Proliferation Of Myeloid-derived Suppressor Cells Research Articles

17β-estradiol promotes myeloid-derived suppressor cells functions and alleviates inflammatory bowel disease by activation of Stat3 and NF-κB signalings

Inflammatory bowel disease (IBD) describes a group of clinically common autoimmune diseases characterized by chronic intestinal inflammation, with gender differences in prevalence. Estrogen has been previously shown to exert anti-inflammatory action in IBD development, however, the mechanisms remain obscure. Recent research has revealed that myeloid-derived suppressor cells (MDSCs) play a protective role in IBD pathogenesis. To investigate the molecular mechanisms of estrogen steroid 17β-estradiol (E2) in IBD progression, we established IBD mouse models (DNB-induced) with or without prior ovariectomy (OVX) and E2 implantation. We found that OVX led to worse IBD symptoms and reduced MDSCs frequency, whereas E2 significantly alleviated these effects in vivo. Moreover, in vitro experiments showed that E2 promoted the proliferation and immunosuppressive function of MDSCs through phosphorylation of Stat3 and p65. Mechanistically, E2-mediated Stat3/p65 phosphorylation depends on the interaction between HOTAIR, a long non-coding RNA that are well-known in MDSCs proliferation, and Stat3/p65 respectively. In conclusion, our study revealed that E2 promotes the expansion and immunosuppressive function of MDSCs, and thus diminished the occurrence and development of IBD.

The Influence of Myeloid-Derived Suppressor Cell Expansion in Neuroinflammation and Neurodegenerative Diseases.

The neuro-immune axis has a crucial function both during physiological and pathological conditions. Among the immune cells, myeloid-derived suppressor cells (MDSCs) exert a pivotal role in regulating the immune response in many pathological conditions, influencing neuroinflammation and neurodegenerative disease progression. In chronic neuroinflammation, MDSCs could lead to exacerbation of the inflammatory state and eventually participate in the impairment of cognitive functions. To have a complete overview of the role of MDSCs in neurodegenerative diseases, research on PubMed for articles using a combination of terms made with Boolean operators was performed. According to the search strategy, 80 papers were retrieved. Among these, 44 papers met the eligibility criteria. The two subtypes of MDSCs, monocytic and polymorphonuclear MDSCs, behave differently in these diseases. The initial MDSC proliferation is fundamental for attenuating inflammation in Alzheimer's disease (AD), Parkinson's disease (PD), and multiple sclerosis (MS), but not in amyotrophic lateral sclerosis (ALS), where MDSC expansion leads to exacerbation of the disease. Moreover, the accumulation of MDSC subtypes in distinct organs changes during the disease. The proliferation of MDSC subtypes occurs at different disease stages and can influence the progression of each neurodegenerative disorder differently.

Noncanonical TRAIL Signaling Promotes Myeloid-Derived Suppressor Cell Abundance and Tumor Growth in Cholangiocarcinoma

Proapoptotic tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) signaling as a cause of cancer cell death is a well-established mechanism. However, TRAIL-receptor (TRAIL-R) agonists have had very limited anticancer activity in human beings, challenging the concept of TRAIL as a potent anticancer agent. Herein, we aimed to define mechanisms by which TRAIL+ cancer cells can leverage noncanonical TRAIL signaling in myeloid-derived suppressor cells (MDSCs) promoting their abundance in murine cholangiocarcinoma (CCA). Multiple immunocompetent syngeneic, orthotopic models of CCA were used. Single-cell RNA sequencing and cellular indexing of transcriptomes and epitopes by sequencing of CD45+ cells in murine tumors from the different CCA models was conducted. In multiple immunocompetent murine models of CCA, implantation of TRAIL+ murine cancer cells into Trail-r-/- mice resulted in a significant reduction in tumor volumes compared with wild-type mice. Tumor-bearing Trail-r-/- mice had a significant decrease in the abundance of MDSCs owing to attenuation of MDSC proliferation. Noncanonical TRAIL signaling with consequent nuclear factor-κB activation in MDSCs facilitated enhanced MDSC proliferation. Single-cell RNA sequencing and cellular indexing of transcriptomes and epitopes by sequencing of immune cells from murine tumors showed enrichment of a nuclear factor-κB activation signature in MDSCs. Moreover, MDSCs were resistant to TRAIL-mediated apoptosis owing to enhanced expression of cellular FLICE inhibitory protein, an inhibitor of proapoptotic TRAIL signaling. Accordingly, cellular FLICE inhibitory protein knockdown sensitized murine MDSCs to TRAIL-mediated apoptosis. Finally, cancer cell-restricted deletion of Trail significantly reduced MDSC abundance and murine tumor burden. Our findings highlight the therapeutic potential of targeting TRAIL+ cancer cells for treatment of a poorly immunogenic cancer.

