TGF-? induces M2-like macrophage polarization via SNAIL-mediated suppression of a pro-inflammatory phenotype.
ABSTRACT: Tumor-associated macrophages (TAMs) are a major component of leukocytic infiltrate in tumors, which facilitates tumor progression and promotes inflammation. TGF-? promotes the differentiation of non-activated macrophages into a TAM-like (M2-like) phenotype; however, the underlying mechanisms are not clear. In this study, we found that TGF-? induces a M2-like phenotype characterized by up-regulation of the anti-inflammatory cytokine IL-10, and down-regulation of the pro-inflammatory cytokines TNF-? and IL-12. In human THP-1 macrophages, overexpression of SNAIL caused M2-like differentiation by inhibiting pro-inflammatory cytokine release and promoting the expression of M2-specific markers. By contrast, SNAIL knockdown promoted M1 polarization through up-regulation of pro-inflammatory cytokines and abolished TGF-?-mediated M2-polarization of THP-1 macrophages. The SMAD2/3 and PI3K/AKT signaling pathways were crucial for TGF-?-induced SNAIL overexpression in THP-1 cells. These findings suggest that TGF-? skews macrophage polarization towards a M2-like phenotype via SNAIL up-regulation, and blockade of TGF-?/SNAIL signaling restores the production of pro-inflammatory cytokines. This study provides new understanding of the role of SNAIL in M2 polarization of macrophages, and suggests a potential therapeutic target for antitumor immunity.
Project description:Feto-placental Hofbauer cells (HBCs) are macrophages residing in placental stroma. They are generally described as anti-inflammatory M2 polarized cells, promoting tolerance and tissue remodeling. In certain pathologies, however, a possible phenotypical switch towards pro-inflammatory M1 macrophages has been proposed. The study aimed to determine if HBCs can acquire an M1 phenotype under pro-inflammatory conditions in vitro. HBCs were isolated from healthy human term placentas. Cells were cultivated upon addition of LPS and INF-? or IL-4 and IL-13 to induce the M1 and M2 phenotype, respectively. Specific cell polarization markers and cytokines, associated with respective phenotypes, were investigated by flow cytometry and ELISA. THP-1 macrophages served as positive control. Pro-inflammatory stimuli reduced M2 markers CD163 and DC-SIGN, but did not induce M1 markers. TNF-? release was increased, but at the same time TGF-? and IL-10 release was upregulated, resembling in part the M2b sub-phenotype. Anti-inflammatory stimuli had no effect on HBC polarization. HBCs maintain their M2 phenotype in vitro despite inflammatory stimuli, which might represent a state of adaption and tolerance to avoid rejection of the semiallogeneic feto-placental unit.
Project description:Fermented dairy products have become attractive functional foods for the delivery of probiotics and their biologically active metabolites. The aim of this study was to examine the immunomodulatory activity of milk fermented with the probiotic lactic acid bacterium Lactobacillus rhamnosus R0011 (LrF) on macrophages challenged with lipopolysaccharide (LPS), a potent pro-inflammatory stimulus. To this end, human THP-1 or U937 monocytes were differentiated into resting macrophages then stimulated with LPS and co-incubated with the LrF or with milk controls. Levels of pro-inflammatory and immunoregulatory cytokines were determined by enzyme-linked immunosorbent assays. Culturing of LPS-stimulated U937 macrophages with either the whole or filtered LrF resulted in an increase in Interleukin (IL)-1Ra production relative to the negative control. THP-1 macrophages cultured with the LrF demonstrated an increase in LPS-induced IL-10 and IL-1? production, while production of LPS-induced IL-6, sCD54, IL-8, IL-1?, TNF-?, IL-12p70 and Transforming Growth Factor-? (TGF-?) was unaffected. Further, the LrF induced the expression of DC-SIGN and CD206, markers of immunoregulatory M2 macrophage polarization, in LPS-challenged THP-1 macrophages. Taken together, milk fermented with L. rhamnosus R0011 increased regulatory cytokine production from LPS-challenged U937 and THP-1 macrophages, while simultaneously up-regulating the production of IL-1? and expression of DC-SIGN and CD206, a profile characteristic of polarization into the immunoregulatory M2 macrophage phenotype.
