Accumulation Of Suppressor Cells Research Articles

Role of Polymorphonuclear Myeloid-Derived Suppressor Cells and Neutrophils in Ischemic Stroke.

Background Immune cells play avital role in the pathology of ischemic stroke. Neutrophils and polymorphonuclear myeloid-derived suppressor cells share a similar phenotype and have attracted increasing attention in immune regulation research, yet their dynamics in ischemic stroke remain elusive. Methods and Results Mice were randomly divided into 2 groups and intraperitoneally treated with anti-Ly6G (lymphocyte antigen 6 complex locus G) monoclonal antibody or saline. Distal middle cerebral artery occlusion and transient middle cerebral artery occlusion were applied to induce experimental stroke, and mice mortality was recorded until 28 days after stroke. Green fluorescent nissl staining was used to measure infarct volume. Cylinder and foot fault tests were used to evaluate neurological deficits. Immunofluorescence staining was conducted to confirm Ly6G neutralization and detect activated neutrophils and CD11b+Ly6G+ cells. Fluorescence-activated cell sorting was performed to evaluate polymorphonuclear myeloid-derived suppressor cell accumulation in brains and spleens after stroke. Anti-Ly6G antibody successfully depleted Ly6G expression in mice cortex but did not alter cortical physiological vasculature. Prophylactic anti-Ly6G antibody treatment ameliorated ischemic stroke outcomes in the subacute phase. Moreover, using immunofluorescence staining, we found that anti-Ly6G antibody suppressed activated neutrophil infiltration into parenchyma and decreased neutrophil extracellular trap formation in penumbra after stroke. Additionally, prophylactic anti-Ly6G antibody treatment reduced polymorphonuclear myeloid-derived suppressor cell accumulation inthe ischemic hemisphere. Conclusions Our study suggested a protective effect of prophylactic anti-Ly6G antibody administration against ischemic stroke by reducing activated neutrophil infiltration and neutrophil extracellular trap formation in parenchyma and suppressing polymorphonuclear myeloid-derived suppressor cell accumulation in thebrain. This study may provide a novel therapeutic approach for ischemic stroke.

In Chronic Hepatitis C Infection, Myeloid-Derived Suppressor Cell Accumulation and T Cell Dysfunctions Revert Partially and Late After Successful Direct-Acting Antiviral Treatment.

Chronic HCV infection is characterized by several immunological alterations, such as the accumulation of suppressor cells and of hyperactivated T lymphocytes. However, it is unclear whether direct-acting antiviral (DAA)-mediated HCV clearance restores immune dysfunctions. We performed a phenotypic characterization by flow cytometry of different immune cell subsets, including monocytic myeloid-derived suppressor cells (M-MDSCs) and T lymphocytes in 168 patients with persistent HCV infection not treated, under DAA therapies and sustained virological responders. Chronic HCV infection prompted the accumulation of M-MDSCs independently of patient and clinical characteristics, and altered their metabolic properties. HCV RNA was undetectable in the majority of patients just after few weeks of DAA therapy, whereas M-MDSC levels normalized only 6 months after therapy. In addition, HCV infection deeply perturbed the T cell compartment since a re-distribution of memory CD4+ and CD8+ T cells was observed at the expenses of naïve cells, and memory T lymphocytes displayed increased activation. Notably, these features were only partially restored by DAA therapies in the CD4, but not in the CD8, compartment as high immune activation levels persisted in the terminally differentiated memory CD8+ T cells even more than 1 year after sustained virological response. Together, these results suggest that successful DAA therapies do not lead to full immunological reconstitution as fast as viral clearance.

Resveratrol ameliorates Lewis lung carcinoma‐bearing mice development, decreases granulocytic myeloid‐derived suppressor cell accumulation and impairs its suppressive ability

Myeloid‐derived suppressor cells (MDSC) are a heterogeneous population of immature myeloid cells which consist of 2 subsets: granulocytic MDSC (G‐MDSC) and monocytic MDSC (M‐MDSC). MDSC expand in tumor‐bearing hosts and contribute to immunotherapeutic resistance by remarkably blocking effector T‐cell activation via different mechanisms. Resveratrol (RSV) is a polyphenol and it has been widely used for its various health benefits. However, the underlying mechanism of its anti‐tumor properties remains unclear. In this study, a transplantable mouse model was used to investigate the effects of RSV on MDSC. The results showed that RSV ameliorated tumor development by decreasing G‐MDSC accumulation, impairing its suppressive ability on CD8+T cells and promoting M‐MDSC differentiation into CD11c+ and F4/80+ cells. Our results indicated that RSV should be considered as a modular of MDSC suppressive function and that RSV is a novel booster for tumor immunotherapy.