27-Hydroxycholesterol/liver X receptor/apolipoprotein E mediates zearalenone-induced intestinal immunosuppression: A key target potentially linking zearalenone and cancer

Zearalenone (ZEN) is a mycotoxin that extensively contaminates food and feed, posing a significant threat to public health. However, the mechanisms behind ZEN-induced intestinal immunotoxicity remain unclear. In this study, Sprague-Dawley (SD) rats were exposed to ZEN at a dosage of 5 mg/kg/day b.w. for a duration of 14 days. The results demonstrated that ZEN exposure led to notable pathological alterations and immunosuppression within the intestine. Furthermore, ZEN exposure caused a significant reduction in the levels of apolipoprotein E (ApoE) and liver X receptor (LXR) (P < 0.05). Conversely, it upregulated the levels of myeloid-derived suppressor cells (MDSCs) markers (P < 0.05) and decreased the presence of 27-hydroxycholesterol (27-HC) in the intestine (P < 0.05). It was observed that ApoE or LXR agonists were able to mitigate the immunosuppressive effects induced by ZEN. Additionally, a bioinformatics analysis highlighted that the downregulation of ApoE might elevate the susceptibility to colorectal, breast, and lung cancers. These findings underscore the crucial role of the 27-HC/LXR/ApoE axis disruption in ZEN-induced MDSCs proliferation and subsequent inhibition of T lymphocyte activation within the rat intestine. Notably, ApoE may emerge as a pivotal target linking ZEN exposure to cancer development.

MicroRNA-211 Regulates Proliferation, Expansion, and Immune Inhibitory Function of Myeloid-Derived Suppressor Cells via Mediation of CHOP Expression

ABSTRACT Background Myeloid-derived suppressor cells (MDSCs) are capable of effectively repressing immune responses against tumors and orchestrating the tumor microenvironment, which can promote tumor angiogenesis and metastasis. The pathway networks used to modulate tumor-expanded MDSC accumulation and function remain unclear. This study identified microRNA-211 (miR-211), whose expression was significantly decreased by factors derived from tumors. Methods miR-211 was assumed to be critical in modulating the accumulation and activity of MDSCs isolated from ovarian cancer (OC)-bearing mice by targeting C/EBP homologous protein (CHOP). Results The upregulation of miR-211 repressed MDSC proliferation, inhibited MDSC immunosuppressive functions, and increased the number of co-incubated CD4+ and CD8+ cells. Furthermore, overexpression of miR-211 led to decreased activities of the NF-κB, PI3K/Akt, and STAT3 pathways and the subsequent downregulation of matrix metalloproteinases to promote tumor cell invasion and metastasis. CHOP overexpression counteracted the effects of miR-211 elevation on these phenotypic changes. Upregulation of miR-211 also dramatically impaired the activity of MDSCs and suppressed OC tumor growth in vivo. Conclusion These results indicated that the miR-211-CHOP axis in MDSCs plays an essential role in the metastasis and proliferation of tumor-expanded MDSCs and might represent a promising cancer treatment target.

Correlation between circulating myeloid-derived suppressor cells and serum miRNA21 in non-small cell lung cancer and its clinical significance

Objective: To investigate the effect and clinical significance of myeloid-derived suppressor cells (MDSCs) levels and serum miRNA-21 expression in advanced non-small cell lung cancer (NSCLC) patients. Methods: Advanced NSCLC patients treated at the First People’s Hospital of Tongxiang City, Zhejiang Province, between July 2021 and June 2022 were included in this study. MDSCs in the peripheral circulation were assessed by flow cytometry, and serum miRNA-21 expression was assessed by q-PCR. The effects of miRNA-21 on MDSC proliferation and ARG-1 and inducible nitric oxide synthase (iNOS) secretion from MDSCs were examined using in vitro cell experiments. In addition, the correlation between MDSC and CEA, CRP and Ki67 was analyzed. Results: Compared with those in the peripheral circulation of individuals in the control group, the expression levels of polymorphonuclear-MDSCs (PMN-MDSCs) and miRNA-21 in the peripheral circulation of NSCLC patients were significantly higher, and the two groups were significantly positively correlated. In in vitro cell experiments, miRNA-21 mimics promoted MDSC proliferation and increased ARG-1 and iNOS secretion and miRNA-21 inhibtor has an opposite result. In addition, PMN-MDSCs in the peripheral circulation of NSCLC patients were significantly positively correlated with carcinoembryonic antigen (CEA), C-reactive protein (CRP), and Ki67. Conclusion: The blood of advanced NSCLC patients contains high levels of MDSCs and miRNA-21, and the two may interact to impact on NSCLC.

SPLUNC1 regulates LPS-induced progression of nasopharyngeal carcinoma and proliferation of myeloid-derived suppressor cells.