Project description:The concept of macrophage polarization toward different phenotypes after CNS injury has been increasingly discussed. Here, we propose that CD200 treatment may help shift pro-inflammatory macrophages to an arginase 1 (Arg1)-, transglutaminase 2 (TGM2)-, and transforming growth factor beta 1 (TGF-?)-positive phenotype. Rat macrophages were stimulated by interferon ? and lipopolysaccharide (LPS) to induce pro-inflammatory phenotypes. Treatment with human CD200-Fc up-regulated expression levels of alternatively activated M2-like markers such as Arg1 and TGM2 but suppressed pro-inflammatory M1-like markers such as toll-like receptor 4, interleukin 1 beta (IL-1?), IL-6, and GM-CSF. Concomitantly, CD200-Fc enhanced (CCAAT/enhancer-binding protein) C/EBP-beta promoter activity, whereas NF-?B activity was suppressed. Treatment with CD200-Fc also up-regulated potentially beneficial TGF-? expression in macrophages. When C/EBP-beta signaling was suppressed with siRNA, the effect of CD200-Fc on Arg1, TGM2 and TGF-? up-regulation was canceled. Taken together, these data provide proof-of-principle that targeting CD200 signaling may be a novel therapeutic approach to shift macrophages toward M2-like polarization via modulating cAMP-response element binding protein-C/EBP-beta transcriptional activity. We showed that CD200 treatment decreased pro-inflammatory cytokines (IL-1?, IL-6, and GM-CSF) along with suppressed inflammatory NF-?B activity in pro-inflammatory M?. On the other hand, CD200 increased Arg1, TGM2, and TGF-? production through CREB-C/EBP? signaling. We think that these findings provide proof-of-concept that CD200 signaling may play a key role in regulating macrophage polarization toward anti-inflammatory phenotypes.
Project description:Abnormal lipid-mediated hepatic inflammatory-immune dysfunction and chronic low grade inflammation play an important role in the pathogenesis of non-alcoholic fatty liver disease (NAFLD). Macrophage polarization is an important mechanism for the regulation of inflammatory response. Since PPAR-? has emerged as a master regulator of macrophage polarization, we aimed to investigate the lipid-induced macrophage/Kupffer cell polarization in vivo and in vitro, and explore the association between PPAR-? activity and macrophages M1/M2 polarization shifting. Here we showed that long-term high-fat diet increased Kupffer cells content with M1-predominant phenotype and increasing production of pro-inflammatory cytokines. Saturated fatty acids polarized Kupffer cells/macrophages to an M1-predominant phenotype while n-3 PUFA polarized Kupffer cells/macrophages to an M2 phenotype, which was associated with activation of NF-?B signal pathway and PPAR-? respectively. Furthermore, up-regulation of PPAR-? shifted lipid-induced macrophages polarization from M1-predominant phenotype to M2 phenotype. Macrophages polarization switch was associated with the interaction between PPAR-? and NF-?Bp65 signal pathway. Rosiglitazone restored high-fat diet-induced imblance of Kupffer cells M1/M2 polarization and alleviated hepatic steatosis as well as local pro-inflammatory response. These findings suggest that manipulation of PPAR-? activity has the potential to balance lipid-induced M1/M2 macrophage/Kupffer cell polarization, and leading to prevent the development of NAFLD.
Project description:Cartilage lesions and osteoarthritis (OA) presents an ever-increasing clinical and socioeconomic burden. Synovial inflammation and articular inflammatory environment are the key factor for chondrocytes apoptosis and hypertrophy, ectopic bone formation and OA progression. To effectively treat OA, it is critical to develop a drug that skews inflammation toward a pro-chondrogenic microenvironment. In this narrative and critical review, we aim to see the potential use of immune cells modulation or cell therapy as therapeutic alternatives to OA patients. Macrophages are immune cells that are present in synovial lining, with different roles depending on their subtypes. These cells can polarize to pro-inflammatory (M1) and anti-inflammatory (M2) phenotypes, being the latter associated with wound-healing by the production of ARG-1 and pro-chondrogenic cytokines, such as IL-10, IL-1RA, and TGF-b. Emerging evidence reveals that macrophage shift can be determined by several stimuli, apart from the conventional in vitro IL-4, IL-13, and IL-10. Evidences show the potential of physical exercise to induce type 2 response, favoring M2 polarization. Moreover, macrophages in contact with oxLDL have effect on the production of anabolic mediators as TGF-b. In the same direction, type II collagen, that plays a critical role in development and maturation process of chondrocytes, can also induce M2 macrophages, increasing TGF-b. The mTOR pathway activation in macrophages was shown to be able to polarize macrophages in vitro, though further studies are required. The possibility to use mesenchymal stem cells (MSCs) in cartilage restoration have a more concrete literature, besides, MSCs also have the capability to induce M2 macrophages. In the other direction, M1 polarized macrophages inhibit the proliferation and viability of MSCs and impair their ability to immunosuppress the environment, preventing cartilage repair. Therefore, even though MSCs therapeutic researches advances, other sources of M2 polarization are attractive issues, and further studies will contribute to the possibility to manipulate this polarization and to use it as a therapeutic approach in OA patients.
