scholarly journals Yu-Ping-Feng Formula Exerts Antilung Cancer Effects by Remodeling the Tumor Microenvironment through Regulating Myeloid-Derived Suppressor Cells

2021 ◽  
Vol 2021 ◽  
pp. 1-16
Author(s):  
Yuli Wang ◽  
Ningyang Sun ◽  
Yingbin Luo ◽  
Zhihong Fang ◽  
Yuan Fang ◽  
...  
Keyword(s):  
Flow Cytometry ◽  
Tumor Microenvironment ◽  
Western Blotting ◽  
Suppressor Cells ◽  
T Cell Subsets ◽  
Control Group ◽  
Spleen Weight ◽  
Myeloid Derived Suppressor Cells ◽  
Rt Pcr

Yu-Ping-Feng (YPF) formula is a classical prescription used for enhancing the body’s immunity function in traditional Chinese medicine (TCM). In clinical practice, the YPF formula has been reported to exhibit antilung cancer and immunomodulatory effect. However, the relationship between them remains unclear. The present study aimed to investigate the antilung cancer effect of the YPF formula and its immune-related mechanisms. The C57BL/6 tumor-bearing mice model was established and randomly divided into the YPF group and the control group. Tumor volume, spleen weight, and survival in both groups were measured and evaluated during 28 days of consecutive intervention. Flow cytometry was used to detect the proportion of immune cell subsets. Myeloid-derived suppressor cells (MDSCs) were induced in vitro from bone marrow cells. After intervention by the YPF formula, CCK-8 and flow cytometry analyses were performed to detect proliferation and apoptosis of MDSCs. A coculture system containing T cells and MDSCs was established to further study the role of MDSCs in the regulation of T-cell subsets proportion by the YPF formula. The expressions of MDSCs-related genes and proteins were detected by RT-PCR and Western blotting. The results showed the YPF formula inhibited tumor growth, reduced spleen weight, and prolonged the survival of mice. Besides, the proportions of MDSCs subsets and Regulatory T (Treg) in the YPF group decreased, whereas those of CD4+T and CD8+T increased both in vitro and in vivo. CCK-8 and flow cytometry demonstrated that the YPF formula could inhibit proliferation and promote apoptosis of MDSCs. The coculture experiments further confirmed that MDSCs served a critical role in regulating the tumor microenvironment by the YPF formula. RT-PCR and Western blotting indicated that the levels of MDSCs’ activation and proliferation-related proteins and genes were downregulated in the YPF group. Therefore, our results demonstrated that the YPF formula could promote apoptosis and inhibit the proliferation of MDSCs. As a result, the negative regulatory effect on the positive immune cells induced by MDSCs was weakened, thus achieving the antilung cancer effect by remodeling the tumor microenvironment.

scholarly journals IMMU-28. TARGETING IMMUNOSUPPRESSIVE MYELOID DERIVED SUPPRESSOR CELLS VIA MIF/CD74 SIGNALING AXIS TO ATTENUATE GBM GROWTH

2019 ◽  
Vol 21 (Supplement_6) ◽  
pp. vi125-vi125
Author(s):  
Tyler Alban ◽  
Defne Bayik ◽  
Balint Otvos ◽  
Matthew Grabowski ◽  
Manmeet Ahluwalia ◽  
...  
Keyword(s):  
Tumor Microenvironment ◽  
Tumor Growth ◽  
Immune Cell ◽  
Suppressor Cells ◽  
Myeloid Derived Suppressor Cells ◽  
Signaling Axis ◽  
Immunosuppressive Microenvironment ◽  
And Function