Nasopharyngeal carcinoma (NPC) is one of the aggressive malignant tumors with high mortality, and the proliferation of myeloid-derived suppressor cells (MDSCs) could promote the metastasis of NPC through inhibiting the function of T cells. Meanwhile, SPLUNC1 was known to inhibit the malignant behavior of NPC cells, while the detailed function of SPLUNC1 in LPS-modified immune microenvironment of NPC remains unclear. To assess the impact of SPLUNC1 in immune microenvironment during the progression of NPC, NPC cells were exposed to LPS and then co-cultured with MDSCs for 48h. RT-qPCR and western blot were performed to evaluate the mRNA and protein level of SPLUNC1, CXCL-2 and CXCR-2, respectively. The level of IL-1β, IL-6, TNF-α, PD-L1, Arg-1 and iNOS were tested by ELISA. Meanwhile, the expression of CD33+ was tested by flow cytometry. The expression of CXCL-2 and CXCR-2 in NPC cells was higher, compared to that in NP69 cells. In contrast, SPLUNC1 level in NPC cells was much lower than that in NP69 cells. SPLUNC1 level was negatively correlated with CXCL-2 and CXCR-2. Overexpression of SPLUNC1 reversed LPS-induced inflammatory responses and proliferation in NPC cells. In addition, SPLUNC1 upregulation could reverse LPS-induced proliferation of MDSCs in tumor microenvironment. Meanwhile, SPLUNC1 overexpression could regulate CXCL-2/CXCR-2 axis through decreasing CXCL-2 and CXCR-2 protein and mRNA expression. SPLUNC1 regulates LPS-induced progression of nasopharyngeal carcinoma and proliferation of MDSCs. Thus, our study might provide a theoretical basis for discovering new strategies against NPC.

Effect of Angelica polysaccharide on mouse myeloid-derived suppressor cells

Angelica polysaccharide (APS) is a polysaccharide extracted from Angelica sinensis and it is one of the main active components of Angelica sinensis. Many studies have demonstrated that APS can promote the activation and function of a variety of immune cells and is recognized as an immune enhancer, but the regulatory effect of APS on myeloid-derived suppressor cells (MDSC) is still unclear. In this study, we investigated the effects of APS on MDSC proliferation, differentiation and function through in vivo and in vitro experiments. In vitro, our results showed that APS promoted the proliferation, differentiation and immunosuppressive function of MDSC through STAT1 and STAT3 signaling pathways, and positively correlated with the expression level of Mannose receptor (MR, also known as CD206) and in a concentration-dependent manner on APS. In vivo, APS up-regulated T cells, γδT cells, CD8+T cells, natural killer cells, monocytes/macrophages, and granulocytes in the peripheral blood and spleen of mice to varying degrees and was accompanied by the same degree of increase in the proportion of MDSC. That reminds to the clinician that when applying APS as treatment they should pay attention to its possible side effects of increasing the quantity and function of MDSC, in order to increase its efficacy.

Pseudolycorine chloride ameliorates Th17 cell-mediated central nervous system autoimmunity by restraining myeloid-derived suppressor cell expansion

Context The alkaloids of Narcissus tazetta L. var. Chinensis Roem (Amaryllidaceae) have antitumor and antiviral activities. However, the immunopharmacological effects of one of its constituents, pseudolycorine chloride (PLY), have not been reported yet. Objective We evaluated the effect of PLY on myeloid-derived suppressor cells (MDSCs) expansion and differentiation into monocyte-like MDSCs (M-MDSCs) and examined whether PLY alleviates Th17 cell-mediated experimental autoimmune encephalomyelitis (EAE), a murine model of multiple sclerosis (MS). Materials and methods In vitro, MDSCs were treated with PLY (0.67, 2 and 6 μM) or solcitinib (10 μM, positive control) for 48 or 96 h, and their proliferation, expansion, and differentiation into M-MDSCs were examined by flow cytometry. Myelin oligodendrocyte glycoprotein (MOG35–55) was used to induce EAE in female C57BL/6 mice, and the mice were treated with 40 mg/kg/d PLY or 1 mg/kg/d FK-506 (tacrolimus, positive control) for 21 days. Inflammatory infiltration, spinal cord demyelination, and MDSCs and Th17 cells infiltration into the spinal cord were examined using haematoxylin and eosin staining, Luxol fast blue staining, and immunofluorescence, respectively. Results In vitro, PLY (IC50/24 h = 6.18 μM) significantly inhibited IL-6 and GM-CSF-induced MDSCs proliferation, expansion and differentiation into M-MDSCs at all concentrations used. However, these concentrations did not show cytotoxicity. In mice, PLY (40 mg/kg) treatment alleviated EAE and inhibited inflammatory infiltration, demyelination, and MDSCs and Th17 cells infiltration into the spinal cord. Discussion and conclusions PLY may be an excellent candidate for the treatment of MS and other autoimmune diseases.

