Bistability in macrophage polarization and metabolic implications

Macrophages are essential innate immune cells characterized by a high diversity and plasticity. In vitro, their full dynamic range of activation profiles include the classical pro-inflammatory (M1) and the alternative anti-inflammatory (M2) program. Bistability usually arises in biological systems that contain a positive-feedback loop or a mutually inhibitory, double-negative-feedback loop, which are common regulatory motifs reported for macrophage transitions from one activation state to the other one. This switch-like behavior of macrophage is observed at four different levels. First, a decision-making module in signal transduction includes mutual inhibitory interactions between M1 (STAT1 and NF-KB/p50-p65) and M2 (STAT3 and NF-KB/p50-p50) signaling pathways. Second, a switch-like behavior at the gene expression level includes complex network motifs of transcription factors and miRNAs. Third, those changes impact metabolic gene expression leading to several switches in energy production, NADPH and ROS production, TCA cycle functionality, biosynthesis and nitrogen metabolism. Fourth, metabolic changes are monitored by specialized metabolic sensors coupled to AMPK and mTOR activity to provide stability by maintaining the signals to promote either M1 or M2 activation. The targeting of robust molecular switches has the potential to treat a broad range of widespread diseases such as sepsis, cancer or chronic inflammatory diseases.

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Stabilization but no functional influence of HIF-1α expression in the intestinal epithelium during Salmonella Typhimurium infection

Infection and Immunity ◽

10.1128/iai.00222-21 ◽

2022 ◽

Author(s):

Laura Robrahn ◽

Aline Dupont ◽

Sandra Jumpertz ◽

Kaiyi Zhang ◽

Christian H. Holland ◽

...

Keyword(s):

Gene Expression ◽

Intestinal Epithelium ◽

Bacterial Infections ◽

Inflammatory Diseases ◽

Protein Stabilization ◽

Inflammatory Factors ◽

Intestinal Epithelial ◽

Data Set ◽

The Impact

The hypoxia-inducible transcription factor 1 (HIF-1) has been shown to enhance microbial killing and to ameliorate the course of bacterial infections. While the impact of HIF-1 on inflammatory diseases of the gut has been studied intensively, its function in bacterial infections of the gastrointestinal tract remains largely elusive. With the help of a publicly available gene expression data set, we could infer significant activation of HIF-1 after oral infection of mice with Salmonella Typhimurium. Immunohistochemistry and western blot analysis confirmed marked HIF-1α protein stabilization, especially in the intestinal epithelium. This prompted us to analyze conditional Hif1a -deficient mice to examine cell type-specific functions of HIF-1 in this model. Our results demonstrate enhanced non-canonical induction of HIF-1 activity upon Salmonella infection in the intestinal epithelium as well as in macrophages. Surprisingly, Hif1a deletion in intestinal epithelial cells did not impact on inflammatory gene expression, bacterial spread or disease outcome. In contrast, Hif1a deletion in myeloid cells enhanced intestinal Cxcl2 expression and reduced the cecal Salmonella load. In vitro , HIF-1α-deficient macrophages showed an overall impaired transcription of mRNA encoding pro-inflammatory factors, however, intracellular survival of Salmonella was not impacted by HIF-1α deficiency.

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Bidirectional Oscillatory Shear Stress Increases Pro-Atherogenic Gene Expressions (I-CAM1, E-Selectin and IL-6) in Endothelial Cells

ASME 2011 Summer Bioengineering Conference, Parts A and B ◽

10.1115/sbc2011-53747 ◽

2011 ◽

Author(s):

Amlan Chakraborty ◽

Venkatakrishna R. Jala ◽

Sutirtha Chakraborty ◽

R. Eric Berson ◽

M. Keith Sharp ◽

...