Abstract The immunosuppressive microenvironment in glioblastoma (GBM) enables persistent tumor growth and evasion from tumoricidal immune cell recognition. Despite a large accumulation of immune cells in the GBM microenvironment, tumor growth continues, and evidence for potent immunosuppression via myeloid derived suppressor cells (MDSCs) is now emerging. In agreement with these observations, we have recently established that increased MDSCs over time correlates with poor prognosis in GBM, making these cells of interest for therapeutic targeting. In seeking to reduce MDSCs in GBM, we previously identified the cytokine macrophage migration inhibitory factor (MIF) as a possible activator of MDSC function in GBM. Here, using a novel in vitro co-culture system to reproducibly and rapidly create GBM-educated MDSCs, we observed that MIF was essential in the generation of MDSCs and that MDSCs generated via this approach express a repertoire of MIF receptors. CD74 was the primary MIF receptor in monocytic MDSCs (M-MDSC), which penetrate the tumor microenvironment in preclinical models and patient samples. A screen of MIF/CD74 interaction inhibitors revealed that MN-166, a clinically relevant blood brain barrier penetrant drug, which is currently fast tracked for FDA approval, reduced MDSC generation and function in vitro. This effect was specific to M-MDSC subsets expressing CD74, and appeared as reduced downstream pERK signaling and MCP-1 secretion. In vivo, MN-166 was able reduce tumor-infiltrating MDSCs, while conferring a significant increase in survival in the syngeneic glioma model GL261. These data provide proof of concept that M-MDSCs can be targeted in the tumor microenvironment via MN-166 to reduce tumor growth and provide a rationale for future clinical assessment of MN-166 to reduce M-MDSCs in the tumor microenvironment. Ongoing studies are assessing the effects of MDSC inhibition in combination with immune activating approaches, in order to inhibit immune suppression while simultaneously activating the immune system.

scholarly journals DOP68 Thiomyristoyl ameliorates colitis by blocking the differentiation of Th17 cells via STAT3/IL-6 signal pathway

2020 ◽  
Vol 14 (Supplement_1) ◽  
pp. S105-S106
Author(s):  
Y XU ◽  
M zhang ◽  
G Xu ◽  
X Zou
Keyword(s):  
Flow Cytometry ◽  
Western Blotting ◽  
Th17 Cells ◽  
Culture Supernatant ◽  
Specific Inhibitor ◽  
Signal Pathway ◽  
T Cell Subsets ◽  
Enzyme Linked Immunosorbent Assay ◽  
Control Group ◽  
Ifn Γ

Abstract Background The pathogenesis of inflammatory bowel disease (IBD), including Crohn’s disease (CD) and ulcerative colitis (UC), has not been fully elucidated, but a strong correlation between IBD and the immune dysregulation of the host has been persistently detected. Sirtuin 2 (SIRT2), a histone deacetylase, has recently been found to play an important role in inflammation, infection and immunomodulatory. As a potent SIRT2-specific inhibitor, thiomyristoyl (TM) has an extensive anticancer activity, but its anti-inflammatory property remains unclear. Methods Human peripheral blood mononuclear cells (PBMC) were differentiated into T helper 17 (Th17) cells in vitro and were treated with TM simultaneously. Interleukin (IL)-17A in the culture supernatant, the ratio of T cells, the levels of phosphorylated-signal transducer and activator of transcription (p-stat3) and phosphorylated-nuclear factor kappa-B (p-NF κB) were determined by enzyme-linked immunosorbent assay (ELISA), flow cytometry or western blotting, respectively. Then, colitis C57BL/6J mice induced by 2.5% dextran sulphate sodium salt (DSS) were treated with TM, and were evaluated by disease activity index (DAI) and haematoxylin-eosin (HE) staining. T-cell subsets in the mice spleen, IL-6, IL-10 and IL-17A in serum, related factors retinoic acid receptor-related orphan receptor-γ t (ROR-γt), forkhead box protein P3 (FOXP3), IL-17A, interferon γ γ (IFN-γ), IL-23, IL-10 and hypoxia-inducible factor (HIF)-1α in mice colon were measured by flow cytometry, ELISA, quantitative real-time polymerase chain reaction (Q-PCR) and western blotting, respectively. Results Compared with the positive control group, the levels of IL-17A in culture supernatant, the percentage of Th17, the levels of p-stat3 and p-NF kappa B in cell lysate were lower in the TM group. Compared with PBS-treated colitis mice, TM-treated colitis mice had longer colons, fewer weight-losses, lower DAI and histopathologic scores. Interestingly, although the expression of IFN-γ, IL-17A, and ROR-γt was inhibited in the colons of TM-treated mice, the level of FOXP3 did not change. Consistently, the percentage of spleen Th17 cells was decreased in the TM group while the percentage of Treg cells was not affected. In addition, the TM group had reduced levels of IL-23 and HIF-1α, and an increased level of IL-10 in the colon, compared with colitis group. Conclusion TM ameliorates DSS-induced experimental colitis by blocking the differentiation of Th17 cells, which may be associated with the STAT3/IL-6 signal pathway. SIRT2 may represent a potential target for the treatment of IBD.