Abstract PO032: Tumor Necrosis Factor-Related Apoptosis-Inducing Ligand (TRAIL)-TRAIL-R Mediates Cholangiocarcinoma Tumor Immune Evasion by Enhancing Myeloid-Derived Suppressive Cell Population

Abstract Background and Aims: Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL), a member of TNF superfamily, is predominantly expressed on immune cells. Engagement of TRAIL with its cognate receptor, TRAIL-R (TR), leads to cancer cells apoptosis. Although the anti-tumor role of TRAIL-TR has been extensively studied, TRAIL agonists have had very limited anti-cancer activity in human clinical trials. TRAIL signaling can facilitate the myeloid response. However, this potential immunosuppressive function of TRAIL has not been examined extensively in cancer biology. Cholangiocarcinoma (CCA), a highly lethal biliary tract cancer, has a dense immunosuppressive microenvironment. Accordingly, CCA provides a model to examine the potential immune regulatory function of TRAIL in cancer biology. Methods: Using syngeneic, orthotopic murine models of CCA, (PMID: 29464042), murine CCA cells (SB cells) that express both Trail and Trail receptor (Tr) were implanted into livers of WT Tr− / − and LyzcreTrf/f mice, the latter have a specific deletion of TRAIL-R on myeloid cells. Hence, in this model the host immune cells express Trail but not the receptor; therefore, they would be capable of inducing TRAIL-mediated apoptosis in CCA cells but would be resistant to TRAIL-mediated immunosuppression. After 4 weeks of tumor growth, mice were sacrificed, and tumors were characterized using flow cytometry. Results: We observed that Tr− / − mice had a significant reduction in tumor burden compared to WT mice. Myeloid-derived suppressor cells (MDSCs) were significantly decreased in Tr− / − tumors compared to WT tumors. Implantation of SB cells into LyzcreTrf/f mice resulted in a marked reduction in tumor burden and attenuation of MDSC infiltration compared to Lyzcre mice. In vitro functional studies employing MDSCs from mice deficient in Tr (MDSC-Tr −/−) were carried out. Compared to WT MDSCs, coculture of MDSC-Tr− /- with SB cells resulted in a significant reduction in the proliferation and immunosuppressive function of MDSCs. Moreover, following cocultured with SB cells, immunofluorescence analysis demonstrated that MDSC-Tr− / − had a reduction in the nuclear translocation of the NFkB subunit P65 compared to WT MDSCs. These data suggest that TRAIL-TR augments MDSC proliferation via NFkB. In conclusion, we have demonstrated that Tr− / − mice have a significant reduction in CCA tumor burden and MDSC infiltration. These results indicate that TRAIL-TR facilitates tumor immune escape and progression by fostering MDSC immunosuppressive function and infiltration, in an NFkB-dependent manner. Direct targeting of TRAIL on CCA cells is a potential anti-tumor strategy. Citation Format: Emilien Loeuillard, Jingchun Yang, Dong Haidong, Gregory Gores, Sumera Ilyas. Tumor Necrosis Factor-Related Apoptosis-Inducing Ligand (TRAIL)-TRAIL-R Mediates Cholangiocarcinoma Tumor Immune Evasion by Enhancing Myeloid-Derived Suppressive Cell Population [abstract]. In: Proceedings of the AACR Special Conference: Advances in the Pathogenesis and Molecular Therapies of Liver Cancer; 2022 May 5-8; Boston, MA. Philadelphia (PA): AACR; Clin Cancer Res 2022;28(17_Suppl):Abstract nr PO032.

Abstract 2191: Alteration of anti-tumoral immunity in the pre-metastatic bone microenvironment via autonomic nerve system dysfunction