Keyword(s):

Gene Expression ◽

Endothelial Cells ◽

Shear Stress ◽

Atherosclerotic Plaque ◽

Inflammatory Diseases ◽

Leukotriene B4 ◽

Oscillatory Shear ◽

Gene Expressions

Wall shear stress (WSS) plays a key role in altering intracellular pathways and gene expression of endothelial cells, and has significant impacts on atherosclerotic plaque development (1–3). Further, the atherogenic regulators Leukotriene B4 (LTB4) and Lipopolysaccharide (LPS) have significant impacts on the pathophysiology of many inflammatory diseases. This study investigates the effects of oscillatory shear directionality on pro-atherogenic gene expression (I-CAM, E-Selectin, and IL-6) in the presence of LTB4 and LPS. An orbital shaker was used to expose the endothelial cells to oscillatory shear in culture dishes, and Computational fluid dynamics (CFD) was applied to quantify the shear stress on the bottom of the orbiting dish. Directionality of oscillatory shear was characterized by a newly developed hemodynamic parameter — Directional oscillatory shear index (DOSI), which was demonstrated in a previous study to significantly impact cell morphology (4). Results showed that DOSI significantly altered gene expression. Therefore, directionality of shear modulates atherosclerotic gene expression in vitro and thus, may influence the formation of atherosclerotic plaque in vivo.

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IRE1α regulates macrophage polarization, PD-L1 expression and tumor survival

10.1101/2020.02.17.952457 ◽

2020 ◽

Author(s):

Alyssa Batista ◽

Jeffrey J. Rodvold ◽

Su Xian ◽

Stephen Searles ◽

Alyssa Lew ◽

...

Keyword(s):

Gene Expression ◽

Tumor Microenvironment ◽

Macrophage Polarization ◽

Immune Surveillance ◽

Surface Expression ◽

Gene Signature ◽

Immune Dysregulation ◽

The Unfolded Protein Response

ABSTRACTIn the tumor microenvironment local immune dysregulation is driven in part by macrophages and dendritic cells that are polarized to a mixed proinflammatory/immune suppressive phenotype. The unfolded protein response (UPR) is emerging as the possible origin of these events. Here we report that the inositol-requiring enzyme 1 (IRE1α) branch of the UPR is directly involved in the polarization of macrophages in vitro and in vivo, including the upregulation of IL-6, IL-23, Arginase1, as well as surface expression of CD86 and PD-L1. Macrophages in which the IRE1α/Xbp1 axis is blocked pharmacologically or deleted genetically have significantly reduced polarization, and CD86 and PD-L1 expression, which was induced independent of IFNγ signaling suggesting a novel mechanism in PD-L1 regulation in macrophages. Mice with IRE1α- but not Xbp1-deficient macrophages showed greater survival than controls when implanted with B16.F10 melanoma cells. Remarkably, we found a significant association between the IRE1α gene signature and CD274 gene expression in tumor-infiltrating macrophages in humans. RNASeq analysis showed that bone marrow derived macrophages with IRE1α deletion lose the integrity of the gene connectivity characteristic of regulated IRE1α-dependent decay (RIDD) and the ability to activate CD274 gene expression. Thus, the IRE1α/Xbp1 axis drives the polarization of macrophages in the tumor microenvironment initiating a complex immune dysregulation leading to failure of local immune surveillance.

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TNF-Like Ligand 1 Aberrance Aggravates Nonalcoholic Steatohepatitis via M1 Macrophage Polarization

Oxidative Medicine and Cellular Longevity ◽

10.1155/2021/3877617 ◽

2021 ◽

Vol 2021 ◽

pp. 1-16

Author(s):

Yuxin Luo ◽

Jinbo Guo ◽

Wenxiu Jia ◽

Mengyao Wu ◽

Fengrong Yin ◽

...

Keyword(s):

Nonalcoholic Steatohepatitis ◽

Lipid Accumulation ◽

Inflammatory Diseases ◽

Macrophage Polarization ◽

Primary Biliary Cholangitis ◽

M2 Macrophage ◽

Hepatic Lipid Accumulation ◽

High Fructose ◽

M1 Macrophage

Nonalcoholic steatohepatitis (NASH) is a progressive, chronic liver disease worldwide which imposes a large economic burden on society. M1/M2 macrophage balance destruction and recruitment of mononuclear immune cells to the liver play critical roles in NASH. Several studies have shown that the expression of TNF-like ligand 1 aberrance (TL1A) increased in macrophages associated with many inflammatory diseases, for example, inflammatory bowel disease, primary biliary cholangitis, and liver fibrosis. One recent research showed that weight, abdominal adipose, and liver leptin, one of the critical fat cytokines, were reduced in TL1A knockout mice. However, the functional and molecular regulatory mechanisms of TL1A on macrophage polarization and recruitment in NASH have yet to be clarified. The authors found that high fructose high fat diet and methionine-choline deficiency diet induced the expression of TL1A in macrophages of liver tissue from murine NASH models. Myeloid-specific TL1A overexpressed mice showed exacerbated steatohepatitis with increased hepatic lipid accumulation, inflammation, liver injury, and apoptosis. M1 macrophages’ infiltration and the production of proinflammatory and chemotactic cytokines increased in liver of NASH mouse models with myeloid-specific TL1A overexpressed. Furthermore, this paper revealed that bone marrow-derived macrophages and Kupffer cells with overexpression of TL1A exacerbated the lipid accumulation and expression of proinflammatory factors in the murine primary hepatocytes after free fatty acid treatment in vitro. In conclusion, TL1A-mediated M1-type macrophage polarization and recruitment into the liver promoted steatohepatitis in murine NASH.

