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CCL17 and IL-10 as Effectors That Enable Alternatively Activated Macrophages to Inhibit the Generation of Classically Activated Macrophages
Abstract
Classically activated macrophages (CAMphi) have been described as a major effector cell on the host's innate immunities. However, CAMphi are not generated in immunocompromised hosts whose alternatively activated macrophages (AAMphi) predominate. In this study, the mechanism by which AAMphi suppress the ability of resident macrophages (RMphi) to generate CAMphi was investigated. AAMphi were isolated from peritoneal exudates of mice 2 days after third-degree thermal injuries affecting 15% total body surface area. CAMphi were generated from RMphi (peritoneal Mphi from normal mice) through stimulation with CpG DNA, a typical CAMphi inducer. RMphi did not polarize to CAMphi when they were cultured with AAMphi in a dual-chamber Transwell even when supplemented with CpG DNA. In addition, RMphi stimulated with CpG DNA did not convert to CAMphi when they were cultured with the culture fluids of AAMphi (AAMphi Culture-Sup). AAMphi Culture-Sup contained IL-6, IL-10, CCL17, PGE(2), and TGF-beta. Among these, CCL17 and IL-10 inhibited CAMphi generation. The ability of AAMphi Culture-Sup to inhibit CAMphi generation was eliminated when the Culture-Sup was treated with a mixture of mAbs directed against CCL17 and IL-10. These results indicate that CCL17 and IL-10 released from AAMphi inhibit CAMphi generation from RMphi stimulated with CpG DNA.
... Other macrophage surface markers (IL-12, CXCL10, CCL17, and CCR7) were analyzed in this study to further characterize human circulating PBMC phenotypes in T2DM and compare them to macrophage phenotypes since information on human macrophage polarization remains sparse. Several studies have attempted to characterize MMe in the adipose tissue of obese and diabetic subjects [26][27][28]. Ex vivo treatment of monocyte-derived macrophages with glucose, insulin, and palmitate induced a different phenotype than the M1 macrophage phenotype [6]. An in vitro study utilizing human monocytic cell lines classified IL-12, CCR7, and CXCL10 as M1 macrophage markers, and CCL17 as an M2 marker [25]. ...
... Metformin treatment reduced IL-12 levels indicating that it reduces the M1 macrophage-like phenotype. CCL17, which is released from alternatively activated macrophages, serves to prevent a generation of classically activated macrophages, and is considered an M2 marker [26], was inhibited in the PBMCs of T2DM. Thus, our previously published CD163 results [7] and current CCL17 results are consistent with an anti-inflammatory role of CD163 and CCL17 (M2 markers) as they are reduced in T2DM [7]. ...
Background and Objectives: Visceral obesity is associated with chronic low-grade inflammation that predisposes to metabolic syndrome. Indeed, infiltration of adipose tissue with immune–inflammatory cells, including ‘classical’ inflammatory M1 and anti-inflammatory ‘alternative’ M2 macrophages, causes the release of a variety of bioactive molecules, resulting in the metabolic complications of obesity. This study examined the relative expression of macrophage phenotypic surface markers, cholesterol efflux proteins, scavenger receptors, and adenosine receptors in human circulating peripheral blood mononuclear cells (PBMCs), isolated from patients with type 2 diabetes mellitus (T2DM), with the aim to phenotypically characterize and identify biomarkers for these ill-defined cells. Materials and Methodology: PBMCs were isolated from four groups of adults: Normal-weight non-diabetic, obese non-diabetic, newly diagnosed with T2DM, and T2DM on metformin. The mRNA expression levels of macrophage phenotypic surface markers (interleukin-12 (IL-12), C-X-C motif chemokine ligand 10 (CXCL10), C-C motif chemokine ligand 17 (CCL17), and C-C motif receptor 7 (CCR7)), cholesterol efflux proteins (ATP-binding cassette transporter-1 (ABCA1), ATP binding cassette subfamily G member 1 (ABCG1), and sterol 27-hydroxylase (CYP27A)), scavenger receptors (scavenger receptor-A (SR-A), C-X-C motif ligand 16 (CXCL16), and lectin-like oxidized LDL receptor-1 (LOX-1)), and adenosine receptors (adenosine A2A receptor (A2AR) and adenosine A3 receptor (A3R)) were measured using qRT-PCR. Results: In PBMCs from T2DM patients, the expression of IL-12, CCR7, ABCA1, and SR-A1 was increased, whereas the expression of CXCL10, CCL17, ABCG1,27-hydroxylase, LOX-1, A2AR and A3R was decreased. On the other hand, treatment with the antidiabetic drug, metformin, reduced the expression of IL-12 and increased the expression of 27-hydroxylase, LOX-1, CXCL16 and A2AR. Conclusion: PBMCs in the circulation of patients with T2DM express phenotypic markers that are different from those typically present in adipose tissue M1 and M2 macrophages and could be representative of metabolically activated macrophages (MMe)-like cells. Our findings suggest that metformin alters phenotypic markers of MMe-like cells in circulation.