Project description:STAMP2 is a counterregulator of inflammation and insulin resistance. The aim of this study is to investigate whether activation of STAMP2 improves insulin resistance by regulating macrophage polarization in adipose tissues. The diabetic ApoE⁻/⁻/LDLR⁻/⁻ mouse model was induced by high-fat diet and low-dose streptozotocin. Samples were obtained from epididymal, subcutaneous and brown adipose tissues. Infiltration of M1/M2 macrophages and inflammatory cytokines were investigated by immunohistochemistry. We then used gene overexpression to investigate the effect of STAMP2 on macrophages infiltration and polarization and inflammatory cytokines expression. Our results showed that infiltration of macrophages, the ratio of M1/M2 macrophages and the expression of pro-inflammatory cytokines were enhanced and STAMP2 was downregulated in adipose tissues of diabetic ApoE⁻/⁻/LDLR⁻/⁻ mice compared with control mice. STAMP2 gene overexpression could significantly reduce macrophages infiltration, the ratio of M1/M2 macrophages and the expression of pro-inflammatory cytokines in epididymal and brown adipose tissues, improving insulin resistance. Our results suggested that STAMP2 gene overexpression may improve insulin resistance via regulating macrophage polarization in visceral and brown adipose tissues.
Project description:BACKGROUND:An uncontrolled inflammatory response is a critical pathophysiological feature of sepsis. Mesenchymal stem cells (MSCs) induce macrophage phenotype polarization and reduce inflammation in sepsis. MSC-secreted transforming growth factor beta (TGF-?) participated in the immune modulatory function of MSCs. However, the underlying mechanism of MSC-secreted TGF-? was not fully elucidated in regulation macrophage M2-like polarization. METHODS:The paracrine effects of MSCs on macrophage polarization were studied using a co-culture protocol with LPS-stimulated RAW264.7 cells/mouse peritoneal macrophages and MSCs. The effect of TGF-? in the co-culture system was blocked by the TGF-? receptor inhibitor. To determine the role of MSC-secreted TGF-?, we used recombinant TGF-? to culture with LPS-stimulated RAW264.7 cells. In addition, we employed antibody microarray analysis to determine the mechanisms of MSC secreted TGF-? on LPS-stimulated RAW264.7 cell/mouse peritoneal macrophage M2-like polarization. Furthermore, we used an Akt inhibitor and a FoxO1 inhibitor to inhibit the Akt/FoxO1 pathway. The nuclear translocation of FoxO1 was detected by Western blot. RESULTS:MSCs induced LPS-stimulated RAW264.7 cell/mouse peritoneal macrophage polarization towards the M2-like phenotype and significantly reduced pro-inflammatory cytokine levels via paracrine, which was inhibited by TGF-? receptor inhibitor. Furthermore, we found that MSC-secreted TGF-? enhanced the macrophage phagocytic ability. The antibody microarray analysis and Western blot verified that TGF-? treatment activated the Akt/FoxO1 pathway in LPS-stimulated macrophages, TGF-?-induced FoxO1 nuclear translocation and obviously expressed in the cytoplasm, the effects of TGF-? regulatory effects on LPS-stimulated macrophage were inhibited by pre-treatment with Akt inhibitor and FoxO1 inhibitor. CONCLUSIONS:TGF-? secreted by MSCs could skew LPS-stimulated macrophage polarization towards the M2-like phenotype, reduce inflammatory reactions, and improve the phagocytic ability via the Akt/FoxO1 pathway, providing potential therapeutic strategies for sepsis.