scholarly journals Hepatic Stellate Cells Enhance Liver Cancer Progression by Inducing Myeloid-Derived Suppressor Cells through Interleukin-6 Signaling

2019 ◽  
Vol 20 (20) ◽  
pp. 5079 ◽  
Author(s):  
Ching-Chuan Hsieh ◽  
Chien-Hui Hung ◽  
Meihua Chiang ◽  
Yu-Chin Tsai ◽  
Jie-Teng He
Keyword(s):  
T Cells ◽  
T Cell ◽  
Tumor Microenvironment ◽  
Hepatic Stellate Cells ◽  
Suppressor Cells ◽  
Stellate Cells ◽  
Myeloid Derived Suppressor Cells ◽  
Term Survival ◽  
Arginase 1

The tumor microenvironment, which consists of fibroblasts, smooth muscle cells, endothelial cells, immune cells, epithelial cells, and extracellular matrices, plays a crucial role in tumor progression. Hepatic stellate cells (HSCs), a class of unique liver stromal cells, participate in immunomodulatory activities by inducing the apoptosis of effector T-cells, generation of regulatory T-cells, and development of myeloid-derived suppressor cells (MDSCs) to achieve long-term survival of islet allografts. This study provides in vitro and in vivo evidences that HSCs induce the generation of MDSCs to promote hepatocellular carcinoma (HCC) progression through interleukin (IL)-6 secretion. HSC-induced MDSCs highly expressed inducible nitric oxide synthase (iNOS) and arginase 1 mRNA and presented potent inhibitory T-cell immune responses in the tumor environment. Wild-type HSC-induced MDSCs expressed lower levels of CD40, CD86, and MHC II, and a higher level of B7-H1 surface molecules, as well as increased the production of iNOS and arginase I compared with MDSCs induced by IL-6-deficient HSCs in vitro. A murine-transplanted model of the liver tumor showed that HCCs cotransplanted with HSCs could significantly enhance the tumor area and detect more MDSCs compared with HCCs alone or HCCs cotransplanted with HSCs lacking IL-6. In conclusion, the results indicated that MDSCs are induced mainly by HSCs through IL-6 signaling and produce inhibitory enzymes to reduce T-cell immunity and then promote HCC progression within the tumor microenvironment. Therapies targeting the pathway involved in MDSC production or its immune-modulating pathways can serve as an alternative immunotherapy for HCC.

Evaluating the Effect of Cinnarizine on Promastigotes and Amastigotes forms of Leishmania major

2020 ◽  
Vol 20 (4) ◽  
pp. 550-555 ◽  
Author(s):  
Lima Asgharpour Sarouey ◽  
Parvaneh Rahimi-Moghaddam ◽  
Fatemeh Tabatabaie ◽  
Khadijeh Khanaliha
Keyword(s):  
Flow Cytometry ◽  
Cutaneous Leishmaniasis ◽  
Leishmania Major ◽  
Inhibitory Effect ◽  
Control Group ◽  
Secondary Infections ◽  
Ic50 Values ◽  
J774 Cells ◽  
Mtt Assays