... flammatory cytokines (IL-1, IL-6 and TNF-α), to kill pathogen-infected cells, to express inducible NO synthase, and to initiate an acute immune response. On the other hand, M2-like macrophages are related with wound healing and tissue repair being responsible for the production of IL-1R antagonist, IL-10, and arginase [71][72][73] . As reported by Katakura et al . (2004) [74] , IL-10 acts as immunosuppressor associated with alternatively activated macrophages (also known as M2-like macrophages), being able to block the generation of classically activated macrophages (also known as M1-like macrophages) then inhibiting the progression of the inflammatory to an acute state. Also, Chao Liu and Jiao Sun [75] ...
... As reported by Katakura et al . (2004) [74] , IL-10 acts as immunosuppressor associated with alternatively activated macrophages (also known as M2-like macrophages), being able to block the generation of classically activated macrophages (also known as M1-like macrophages) then inhibiting the progression of the inflammatory to an acute state. Also, Chao Liu and Jiao Sun [75] reported that fish collagen hydrolysates have the potential to polarize human macrophages into M2-like phenotype through inhibition of the expression of M1 macrophage markers (IL1 β and TNF-α) and enhancing the expression of M2 macrophage marker (Arg1 and IL-10). ...
The host immunologic response to a specific material is a critical aspect when considering it for clinical implementation. Collagen and gelatin extracted from marine sources have been proposed as biomaterials for tissue engineering applications, but there is a lack of information in the literature about their immunogenicity. In this work, we evaluated the immune response to collagen and/or gelatin from blue shark and codfish, previously extracted and characterized. After endotoxin evaluation, bone marrow-derived macrophages were exposed to the materials and a panel of pro- and anti-inflammatory cytokines were evaluated both for protein quantification and gene expression. Then, the impact of those materials in the host was evaluated through peritoneal injection in C57BL/6 mice. The results suggested shark collagen as the less immunogenic material, inducing low expression of pro-inflammatory cytokines as well as inducible nitric oxide synthase (encoded by Nos2) and high expression of Arginase 1 (encoded by Arg1). Although shark gelatin appeared to be the material with higher pro-inflammatory expression, it also presents a high expression of IL-10 (anti-inflammatory cytokine) and Arginase (both markers for M2-like macrophages). When injected in the peritoneal cavity of mice, our materials demonstrated a transient recruitment of neutrophil, being almost non-existent after 24 hours of injection. Based on these findings, the studied collagenous materials can be considered interesting biomaterial candidates for regenerative medicine as they may induce an activation of the M2-like macrophage population, which is involved in suppressing the inflammatory processes promoting tissue remodeling.
Statement of significance
Marine-origin biomaterials are emerging in the biomedical arena, namely the ones based in marine-derived collagen/gelatin proposed as cell templates for tissue regeneration. Nevertheless, although the major cause of implant rejection in clinical practice is the host's negative immune response, there is a lack of information in the literature about the immunological impact of these marine collagenous materials. This work aims to contribute with knowledge about the immunologic response to collagen/gelatin extracted from blue shark and codfish skins. The results demonstrated that despite some differences observed, all the materials can induce a macrophage phenotype related with anti-inflammation resolution and then act as immuno-modulators and anti-inflammatory inducible materials.