HLA-G promotes myeloid-derived suppressor cell accumulation and suppressive activity during human pregnancy through engagement of the receptor ILT4.

Establishing and maintaining maternal-fetal tolerance is essential for a successful pregnancy; failure of immunological adaptation to pregnancy leads to severe complications such as abortion or preterm delivery. Myeloid-derived suppressor cells (MDSCs) are innate immune cells that suppress T-cell responses, expand during pregnancy and thus may play a role in tolerance induction. Human leucocyte antigen G (HLA-G) is a major histocompatibility complex (MHC) I molecule with immune-modulatory properties, which is expressed during pregnancy. Here, we investigated the impact of HLA-G on MDSCs accumulation and activation in pregnant women. We demonstrate that granulocytic MDSCs (GR-MDSCs) express receptors for HLA-G, namely immunoglobulin-like transcript (ILT) 2 and 4, and that ILT4-expression by GR-MDSCs is regulated during pregnancy. Stimulation with soluble HLA-G (sHLA-G) increased suppressive activity of GR-MDSCs, induced MDSCs from peripheral blood mononuclear cells (PBMCs) and led to phosphorylation of the signal transducer and activator of transcription 3 (STAT3) and induction of indoleamine-2,3-dioxygenase (IDO) in myeloid cells. Effects of sHLA-G on MDSC accumulation were mediated through ILT4. These results suggest an interaction between MDSCs and HLA-G in humans as a potential mechanism for maintaining maternal-fetal tolerance. Modulating MDSC function during pregnancy via HLA-G might provide new opportunities for a therapeutic manipulation of immunological pregnancy complications.

Abstract 2877: Exercise, alone and in combination with a whole tumor cell vaccine reduces mammary tumor cell growth and enhances anti-tumor immunity

Abstract Regular, moderate exercise (EX) can reduce both the incidence and recurrence of breast cancer (BC), and improve survival. Numerous biological mechanism(s) have been proposed to explain these beneficial clinical effects of EX. However, little work has been done to examine the effect of EX on immunomodulation, i.e. the balance between anti-tumor immunity and the emergence of immunosuppressive cells, in particular myeloid derived suppressor cells (MDSC). We have previously demonstrated that moderate EX significantly enhances antigen-specific CD4+ and CD8+ T cell responses and reduces regulatory T cell function in tumor free-mice. Thus, the goal of the current study was to determine if EX could enhance effector function and reduce immunosuppression in tumor bearing mice, and determine if there were any synergistic effects of EX and the administration of a whole tumor cell vaccine on the aforementioned outcomes. Female Balb/c mice were randomized into EX or sedentary control (SED) groups (n = 14-16/group) and had access to running wheel or standard cages, respectively, for 12 weeks prior to the injection of 5×10^4 lucerifase-transfected 4T1.2 tumor cells into the 4th mammary fat pad. Mice were further randomized into vaccination (n = 6-8/group) or vehicle control (n = 4/group) and administered 1×10^6 irradiated 4T1.2 cells (VAC) or HBSS (VEH) at day 7, 14, 21, and 28 post tumor injection. All mice were fed the AIN-76A diet; however, the EX mice (n = 14) were fed 90% of caloric intake of SED animals to remain in energy balance (prevent weight gain) over the course of the study. Primary tumor growth was quantified, and at sacrifice (day 35) organs were collected to assess the following endpoints: splenic antigen-specific CD8+ effector function and myeloid derived suppressor cell (MDSC) accumulation, and metastatic burden in femurs. EX mice, with or without vaccine, weighed significantly less than SED mice (p<0.001). There was a significant effect of both vaccination and EX on primary tumor growth (F(24,200) = 7.386, p<0.001), splenic IFN-γ production (KW = 11.43, p = 0.010); and the accumulation of MDSC (F(3,28) = 6.486, p = 0.021). However, there was only an EX effect on metastatic burden (p = 0.027). There was a synergistic effect of the combination of vaccine+EX on tumor growth, but no other endpoint. These results demonstrate that exercise alone (i.e. in a prevention model, 12 weeks prior to tumor implantation) is highly effective in reducing primary tumor growth and metastases in an aggressive tumor, and significantly shifted the balance of effector and immunosuppressive factors in the direction of anti-tumor immunity. Furthermore, these results demonstrate that exercise is a viable intervention that may yield significant clinical benefit when used in combination with therapeutic cancer vaccines. This work is supported by internal PSU pilot funds for CJR; T32AI074551-03 for WJT. Citation Format: William J. Turbitt, Donna Sosnoski, Andrea Mastro, Connie Rogers. Exercise, alone and in combination with a whole tumor cell vaccine reduces mammary tumor cell growth and enhances anti-tumor immunity. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 2877. doi:10.1158/1538-7445.AM2015-2877