: As an important global disease, cutaneous leishmaniasis is associated with complications such as secondary infections and atrophic scars. The first line treatment with antimonials is expensive and reported to have serious side effects and enhance resistance development. The main objective of this study was to evaluate the effect of Cinnarizine on standard strains of Leishmania major because of paucity of information on this subject. Methods: In this experimental study, four concentrations of the drug (5, 10, 15 and 20 μg/ml) were added to Leishmania major cultures at 24, 48 and 72 hours intervals. MTT assays were performed to determine parasite viability and drug toxicity. Leishmania major promastigotes were augmented to the in vitro cultured macrophages (J774 cells) and then incubated for 72 hours. Half maximal inhibitory concentration (IC50) was ascertained by counting parasites. The inhibitory effect of the drug was compared with that of Glucantime. Flow-cytometry was performed to investigate apoptosis. Each test was repeated thrice. Results: The IC50 values of Cinnarizine after 72 hours were calculated to be 34.76 μg/ml and 23.73 μg/ml for promastigotes and amastigotes, respectively. The results of MTT assays showed 48 % promastigote viability after 72 hour-exposure to Cinnarizine at 20 μg/ml concentration. Programmed cell death in promastigote- and amastigote-infected macrophages was quantified to be 13.66 % and 98.7 %, respectively. Flow- cytometry analysis indicated that Cinnarizine induced early and late apoptosis in parasites. All treatments produced results which differed significantly from control group (P<0.05). Conclusion: Cinnarizine showed low toxicity with anti-leishmanial and apoptosis effects on both promastigote and intracellular amastigote forms. Therefore, we may suggest further assessment on animal models of this drug as candidates for cutaneous leishmaniasis therapy.

scholarly journals A Novel Mouse Monoclonal Antibody C42 against C-Terminal Peptide of Alpha-1-Antitrypsin

2021 ◽  
Vol 22 (4) ◽  
pp. 2141
Author(s):  
Srinu Tumpara ◽  
Elena Korenbaum ◽  
Mark Kühnel ◽  
Danny Jonigk ◽  
Beata Olejnicka ◽  
...  
Keyword(s):  
Monoclonal Antibody ◽  
Flow Cytometry ◽  
Western Blotting ◽  
Complex Mixture ◽  
Immunoglobulin M ◽  
Confocal Laser ◽  
Enzyme Linked Immunosorbent Assay ◽  
Dot Blot

The C-terminal-fragments of alpha1-antitrypsin (AAT) have been identified and their diverse biological roles have been reported in vitro and in vivo. These findings prompted us to develop a monoclonal antibody that specifically recognizes C-36 peptide (corresponding to residues 359–394) resulting from the protease-associated cleavage of AAT. The C-36-targeting mouse monoclonal Immunoglobulin M (IgM) antibody (containing κ light chains, clone C42) was generated and enzyme-linked immunosorbent assay (ELISA)-tested by Davids Biotechnologie GmbH, Germany. Here, we addressed the effectiveness of the novel C42 antibody in different immunoassay formats, such as dot- and Western blotting, confocal laser microscopy, and flow cytometry. According to the dot-blot results, our novel C42 antibody detects the C-36 peptide at a range of 0.1–0.05 µg and shows no cross-reactivity with native, polymerized, or oxidized forms of full-length AAT, the AAT-elastase complex mixture, as well as with shorter C-terminal fragments of AAT. However, the C42 antibody does not detect denatured peptide in SDS-PAGE/Western blotting assays. On the other hand, our C42 antibody, unconjugated as well as conjugated to DyLight488 fluorophore, when applied for immunofluorescence microscopy and flow cytometry assays, specifically detected the C-36 peptide in human blood cells. Altogether, we demonstrate that our novel C42 antibody successfully recognizes the C-36 peptide of AAT in a number of immunoassays and has potential to become an important tool in AAT-related studies.