... The cytokines IL-4, IL-5, and IL-13 promote alternatively activated anti-inflammatory macrophages (25,26). Furthermore, the potent antiinflammatory cytokine IL-10 inhibits the generation of proinflammatory macrophages (27). IL-11 promotes the T helper cell type 2 lymphocyte response and inhibits monocyte/macrophage inflammation (22). ...
... The expression and secretion of CXCL5 are high in the macrophage fraction of adipose tissue, plasma concentrations of the chemokine are abnormally high in obese individuals, and it impairs the action of insulin (32). On the other hand, CCL17, like IL-10, is secreted by alternatively activated macrophages and inhibits the formation of activated, proinflammatory macrophages (27). IL-4 also stimulates T cells to secrete IL-9, a pleiotropic cytokine that affects the development of immunity and inflammation (33). ...
Context
Glucagon-like polypeptide-1 (GLP-1) has anti-inflammatory activity but its effect on increased inflammation in adipose tissue from obese individuals is unclear.
Objective
To determine the effect of GLP-1 infusion on expression of genes encoding cytokines, chemokines and their receptors and activated monocyte/macrophages in adipose tissue stromal vascular fraction (SVF) and peripheral blood monocytes (PBMC) in obese women.
Design
A single blind, randomized, and crossover comparison of infusions with GLP-1 and saline.
Participants
Eleven obese women (BMI > 30 kg/m²) ages 28-63 years were recruited.
Intervention
Infusion of active GLP-1 (7-36) and saline for 4 h on different months. Total RNA and stromal vascular fraction (SVF) were isolated from subcutaneous adipose tissue biopsies and PBMC were isolated from blood collected before and immediately after the infusions.
Main outcomes
Expression of genes controlling inflammation in adipose tissue and proportions of CD14⁺ CD16⁺cells in adipose tissue SVF and PBMC.
Results
Infusion with GLP-1 compared with saline significantly decreased the expression of pro-inflammatory genes TNFα, IL1RN, CCL5, CXCL5, CCR2, CCR4, CCR7 and CXCR2 and significantly increased expression of anti-inflammatory genes IL4, IL5, IL10, IL11, IL13, CCL17 and pro-inflammatory genes IL1A, IL7, IL12B, CCL7, CXCL6 and CXCL10 and significantly increased the proportion of CD14⁺ CD16⁺ cells in adipose tissue SVF (P=0.03).
Conclusions
These data suggest that increased circulating GLP-1 may predominantly attenuate adipose tissue inflammation in obese women. The increased expression of a few pro-inflammatory genes and proportions of activated monocytes may conceivably reflect continued immune response against unresolved adipocyte hypertrophy.
... Type 2 immune responses, which have a key role in the impaired host antibacterial resistance, are predominant in severely burned patients [16][17][18]. Type 2 cytokines (IL-4, IL-5, IL-9, IL-10, and IL-13), produced by type 2 cells (Th2 cells [19], M2Mf [20][21][22], and PMN-II [23]), play a role in the increased susceptibility of severely burned hosts to opportunistic infections. In addition, ILC2 have recently been described to be critical in the development of type 2 immune responses [24][25][26][27]. ...
... Thermal injury was produced with a Bunsen burner for 9 s using an insulated mold with a 4.5 3 5.0 cm window that produced a thirddegree burn on ;25% of the TBSA for a 26 g mouse. The body surface of each animal was calculated by the formula: A = KW2/3 (A = surface area in square centimeters, K = 10, and W = body weight in grams) [21,22]. For fluid resuscitation, 1 ml physiologic saline per mouse was injected i.p. after burn injury. ...