Project description:Adipose tissue resident macrophages have important roles in the maintenance of tissue homeostasis and regulate insulin sensitivity for example by secreting pro-inflammatory or anti-inflammatory cytokines. Here, we show that M2-like macrophages in adipose tissue regulate systemic glucose homeostasis by inhibiting adipocyte progenitor proliferation via the CD206/TGF? signaling pathway. We show that adipose tissue CD206+ cells are primarily M2-like macrophages, and ablation of CD206+ M2-like macrophages improves systemic insulin sensitivity, which was associated with an increased number of smaller adipocytes. Mice genetically engineered to have reduced numbers of CD206+ M2-like macrophages show a down-regulation of TGF? signaling in adipose tissue, together with up-regulated proliferation and differentiation of adipocyte progenitors. Our findings indicate that CD206+ M2-like macrophages in adipose tissues create a microenvironment that inhibits growth and differentiation of adipocyte progenitors and, thereby, control adiposity and systemic insulin sensitivity.Adipose tissue contains macrophages that can influence both local and systemic metabolism via the secretion of cytokines. Here, Nawaz et al. report that M2-like macrophages, present in adipose tissue, create a microenvironment that inhibits proliferation of adipocyte progenitors due to the secretion of TGF-?1.
Project description:Macrophages play an essential role in host defense and display remarkable plasticity in switching between classically (pro-inflammatory-M1) and alternatively activated (anti-inflammatory-M2) phenotypes. The molecular mechanisms of macrophage polarization are not fully understood. Long non-coding RNAs (lncRNAs) with a length of?>?200 nucleotides have been shown to play diverse roles in biological processes. Aberrant expression of lncRNAs is associated with a variety of pathophysiological conditions such as cancer, diabetes, cardiovascular, pulmonary diseases, and tissue fibrosis. In this study, we investigated the role of lncRNA FENDRR in human and mouse macrophage polarization. Human THP-1 monocytes were activated with phorbol-12-myristate-13-acetate (PMA) and differentiated into M1 macrophages with IFN? or M2 macrophages with IL4. Real-time PCR analysis revealed that FENDRR was expressed 80-fold higher in M1 macrophages than that in M2 macrophages. Overexpression of FENDRR in PMA-activated THP-1 cells increased the IFN?-induced expression of M1 markers, including IL1? and TNF? at both mRNA and protein levels. Knockdown of FENDRR had an opposite effect. Similarly, FENDRR overexpression in primary mouse bone marrow-derived macrophages increased mRNA expression of M1 markers. FENDRR overexpression increased, while FENDRR knock-down decreased, the IFN?-induced phosphorylation of STAT1 in PMA-activated THP-1 cells. Our studies suggest that FENDRR enhances IFN?-induced M1 macrophage polarization via the STAT1 pathway.
Project description:Polychlorinated biphenyls (PCBs) are persistent organic pollutants that contribute to inflammatory diseases such as atherosclerosis, and macrophages play a key role in the overall inflammatory response. Depending on specific environmental stimuli, macrophages can be polarized either to pro-inflammatory (e.g., M1) or anti-inflammatory (e.g., M2) phenotypes. We hypothesize that dioxin-like PCBs can contribute to macrophage polarization associated with inflammation. To test this hypothesis, human monocytes (THP-1) were differentiated to macrophages and subsequently exposed to PCB 126. Exposure to PCB 126, but not to PCB 153 or 118, significantly induced the expression of inflammatory cytokines, including TNF? and IL-1?, suggesting polarization to the pro-inflammatory M1 phenotype. Additionally, monocyte chemoattractant protein-1 (MCP-1) was increased in PCB 126-activated macrophages, suggesting induction of chemokines which regulate immune cell recruitment and infiltration of monocytes/macrophages into vascular tissues. In addition, oxidative stress sensitive markers including nuclear factor (erythroid-derived 2)-like 2 (NFE2L2; Nrf2) and down-stream genes, such as heme oxygenase 1 (HMOX1) and NAD(P)H quinone oxidoreductase 1 (NQO1), were induced following PCB 126 exposure. Since dioxin-like PCBs may elicit inflammatory cascades through multiple mechanisms, we then pretreated macrophages with both aryl hydrocarbon receptor (AhR) and NF-?B antagonists prior to PCB treatment. The NF-?B antagonist BMS-345541 significantly decreased mRNA and protein levels of multiple cytokines by approximately 50% compared to PCB treatment alone, but the AhR antagonist CH-223191 was protective to a lesser degree. Our data demonstrate the involvement of PCB 126 in macrophage polarization and inflammation, indicating another important role of dioxin-like PCBs in the pathology of atherosclerosis.