Abstract Bone metastasis is a major cause of morbidity and mortality for breast cancer patients. Bone is a unique metastatic microenvironment because of complex interactions among numerous distinct cell types such as osteo-blasts, -clasts, -cytes and marrow immune cells in physiological and pathological conditions. Imbalanced bone homeostasis by diverse factors such as the sympathetic nerve system (SNS) activation, potentially contribute to the progression of bone metastasis, yet precise mechanisms remain unclear. We investigated the effects of beta 2 adrenergic receptor (β2AR) activation in osteoblasts induced by prolonged SNS stimulation on anti-tumoral immunity in bone metastasis. We treated female Balb/C with chronic immobilization stress (CIS; 2 h. daily) and found that CIS increased syngeneic intra-tibial 4T1 bone metastasis growth as well as the number of CD11b+Gr1+ myeloid-derived suppressor cells (MDSC) counterbalancing anti-tumoral cytotoxic T cells in bone microenvironment. A β2AR-specific inhibitor (ICI-118551) or MDSC depletion using anti-Gr1 antibodies reversed CIS-induced bone metastatic tumor growth. More importantly, 2-week CIS pre-treatment increased EDU+ MDSC proliferation in the bone marrow, contributing to subsequent bone metastatic tumor growth and micro-metastatic seeding in intra-tibial and intra-cardiac tumor injection models, respectively. When CIS was administered after the establishment of bone metastatic tumors, the pro-tumorigenic effects were marginal. In osteoblast-specific β2AR knockout mice (Adrb2flox/flox; murine 2.3kb Col1-CreC57BL6 mice), CIS did not increase MDSC proliferation compared with littermate floxed controls, suggesting that β2AR in osteoblasts upregulates MDSC in bone microenvironment. RNA-sequencing transcriptome analysis of saline- or clenbuterol (a selective β2AR agonist)-treated murine osteoblasts showed that interleukin-6 (IL6) was the most significantly increased cytokine. Indeed, CIS or clenbuterol treatment increased IL-6 expression in vivo (immunohistochemistry) and in vitro (cultured osteoblasts), respectively. MDSC isolated from the bone marrow of CIS-treated mice had significantly increased STAT3 phosphorylation by flow cytometry and the expression of immuno-suppressive functional molecules such as arginase 1 (Arg1) and indolamine 2,3-dioxygenase 1 (Ido1) compared to those from control mice. Our data collectively demonstrated that SNS-dependent β2AR in osteoblasts expands and activates MDSC in the pre-metastatic bone microenvironment via the IL6-STAT3-Arg1/IDO1 pathway, creating a fertile soil for bone metastasis progression. Our data provided the first direct evidence for anti-tumoral immunity and pro-tumorigenic effects of autonomous nerve system dysfunction specific to the metastatic bone microenvironment. Citation Format: Eun Jung Lee, Da Hyeon Yun, Seungpil Jung, Serk In Park. Alteration of anti-tumoral immunity in the pre-metastatic bone microenvironment via autonomic nerve system dysfunction [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 2191.

Abstract P072: Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) foster myeloid-derived suppressor cell-mediated tumor immune evasion in cholangiocarcinoma

Abstract Background and Aims: Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) is predominantly expressed on immune cells. Although TRAIL biology has garnered considerable interest as a potential anti-cancer strategy, TRAIL agonists have had very limited anti-cancer activity in humans. TRAIL signaling in T cells may also potentially provide an immune checkpoint function as it can inhibit T cell activation and proliferation by interfering with T cell receptor signaling. However, this potential immune checkpoint function of TRAIL has not been examined in cancer biology. Cholangiocarcinoma (CCA), a highly lethal biliary tract cancer, provides a model to examine the potential immune checkpoint function of TRAIL. Methods: Using a syngeneic, orthotopic murine model of CCA (PMID: 29464042), murine CCA cells (SB cells) that express both TRAIL and TRAIL receptor (TR) were implanted into livers of WT and Tr−/− mice. Hence, in this model the host immune cells express TRAIL but not the receptor; therefore, they would be capable of inducing TRAIL-mediated apoptosis in CCA cells but would be resistant to TRAIL-mediated immunosuppression. After 4 weeks of tumor growth, mice were sacrificed, and tumors were characterized using flow cytometry. Results: We observed that Tr−/− mice had a significant reduction in tumor burden compared to WT mice. Moreover, tumor bearing Tr−/− mice had a significant increase in cytotoxic T lymphocytes (CTLs) and enhanced CTL effector function. However, coculture of T cells with SB cells or SB cells deficient in Tr (SB-Tr−/−) did not result in a significant difference in T cell apoptosis or function, implying that TRAIL-TR is not a direct T cell checkpoint. Myeloid derived suppressor cells (MDSCs) were significantly decreased in Tr−/− tumors compared to WT tumors. Furthermore, implantation of SB cells devoid of Trail (SB-Trail−/−) into WT mice resulted in a significant reduction in tumor burden and MDSC infiltration. Coculture of SB cells with MDSCs from Tr−/− mice attenuated MDSC proliferation and immunosuppression compared to WT MDSCs. Moreover, treatment of MDSCs from Tr−/− mice with TRAIL recombinant protein resulted in a reduction in their proliferation and immunosuppressive function compare to MDSCs from WT mice implying that TRAIL-TR fosters MDSC growth and immunosuppressive function. In conclusion, we have demonstrated that Tr−/− mice have a significant reduction in CCA tumor burden and MDSC infiltration. Consequently, Tr−/− mice bearing tumors have enhanced CTL infiltration and function. These data suggest that the TRAIL-TR system mediates tumor immune evasion via MDSCs. Herein, we suggest that TRAIL-TR appears to function as an indirect T cell checkpoint by augmenting the immunosuppressive effects of MDSCs. Citation Format: Emilien Loeuillard, Juan Wang, Jingchun Yang, Haidong Dong, Gregory Gores, Sumera Ilyas. Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) foster myeloid-derived suppressor cell-mediated tumor immune evasion in cholangiocarcinoma [abstract]. In: Abstracts: AACR Virtual Special Conference: Tumor Immunology and Immunotherapy; 2021 Oct 5-6. Philadelphia (PA): AACR; Cancer Immunol Res 2022;10(1 Suppl):Abstract nr P072.