755 CXCR1 and CXCR2 chemokine receptor agonists produced by tumors induce neutrophil extracellular traps that interfere with immune cytotoxicity

2020 ◽  
Vol 8 (Suppl 3) ◽  
pp. A803-A803 ◽  
Author(s):  
Alvaro Teijeira ◽  
Saray Garasa ◽  
Itziar Migueliz ◽  
Assunta Cirella ◽  
Ignacio Melero
Keyword(s):  
Cancer Patients ◽  
Tumor Cells ◽  
Mouse Models ◽  
Chemokine Receptor ◽  
Suppressor Cells ◽  
Neutrophil Extracellular Traps ◽  
Myeloid Derived Suppressor Cells ◽  
Extracellular Traps ◽  
Tumor Associated Neutrophils

BackgroundNeutrophils are expanded and abundant in an important fraction (up to 35% of patients) in cancer-bearing hosts. When neutrophils are expanded, they usually promote exert immunomodulatory functions promoting tumor progression and the generation of metastases. Neutrophils can undergo a specialized form of cell death called NETosis that is characterized by the extrusion of their DNA to contain infections. In cancer NETs have been described to promote metastases in mouse models. IL-8, a CXCR1/2 ligand clinically targeted by blocking antibodies, has been described to induce NETosis and is upregulated in many cancer patients. Our hypothesis is that chemokines secreted by cancer cells can mediate NETosis in tumor associated neutrophils and that NETs can be one of the immunomodulatory mechanisms provided by tumor associated neutrophils.MethodsNETosis induction of peripheral neutrophils and granulocytic myeloid derived suppressor cells by different chemotactic stimuli, tumor cell supernatants and cocultures upon CXCR1/2 blockade. NET immunodetection in mouse models and xenograft tumors upon CXCR1/2 blockade. In vitro tumor cytotoxicity assays in the presence/absence of NETs, and videomicroscopy studies in vitro and by intravital imaging to test NETs inhibition of immune cytotoxicity by immune-cell/target-cell inhibition. Tumor growth studies and metastases models in the presence of NETosis inhibitors and in combination with checkpoint blockade in mouse cancer models.ResultsUnder the influence of CXCR1 and CXCR2 chemokine receptor agonists and other chemotactic factors produced by tumors, neutrophils, and granulocytic myeloid-derived suppressor cells (MDSCs) from cancer patients extrude their neutrophil extracellular traps (NETs). In our hands, CXCR1 and CXCR2 agonists proved to be the major mediators of cancer-promoted NETosis. NETs wrap and coat tumor cells and shield them from cytotoxicity, as mediated by CD8+ T cells and natural killer (NK) cells, by obstructing contact between immune cells and the surrounding target cells. Tumor cells protected from cytotoxicity by NETs underlie successful cancer metastases in mice and the immunotherapeutic synergy of protein arginine deiminase 4 (PAD4) inhibitors, which curtail NETosis with immune checkpoint inhibitors. Intravital microscopy provides evidence of neutrophil NETs interfering cytolytic cytotoxic T lymphocytes (CTLs) and NK cell contacts with tumor cells.ConclusionsCXCR1 and 2 are the main receptors mediating NETosis of tumor associated neutrophils in our in-vitro and in vivo systems expressing high levels of CXCR1 and 2 ligands. NETs limit cancer cell cytotoxicity by impeding contacts with cancer cells.

scholarly journals Rgs2 Mediates Pro-Angiogenic Function of Myeloid Derived Suppressor Cells in the Tumor Microenvironment via Upregulation of MCP-1

PLoS ONE ◽  
2011 ◽  
Vol 6 (4) ◽  
pp. e18534 ◽  
Author(s):  
Kimberly C. Boelte ◽  
Laura E. Gordy ◽  
Sebastian Joyce ◽  
Mary Ann Thompson ◽  
Li Yang ◽  
...  
Keyword(s):  
Tumor Microenvironment ◽  
Suppressor Cells ◽  
Myeloid Derived Suppressor Cells
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