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Role of LpL (Lipoprotein Lipase) in Macrophage Polarization In Vitro and In Vivo

Arteriosclerosis Thrombosis and Vascular Biology ◽

10.1161/atvbaha.119.312389 ◽

2019 ◽

Vol 39 (10) ◽

pp. 1967-1985 ◽

Cited By ~ 4

Author(s):

Hye Rim Chang ◽

Tatjana Josefs ◽

Diego Scerbo ◽

Namrata Gumaste ◽

Yunying Hu ◽

...

Keyword(s):

Gene Expression ◽

Lipoprotein Lipase ◽

Lipid Accumulation ◽

Macrophage Polarization ◽

Myeloid Cell ◽

Acid Uptake ◽

Lipid Uptake ◽

Very Low Density Lipoproteins

Objective: Fatty acid uptake and oxidation characterize the metabolism of alternatively activated macrophage polarization in vitro, but the in vivo biology is less clear. We assessed the roles of LpL (lipoprotein lipase)-mediated lipid uptake in macrophage polarization in vitro and in several important tissues in vivo. Approach and Results: We created mice with both global and myeloid-cell specific LpL deficiency. LpL deficiency in the presence of VLDL (very low-density lipoproteins) altered gene expression of bone marrow–derived macrophages and led to reduced lipid uptake but an increase in some anti- and some proinflammatory markers. However, LpL deficiency did not alter lipid accumulation or gene expression in circulating monocytes nor did it change the ratio of Ly6C high /Ly6C low . In adipose tissue, less macrophage lipid accumulation was found with global but not myeloid-specific LpL deficiency. Neither deletion affected the expression of inflammatory genes. Global LpL deficiency also reduced the numbers of elicited peritoneal macrophages. Finally, we assessed gene expression in macrophages from atherosclerotic lesions during regression; LpL deficiency did not affect the polarity of plaque macrophages. Conclusions: The phenotypic changes observed in macrophages upon deletion of Lpl in vitro is not mimicked in tissue macrophages.

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Metabolic Switch and Cytotoxic Effect of Metformin on Burkitt Lymphoma

Frontiers in Oncology ◽

10.3389/fonc.2021.661102 ◽

2021 ◽

Vol 11 ◽

Author(s):

Irene Bagaloni ◽

Axel Visani ◽

Sara Biagiotti ◽

Annamaria Ruzzo ◽

Mohsen Navari ◽

...

Keyword(s):

Gene Expression ◽

Glucose Metabolism ◽

Gene Expression Profiling ◽

Burkitt Lymphoma ◽

Expression Profiling ◽

Aerobic Glycolysis ◽

Tca Cycle ◽

Metabolic Switch ◽

Biochemical Assays

Altered cellular energetic metabolism has recently emerged as important feature of neoplastic cells. Indeed, interfering with cancer cell metabolism might represent a suitable therapeutic strategy. In this study, we aimed to assess glucose metabolism activation in human lymphomas and evaluate how metformin can exert its action on lymphoma cells. We studied a large series of human lymphomas (N = 252) and an in vitro model of Burkitt lymphoma (BL) cells. We combined molecular biology techniques, including global gene expression profiling (GEP) analysis, quantitative PCR (qPCR) and Western blotting, and biochemical assays, aimed to assess pentose phosphate pathway, tricarboxylic acid (TCA) cycle, and aerobic glycolysis rates. We found that glucose metabolism is overall enhanced in most lymphoma subtypes, based on gene expression profiling (GEP), with general shift to aerobic glycolysis. By contrast, normal B cells only showed an overall increase in glucose usage during germinal center transition. Interestingly, not only highly proliferating aggressive lymphomas but also indolent ones, like marginal zone lymphomas, showed the phenomenon. Consistently, genes involved in glycolysis were confirmed to be overexpressed in BL cells by qPCR. Biochemical assays showed that while aerobic glycolysis is increased, TCA cycle is reduced. Finally, we showed that metformin can induce cell death in BL cells by stressing cellular metabolism through the induction of GLUT1, PKM2, and LDHA. In conclusion, we unveiled glucose metabolism abnormalities in human lymphomas and characterized the mechanism of action of metformin in Burkitt lymphoma model.