179 Association of CX3CR1 and Myeloid-derived Suppressor Cell with Survival of Sepsis Patients and Risk of Acute Respiratory Distress Syndrome

Peripheral blood immune micro-environment has been shown to influence survival outcomes in sepsis patients. However, the association of immune micro-environment with clinical outcomes of immune-suppression induced by sepsis remains unclear. Here we performed comprehensive analysis of immune cell infiltration and clinically validated association of CX3CR1 and Myeloid-derived suppressor cell (MDSC) with survival of sepsis patients and risk of acute respiratory distress syndrome (ARDS). This study analyzed discovering cohort including all sepsis patients, which grouped by survival status and ARDS occurrence, from the Gene Expression Omnibus database (GSE10474, GSE32707, GSE66890). Core differential expressed genes (DEGs) and immune cell were selected between groups using LASSO and Random Forest machine learning algorithm. Most survival correlated gene and immune cell were validated in validating cohort (GSE65682) using Kaplan-Meier survival analysis. Since gene and immune cell were selected, patients from discovering cohort were divided into higher and lower group. To further explore underlying mechanism, weighted gene co-expression network analysis (WGCNA) were applied. For functional and pathway enrichment analysis, clusterProfiler R package was used for Gene Ontology (GO), Kyoto encyclopedia of genes and genomes (KEGG) analyses. Among 133 sepsis patients from discovering cohort, 110 down-regulated genes and 76 up-regulated genes were identified as DEGs based on the survival status, while 177 down-regulated genes and 89 up-regulated genes were identified as DEGs based on the ARDS risk, respectively. After combining the DEGs screened out via the LASSO and RF algorithms, 13 DEGs between survival status and 16 DEGs between ARDS risk, were selected simultaneously by these two algorithms. The gene of CX3CR1 and immune cell of MDSC were significantly higher in alive group and none-ARDS group, and they two were correlated most (Spearman r = 0.69 p<0.01). Among 479 patients in validating cohort, higher CX3CR1 was associated with better 28-day survival benefit (hazard ratio [HR] 2.657, 95% CI 1.838-3.843) and higher MDSC associated with better 28-day survival benefit (HR 2.205, 95% CI 1.516-2.206). Using the gene modules of WGCNA that correlated most with CX3CR1 expression level and MDSC proliferation level, biological processes of T cell cytokine production and regulation of glial cell migration were significantly enriched. We demonstrated that CX3CR1 and MDSC can predict ARDS risk and contribute survival benefit for sepsis patients. MDSC might be important component of the immune micro-environment and should be integrated into predictive biomarker panels for immune therapy and be validated in future prospective clinical trials.

Swertianolin ameliorates immune dysfunction in sepsis via blocking the immunosuppressive function of myeloid- derived suppressor cells.

In this study, we studied the long-term proliferation trajectory of myeloid-derived suppressor cells (MDSCs) in murine sepsis model and investigated whether swertianolin could modulate the immunosuppressive function of MDSCs. A murine sepsis model was established by cecal ligation and perforation (CLP), according to the Minimum Quality Threshold in Pre-Clinical Sepsis Studies (MQTiPSS) guidelines. The bone marrow and spleen of the mice were collected at 24 h, 72 h, 7 and 15 d after sepsis induction. The proportions of monocytic- MDSCs (M-MDSCs; CD11b+LY6G–LY6Chi) and granulocytic-MDSCs (G-MDSC, CD11b+ Ly6G+ Ly6Clow) were analyzed by flow cytometry. Then, we have investigated whether swertianolin could modulate the immunosuppressive function of MDSCs in in vitro experiments. G-MDSCs and M-MDSCs increased acutely after sepsis with high levels sustained over a long period of time. G-MDSCs were the main subtype identified in the murine model of sepsis with polymicrobial peritonitis. Furthermore, it was found that swertianolin reduced significantly interleukin-10 (IL-10), nitric oxide (NO), reactive oxygen species (ROS), and arginase production in MDSCs, while reducing MDSC proliferation and promoting MDSC differentiation into dendritic cells. Swertianolin also improved T-cell activity by blocking the immunosuppressive effect of MDSCs. Both subsets of MDSCs significantly increased in the bone marrow and spleen of the mice with sepsis, with GMDSCs being the main subtype identified. Swertianolin effectively regulated the functions of MDSCs and reduced immune suppression.