IT-18 * GLIOBLASTOMA STAT3 AND PD-L1 EXPRESSION DRIVES ACCUMULATION OF IMMUNOSUPPRESSIVE LEUKOCYTES

BACKGROUND: Glioblastoma (GBM) is a devastating disease with a median survival of 14 months. Immunotherapies hold promise but are compromised by tumor-mediated immunosuppression including accumulation of regulatory T-cells (TREG's) and myeloid-derived suppressor cells (MDSC's). Mechanisms leading to TREG and MDSC expansion in GBM patients are unclear. Tumor-derived STAT3 and PD-L1 have been implicated in direct GBM-mediated immunosuppression. We hypothesized that they might also be critical for accumulation of TREG's and MDSC's. METHODS: STAT3 and PD-L1 expression was knocked down (KD) in primary human GBM cell cultures using lentiviral constructs expressing shRNA and confirmed by western blot. Wild type, PD-L1-KD, or STAT3-KD GBM cells were co-cultured with healthy donor human CD4+ lymphocytes or CD14+ monocytes. TREG (CD4 + /CD25 + /FoxP3+), monocytic MDSC's (CD14 + /HLA-DR-), and granulocytic MDSC's (CD11b + /CD14-/CD15+/-/CD33+) frequencies were measured by flow cytometry. RESULTS: Lentiviral shRNA produced efficient STAT3 and PD-L1 KD in primary human GBM cell cultures. STAT3-KD did not alter PD-L1 expression, suggesting they are independently regulated. GBM co-culture produced TREG expansion from naive CD4+ cells (2.01% + /-1,03% vs. 0.77% + /-0.10% of CD4+) but this was markedly reduced with GBM cell STAT3-KD and PD-L1-KD (1.15% + /-0.04%, 1.35% + /-0.04%; P < 0.05). Monocytic MDSC's were 1.24% of naive CD14+. This increased with GBM co-culture (7.61% + /-1.87%) but decreased to 0.48% + /-0.34% with STAT3-KD and 4.43% + /-0.64% with PD-L1-KD (P< 0.05). Similar results were seen for granulocytic MDSC's. CONCLUSION: These findings provide clear evidence that expression of STAT3 and PD-L1, two important immunosuppressive signals, are independently regulated in human GBM cells. Furthermore, GBM cell STAT3 and PD-L1 expression are critical drivers of immunosuppressive leukocyte (TREG and MDSC) accumulation from naive CD4+ and CD14+ cells in addition to their more widely appreciated direct immunosuppressive effects on activated T cells. These findings suggest that targeting both STAT3 and PD-L1 may have synergistic immunomodulatory effects of potential benefit to GBM patients.

Functional and Molecular Characterization of Myeloid Suppressor Cells Expanded During Lymphoma Tumour Progression

We previously reported that in mice with large progressing T‐cell lymphoma tumours, dysfunctions in the antitumour CTL activity occur, associated with an accumulation of splenic arginase‐producing myeloid suppressor cells (MSCs). In this study, we first demonstrate that both the presence and the activation state of these MSC depends on tumour evolution. While in tumour regressors hardly any arginase‐producing MSC can be found, both the amount and the arginase activity of this population expands from early over late progressors. This gradual induction of MSCs is paralleled by an increasing suppression of CTL activity and Th1, but not Th2, cytokine production. Upon analysing the molecular repertoire of MSC in vitro, we found, besides arginase1, a well‐established marker for alternatively activated myeloid cells or M2, a strong upregulation of FIZZ1 and Ym, two additional recently identified markers for M2. Further evaluation of molecular markers by microarray analysis in MSC yielded genes involved in wound healing (e.g. coagulation factor XIIIa), anti‐inflammation (e.g. selenoprotein P), immunomodulation (e.g. PD‐L2) and fat and sugar metabolism (e.g. leptin receptor). Of note, many of these genes are regulated by type 2 cytokines (IL‐4, IL‐13 and IL‐10) and are therefore rather M2 associated. Overall, our data provide new markers for MSC in cancer and further establish their M2 activation state.