Neutrophil Extracellular Traps (NETs) in Venous Thrombosis

Blood ◽

10.1182/blood.v128.22.1424.1424 ◽

2016 ◽

Vol 128 (22) ◽

pp. 1424-1424

Author(s):

François-René Bertin ◽

Sandrine Laurance ◽

Catherine Lemarie ◽

Mark Blostein

Keyword(s):

Venous Thrombosis ◽

Nk Cells ◽

Natural Killer ◽

Immune Cells ◽

Neutrophil Extracellular Traps ◽

Innate Immune ◽

Innate Immune Cells ◽

Flow Restriction ◽

Extracellular Traps

Abstract Thrombosis is considered to be a pathological deviation of physiologic hemostasis involving similar mechanisms. Interestingly, recent work demonstrates that innate immune cells promote venous thrombosis. Innate immune cells were shown to collaborate to induce the activation of the coagulation cascade and platelets. In particular, neutrophils contribute to venous thrombosis through the release of neutrophil extracellular traps (NETs). However, the mechanism triggering the formation of NETs during venous thrombosis remain unknown. Of interest, a study showed that IFNγ induced the formation of NETs. Thus, we investigated the role of IFNγ-producing cells in the development of thrombosis. We used mice lacking IFNγ, Tbet (the transcription factor regulating the expression of IFNγ) or wild type mice. Venous thrombosis was induced using the flow restriction model in the inferior vena cava , as has been previously published. In Tbet-/-, IFNγ-/- and WT mice, we show that the absence of Tbet or IFNγ decreases the formation of thrombi after venous thrombosis induction, suggesting that the Tbet+/IFNγ producing cells are required for the early development of venous thrombosis. Comparing the composition of the thrombi from Tbet-/-, IFNγ-/- and WT mice, we show that, in all mice, neutrophils are the main cellular component of thrombi followed by monocytes; however, the number of neutrophil extracellular traps (NETs) formed during thrombosis is significantly lower in Tbet-/- and IFNγ-/- mice. Furthermore, NET formation is also decreased in WT mice specifically depleted of IFNγ and increases in Tbet-/- and IFNγ-/- mice injected with recombinant IFNγ. In vitro, we show that stimulation of WT murine neutrophils with recombinant IFNγ triggers the formation of NETs demonstrating that Tbet and IFNγ are crucial for NET formation by neutrophils. Natural killer (NK) cells are the main producers of IFNγ . Thus, we investigated the role of NK cells in venous thrombosis induced by flow restriction. NK cells were specifically depleted with an antibody during the development of venous thrombosis. The absence of NK cells results in smaller thrombi suggesting that NK cells are required for early thrombus development. Additionally, depletion in NK cells results in decreased in-situ IFNγ production and decreased NET formation. To directly link NK cells to the formation of NETs, WT neutrophils were co-cultured with Tbet-/- and IFNγ-/- NK cells. We show that WT neutrophils release less NETs when cultured with Tbet-/- and IFNγ-/- NK cells as compared to WT NK cells. These data suggest that NK cells trigger the formation of NETs by neutrophils through the production of IFNγ. Hence, we demonstrate that, in a partial flow restriction model of venous thrombosis, Tbet and IFNγ are crucial for thrombus development by promoting the formation of NETs by neutrophils and that NK cells are key effector cells in this process. Disclosures Blostein: boehringer-ingelheim: Research Funding.

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Iberdomide (CC-220) Has Synergistic Anti-Tumor and Immunostimulatory Activity Against Multiple Myeloma in Combination with Both Bortezomib and Dexamethasone, or in Combination with Daratumumab in Vitro

Blood ◽

10.1182/blood-2018-99-113383 ◽

2018 ◽

Vol 132 (Supplement 1) ◽

pp. 1935-1935 ◽

Cited By ~ 7

Author(s):

Michael Amatangelo ◽

Chad C. Bjorklund ◽

Jian Kang ◽

Ann Polonskaia ◽

Sridevi Viswanatha ◽

...

Keyword(s):

Multiple Myeloma ◽

Nk Cells ◽

Cell Lines ◽

Immune Cells ◽

Antiproliferative Activity ◽

Mononuclear Cells ◽

Interleukin 2 ◽

Immunomodulatory Activity ◽

Employment Equity ◽

Equity Ownership

Abstract Despite significant progress in the treatment of multiple myeloma (MM), the disease remains incurable. Multiple targeted and biologics-based therapies, including immunomodulatory agent IMiD® compounds (lenalidomide and pomalidomide), proteasome inhibitors (bortezomib and carfilzomib), and monoclonal antibodies (daratumumab and elotuzumab) have shown impressive activity in treating advanced MM. Moreover, triplet regimens combining these agents have consistently proven to be more efficacious than doublets in heavily pretreated patients with limited additional toxic effects. Iberdomide (CC-220) is a novel compound being investigated in a phase I/II study (clinicaltrials.gov NCT02773030) for treatment of lenalidomide- and pomalidomide-relapsed/refractory MM (RRMM) in combination with dexamethasone. Preclinical studies of iberdomide have shown that it more potently binds to cereblon than other cereblon-binding compounds, is more efficient at degrading Aiolos and Ikaros, and has enhanced immunomodulatory activity, inducing greater interleukin-2 secretion and granzyme-b degranulation in immune cells (Matyskiela et al and Bjorklund et al, submitted abstract). Clinical studies of bortezomib and daratumumab in combination with other cereblon-binding agents have demonstrated high tolerability with notable efficacy in the RRMM setting; however, these combinations with iberdomide have not been investigated. Here we show in MM cell lines that iberdomide induces deep Aiolos and Ikaros degradation in the presence of bortezomib at clinically relevant concentrations as determined in healthy volunteers. Furthermore, iberdomide treatment in combination with bortezomib produced synergistic antiproliferative activity and deeper induction of apoptosis than combinations of other clinically approved cereblon-binding compounds with bortezomib across multiple cell lines. In addition, adding dexamethasone resulted in further synergistic antiproliferative activity. In combination with daratumumab, iberdomide also had synergistic anti-MM activity in Complement-Dependent Cytotoxicity (CDC) assays. In co-culture systems using myeloma and immune cells, iberdomide significantly increased the antibody-dependent cellular cytotoxic activity of daratumumab. While iberdomide treatment of MM cell lines resulted in increased CD38 surface expression, combinations were more effective when peripheral blood mononuclear cells (PBMCs) were pretreated, suggesting that iberdomide immunomodulatory activity is a significant contributor to the synergy observed. Interestingly, pretreatment of PBMCs with daratumumab resulted in reduced efficacy of the combination. We observed that the treatment of PBMCs with daratumumab resulted in killing of natural killer (NK) cells in the PBMC culture. In contrast, treatment of PBMCs with iberdomide resulted in proliferation of NK cells, possibly helping to rescue the antagonistic effect of daratumumab on NK cell-mediated antibody‐dependent cellular cytotoxicity. Taken together, these preclinical data support further investigation of iberdomide in combination with both bortezomib/dexamethasone and daratumumab in the clinic. Disclosures Amatangelo: Celgene Corporation: Employment, Equity Ownership. Bjorklund:Celgene Corporation: Employment, Equity Ownership. Kang:Celgene Corporation: Employment, Equity Ownership. Polonskaia:Celgene Corporation: Employment, Equity Ownership. Viswanatha:Celgene Corporation: Employment, Equity Ownership. Thakurta:Celgene Corporation: Employment, Equity Ownership.

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