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Role of tissue factor in delayed bone repair induced by diabetic state in mice

PLoS ONE ◽

10.1371/journal.pone.0260754 ◽

2021 ◽

Vol 16 (12) ◽

pp. e0260754

Author(s):

Hiroki Ehara ◽

Kohei Tatsumi ◽

Yoshimasa Takafuji ◽

Naoyuki Kawao ◽

Masayoshi Ishida ◽

...

Keyword(s):

Gene Expression ◽

Tissue Factor ◽

Bone Repair ◽

Inflammatory Diseases ◽

Nuclear Factor Κb ◽

Osteoclast Formation ◽

Protective Effects ◽

Diabetic State ◽

Related Transcription Factor

Background Tissue factor (TF) is the primary activator of the extrinsic coagulation protease cascade. Although TF plays roles in various pathological states, such as thrombosis, inflammatory diseases, cancer, and atherosclerosis, its involvement in bone metabolism remains unknown. Materials and methods The present study examined the roles of TF in delayed bone repair induced by a diabetic state in mice using wild-type (WT) and low TF-expressing (LTF) male mice. A diabetic state was induced by intraperitoneal injections of streptozotocin (STZ). Results A prolonged diabetic state significantly reduced total and trabecular bone mineral densities (BMD) as well as cortical bone thickness in WT and LTF mice; these BMD parameters were similar between WT and LTF mice treated with or without STZ. The diabetic state induced in WT mice delayed the repair of the femur following injury. The diabetic state induced in LTF mice was associated with further delays in bone repair. In in vitro experiments, TF significantly decreased receptor activator of nuclear factor-κB ligand-induced osteoclast formation and osteoclastogenic gene expression in RAW264.7 cells. However, it did not affect the gene expression levels of runt-related transcription factor 2 and osterix as well as alkaline phosphatase activity in mouse primary osteoblasts. Conclusion Low TF state was associated with enhanced bone repair delay induced by diabetic state in mice. The TF-induced suppression of bone remodeling may be a contributing factor to the protective effects of TF against delayed bone repair in a diabetic state.

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Contribution of Thrombospondin-1 and -2 to Lipopolysaccharide-Induced Acute Respiratory Distress Syndrome

Mediators of Inflammation ◽

10.1155/2021/8876484 ◽

2021 ◽

Vol 2021 ◽

pp. 1-22

Author(s):

Qiang Li ◽

Xiaoxiao Fu ◽

Jiang Yuan ◽

Shu Han

Keyword(s):

T Regulatory Cells ◽

Inflammatory Diseases ◽

Macrophage Polarization ◽

Endothelial Permeability ◽

M2 Macrophage ◽

Vascular Endothelial ◽

Macrophage Phenotype ◽

Thrombospondin 1

Thrombospondin (TSP) proteins have been shown to impact T-cell adhesion, migration, differentiation, and apoptosis. Thrombospondin-1 (TSP-1) is specifically upregulated in several inflammatory diseases and can effectively promote lipopolysaccharide- (LPS-) induced inflammation. In contrast, thrombospondin-2 (TSP-2) has been associated with activation of “anti-inflammatory” T-regulatory cells (Tregs). In this study, we investigated the effects of both TSP-1 and TSP-2 overexpression on macrophage polarization and activation in vitro and in vivo. We analyzed the effects of TSP-1 and TSP-2 on inflammation, vascular endothelial permeability, edema, ultrastructural morphology, and apoptosis in lung tissues of an ARDS mouse model and cultured macrophages. Our results demonstrated that TSP-2 overexpression effectively attenuated LPS-induced ARDS in vivo and promoted M2 macrophage phenotype polarization in vitro. Furthermore, TSP-2 played a role in regulating pulmonary vascular barrier leakage by activating the PI3K/Akt pathway. Overall, our findings indicate that TSP-2 can modulate inflammation and could therefore be a potential therapeutic target against LPS-induced ARDS.