Realization of Osteolysis, Angiogenesis, Immunosuppression, and Drug Resistance by Extracellular Vesicles: Roles of RNAs and Proteins in Their Cargoes and of Ectonucleotidases of the Immunosuppressive Adenosinergic Noncanonical Pathway in the Bone Marrow Niche of Multiple Myeloma.

Simple SummaryMultiple myeloma (MM) is a disease that extensively involves bone, and angiogenesis and immunosuppression are important processes in the development of MM. Proteasome inhibitors and immunomodulatory drugs remarkably improve the survival of MM patients. However, MM is still an incurable disease that rapidly becomes resistant to these drugs. There is robust evidence that extracellular vesicles (EVs) contribute to cancer metastasis. Osteoclasts, in addition to immunosuppressive cells in the bone marrow (BM), are key players in osteolysis and immunosuppression. BM stromal cells and MM cells secrete EVs through which they communicate with each other: EVs, in fact, contain proteins, small RNAs, and long non-coding RNAs that mediate this communication and contribute to angiogenesis, osteolysis, and cancer dissemination and drug resistance. Ectoenzymes are expressed in myeloma cells, osteoclasts, and stromal cells and produce immunosuppressive adenosine. Recently, an antibody targeting CD38, an ectoenzyme, has been shown to improve the survival of patients with MM. Thus, understanding the properties of EV and ectoenzymes will help elucidate key processes of MM development.Angiogenesis and immunosuppression promote multiple myeloma (MM) development, and osteolysis is a primary feature of MM. Although immunomodulatory drugs and proteasome inhibitors (PIs) markedly improve the survival of patients with MM, this disease remains incurable. In the bone marrow niche, a chain of ectoenzymes, including CD38, produce immunosuppressive adenosine, inhibiting T cell proliferation as well as immunosuppressive cells. Therefore, anti-CD38 antibodies targeting myeloma cells have the potential to restore T cell responses to myeloma cells. Meanwhile extracellular vesicles (EVs) containing microRNAs, proteins such as cytokines and chemokines, long noncoding RNAs, and PIWI-interacting RNAs have been shown to act as communication tools in myeloma cell/microenvironment interactions. Via EVs, mesenchymal stem cells allow myeloma cell dissemination and confer PI resistance, whereas myeloma cells promote angiogenesis, myeloid-derived suppressor cell proliferation, and osteoclast differentiation and inhibit osteoblast differentiation. In this review, to understand key processes of MM development involving communication between myeloma cells and other cells in the tumor microenvironment, EV cargo and the non-canonical adenosinergic pathway are introduced, and ectoenzymes and EVs are discussed as potential druggable targets for the treatment of MM patients.

Repression of MUC1 Promotes Expansion and Suppressive Function of Myeloid-Derived Suppressor Cells in Pancreatic and Breast Cancer Murine Models.

Myeloid-derived suppressor cells (MDSCs) are immature myeloid cells that are responsible for immunosuppression in tumor microenvironment. Here we report the impact of mucin 1 (MUC1), a transmembrane glycoprotein, on proliferation and functional activity of MDSCs. To determine the role of MUC1 in MDSC phenotype, we analyzed MDSCs derived from wild type (WT) and MUC1-knockout (MUC1KO) mice bearing syngeneic pancreatic (KCKO) or breast (C57MG) tumors. We observed enhanced tumor growth of pancreatic and breast tumors in the MUC1KO mice compared to the WT mice. Enhanced tumor growth in the MUC1KO mice was associated with increased numbers of suppressive MDSCs and T regulatory (Tregs) cells in the tumor microenvironment. Compared to the WT host, MUC1KO host showed higher levels of iNOS, ARG1, and TGF-β, thus promoting proliferation of MDSCs with an immature and immune suppressive phenotype. When co-cultured with effector T cells, MDSCs from MUC1KO mice led to higher repression of IL-2 and IFN-γ production by T cells as compared to MDSCs from WT mice. Lastly, MDSCs from MUC1KO mice showed higher levels of c-Myc and activated pSTAT3 as compared to MDSCs from WT mice, suggesting increased survival, proliferation, and prevention of maturation of MDSCs in the MUC1KO host. We report diminished T cell function in the KO versus WT mice. In summary, the data suggest that MUC1 may regulate signaling pathways that are critical to maintain the immunosuppressive properties of MDSCs.