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Phenylbutyrate facilitates homeostasis of non-resolving inflammatory macrophages

Innate Immunity ◽

10.1177/1753425919879503 ◽

2019 ◽

Vol 26 (1) ◽

pp. 62-72 ◽

Cited By ~ 1

Author(s):

Allison Rahtes ◽

Kisha Pradhan ◽

Mimosa Sarma ◽

David Xie ◽

Chang Lu ◽

...

Keyword(s):

Gene Expression ◽

Low Dose ◽

Inflammatory Diseases ◽

Macrophage Polarization ◽

Related Factor ◽

Inflammatory Monocytes ◽

Inflammatory Protein ◽

Anti Inflammatory ◽

Inflammation Resolution ◽

Ccl2 Gene

Non-resolving inflammatory monocytes/macrophages are critically involved in the pathogenesis of chronic inflammatory diseases. However, mechanisms of macrophage polarization are not well understood, thus hindering the development of effective strategies to promote inflammation resolution. In this study, we report that macrophages polarized by subclinical super-low dose LPS preferentially expressed pro-inflammatory mediators such as ccl2 (which encodes the protein monocyte chemo attractant protein-1) with reduced expression of anti-inflammatory/homeostatic mediators such as slc40a1 (which encodes the protein ferroportin-1). We observed significantly elevated levels of the autophagy-associated and pro-inflammatory protein p62 in polarized macrophages, closely correlated with the inflammatory activation of ccl2 gene expression. In contrast, we noted a significant increase of ubiquitinated/inactive nuclear-erythroid-related factor 2 (NRF2), consistent with reduced slc40a1 gene expression in polarized macrophages. Addition of the homeostatic restorative agent phenylbutyrate (4-PBA) effectively reduced cellular levels of p62 as well as ccl2 gene induction by super-low dose LPS. On the other hand, application of 4-PBA also blocked the accumulation of ubiquitinated NRF2 and restored anti-inflammatory slc40a1 gene expression in macrophages. Together, our study provides novel insights with regard to macrophage polarization and reveals 4-PBA as a promising molecule in restoring macrophage homeostasis.

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Development of a Cell-Based Reporter Assay for Screening of Inhibitors of Hypoxia-Inducible Factor 2-Induced Gene Expression

CrossRef Listing of Deleted DOIs ◽

10.1177/1087057106289234 ◽

2006 ◽

Vol 11 (6) ◽

pp. 678-687 ◽

Cited By ~ 20

Author(s):

Girma M. Woldemichael ◽

James R. Vasselli ◽

Roberta S. Gardella ◽

Tawnya C. Mckee ◽

W. Marston Linehan ◽

...

Keyword(s):

Gene Expression ◽

Cell Lines ◽

High Throughput ◽

High Throughput Screening ◽

Dynamic Range ◽

Clear Cell ◽

Hypoxia Inducible Factor ◽

Luciferase Reporter ◽

Reporter Cell

Reporter cell lines have been developed for the identification of inhibitors of gene expression enhanced by hypoxia-inducible factor 2, which has been implicated as a transcription factor involved in the tumorigenesis of clear cell renal carcinoma. Stably transformed reporter clones of the human renal clear cell carcinoma cell line 786-O were generated by transfection or retroviral infection. Luciferase reporter expression in the vectors used was driven by either the natural human vascular endothelial growth factor (VEGF) promoter-enhancer or by the VEGF and the human endothelial nitric oxide synthase enhancers modulating minimal human cytomegalovirus promoter. Utility of the generated reporter cell lines was validated by introducing the von Hippel-Lindau protein complex and testing for reporter inducibility by hypoxia. The dynamic range in reporter activity under hypoxic stress was found to be at least 30- to 40-fold, with a signal-to-noise ratio of 60:1. Properties of the cell lines such as tolerance to up to 3% DMSO, signal stability with multiple in vitro passages, and utility in both 96- and 384-well plate formats indicated their suitability for use in a high-throughput screen. In addition, the potential use of these reporter lines in the evaluation of high-throughput screening hits in vivo in various mice models has been demonstrated.

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