MicroRNA-150 inhibits myeloid-derived suppressor cells proliferation and function through negative regulation of ARG-1 in sepsis

AimsSepsis is defined as life-threatening organ dysfunction caused by a dysregulated host response to infection. The majority of sepsis-related deaths occur during late sepsis, which presents as a state of immunosuppression. Myeloid-derived suppressor cells (MDSCs) have been reported to promote immunosuppression during sepsis. Here we aim to understand the role of microRNAs in regulating MDSCs proliferation and immunosuppression function during sepsis. Main methodsMurine sepsis model was established using cecal ligation and puncture (CLP). A microarray was used to identify microRNAs with differential expression in murine sepsis. The effect of microRNA-150 on MDSCs proliferation and function was then evaluated. 140 multiple trauma patients from Tongji Hospital and 10 healthy controls were recruited. Peripheral blood samples were taken and the serum level of miR-150 was measured. Key findingsIn the murine model of sepsis, MDSCs expansion was noted in the spleen and bone marrow, while expression of miR-150 in MDSCs decreased. Replenishing miR-150 inhibited the expansion of MDSCs in both monocytic and polymorphonuclear subpopulations, as well as decreasing the immunosuppressive function of MDSCs, through down-regulation of ARG1. Both pro-inflammatory cytokine IL-6 and anti-inflammatory cytokines TGF-β and IL-10 were reduced by miR-150. In human, the serum level of miR-150 was down-regulated in septic patients and elevated in non-septic trauma patients compared to healthy controls. SignificanceOur study showed that MiR-150 is down-regulated during sepsis. Replenishing miR-150 reduces the immunosuppression function of MDSCs by down-regulating ARG1 in late sepsis. MiR-150 might serve as a potential therapeutic option for sepsis.

A single cell sequencing approach to understand the nature of myeloid-derived suppressor cells in tumor-bearing mice

Abstract Peripheral T Cell Lymphoma (PTCL), a type of non-Hodgkin’s lymphoma, is a heterogeneous group of mature T cells that have undergone transformation into a malignant form. The tumor microenvironment (TME) is very heterogeneous and is comprised of different cell types. Myeloid-derived suppressor cells (MDSC) are one of the vital components of the TME. Further, MDSC are associated with tumor growth, angiogenesis and enhanced metastasis. The aim of this study was to compare MSDCs isolated from TME and splenic MDSCs from tumor-bearing hosts (TBHs) as well as splenic MDSCs from naïve mice by using single cell RNA sequencing (scRNASeq). For this reason, C57BL/6 mice were injected with 1×106 EL4 cancer cells subcutaneously to induce tumor growth and 14 days later, cells were isolated from these three tissues, and flow cytometry and scRNASeq were performed. The flow cytometry results showed an increase in MDSC recruitment in the spleens from TBHs in comparison with spleens from naïve mice. scRNASeq analysis showed most of the genes upregulated in MDSCs from TME and spleen of TBHs were responsible for generation, recruitment and proliferation of MDSCs (Lgals5, CXCL10, Alox5ap, CCL6, IL1b, S100A8/A9/A11, CCL4, CCL5). Furthermore, we found several genes overexpressed in MDSCs promoted the proliferation and migration of cancer cells as well as tumor formation (Lyz2, Plac8, Ifitm3, Vim, S100A6, Crip1). In addition, we found some genes have anti-tumor effects by suppressing T and B cells (Lgals5, Ly6c2). In summary, these studies demonstrated that MDSCs from TME and spleens of TBHs have unique features for inducing immunosuppression, as well as recruitment and expansion of MDSCs in TBHs, which make these cells critical for target for immunotherapy.

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

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.

Single-Cell RNA-seq of Human Myeloid-Derived Suppressor Cells in Late Sepsis Reveals Multiple Subsets With Unique Transcriptional Responses: A Pilot Study.

Increased circulating myeloid-derived suppressor cells (MDSCs) are independently associated with poor long-term clinical outcomes in sepsis. Studies implicate subsets of MDSCs having unique roles in lymphocyte suppression; however, characterization of these cells after sepsis remains incomplete. We performed a pilot study to determine the transcriptomic landscape in MDSC subsets in sepsis using single-cell RNAseq (scRNA-seq). A mixture of whole blood myeloid-enriched and Ficoll-enriched PBMCs from two late septic patients on post-sepsis day 21 and two control subjects underwent Cellular Indexing of Transcriptomes and Epitopes by Sequencing (CITE-seq). We successfully identified the three MDSC subset clusters-granulocytic (G-), monocytic (M-), and early (E-) MDSCs. Sepsis was associated with a greater relative expansion of G-MDSCs versus M-MDSCs at 21 days as compared to control subjects. Genomic analysis between septic patients and control subjects revealed cell-specific and common differential expression of genes in both G-MDSC and M-MDSC subsets. Many of the common genes have previously been associated with MDSC proliferation and immunosuppressive function. Interestingly, there was no differential expression of several genes demonstrated in the literature to be vital to immunosuppression in cancer-induced MDSC. This pilot study successfully demonstrated that MDSCs maintain a transcriptomic profile that is immunosuppressive in late sepsis. Interestingly, the landscape in chronic critical illness is partially dependent on the original septic insult. Preliminary data would also indicate immunosuppressive MDSCs from late sepsis patients appear to have a somewhat unique transcriptome from cancer and/or other inflammatory diseases.