Gut microbiota that invades to the defective mucosal barrier is one of the major sources of infectious complications in severely burned hosts. In this study, a role of group 2 innate lymphoid cells (ILC2) and effects of N-{4-[2,2,2-trifluoro-1-hydroxy-1-(trifluoromethyl)ethyl]phenyl}-2-thiophenesulfonamide (SR3335) on the host antibacterial resistance against infectious complications caused by Enterococcus faecalis oral infection were investigated in burned mice. Retinoic acid receptor-related orphan receptor α (RORα) is a transcription factor required for the development of ILC2, and SR3335 is an RORα-selective inverse agonist. All of burned mice died within 6 d of E. faecalis infection (5 × 10(6) CFU/mouse), whereas 100% of the same mice treated with SR3335 survived. The increased ILC2 and their cytokine products (IL-5 and IL-13) were detected in the lamina propria of mice, 1-7 d after burn injury. However, the number of ILC2 did not increase in the lamina propria of burned mice treated with SR3335. The antibacterial resistance of SCID-beige (SCIDbg) mice to E. faecalis infection was impaired by the inoculation of ILC2. BALB/c, SCIDbg, and polymorphonuclear leukocyte (PMN)-depleted SCIDbg mice were shown to be resistant against E. faecalis infection. However, all Mϕ depleted SCIDbg mice died after the infection. These results indicate that host antibacterial effector Mϕ against enterococcal translocation are influenced by ILC2, increased in the bacterial translocation site of burned mice, and sepsis stemming from E. faecalis oral infection was amazingly mitigated in these mice after treatment with SR3335, an inhibitor of cellular differentiation from an ILC precursor (ILCP) to ILC2.
... The widespread differences observed between M2a MΦs and M2b, M2c, and M2d MΦs may be due in part to M2a MΦs expressing CCL17, which explicitly inhibits activation towards an M1, or pro-inflammatory, state and is not expressed in the other M2 MΦ subtypes [14,53]. Our metabolome profiles correlate with MΦ specialized functions, such as phagocytosis, and are consistent with other published studies [23]. ...
The complexity of macrophage (MΦ) plasticity and polarization states, which include classically activated pro-inflammatory (M1) and alternatively activated anti-inflammatory (M2) MΦ phenotypes, is becoming increasingly appreciated. Within the M2 MΦ polarization state, M2a, M2b, M2c, and M2d MΦ subcategories have been defined based on their expression of specific cell surface receptors, secreted cytokines, and specialized immune effector functions. The importance of immunometabolic networks in mediating the function and regulation of MΦ immune responses is also being increasingly recognized, although the exact mechanisms and extent of metabolic modulation of MΦ subtype phenotypes and functions remain incompletely understood. In this study, proton (1H) nuclear magnetic resonance (NMR) metabolomics was employed to determine the polar metabolomes of M2 MΦ subtypes and to investigate the relationship between aqueous metabolite profiles and M2 MΦ functional phenotypes. Results from this study demonstrate that M2a MΦs are most distinct from M2b, M2c, and M2d MΦ subtypes, and that M2b MΦs display several metabolic traits associated with an M1-like MΦ phenotype. The significance of metabolome differences for metabolites implicated in glycolysis, the tricarboxylic acid (TCA) cycle, phospholipid metabolism, and creatine–phosphocreatine cycling is discussed. Altogether, this study provides biochemical insights into the role of metabolism in mediating the specialized effector functions of distinct M2 MΦ subtypes and supports the concept of a continuum of macrophage activation states rather than two well-separated and functionally distinct M1/M2 MΦ classes, as originally proposed within a classical M1/M2 MΦ framework.
... Higher levels of IL-6 and IL-10 were found to be associated with more severe cases of COVID-19 (28). In addition, a previous study revealed that IL-10 produced by macrophages inhibited the adjacent cells to differentiate into classically activated macrophages, thereby allowing the macrophage population to self-regulate (29). This suggested that the SARS-CoV-2 N protein may regulate the proportion of macrophages of the M1/M2 type in the macrophage population. ...
The present study aimed to explore the immune regulatory function of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) nucleocapsid (N) protein and related mechanisms. In a series of protein activity experiments, SARS-CoV-2 N protein promoted proliferation of three immune cell lines: mouse Raw264.7, human Jurkat and human Raji in a dose-dependent manner. A total of 10 µg/ml N protein could significantly change cell cycle progression of the aforementioned three immune cell lines and could promote quick entry of Raw264.7 cells into G2/M phase from S phase to achieve rapid growth. Additionally, the N protein could also stimulate Raw264.7 cells to secrete a number of proinflammatory factors such as TNF-α, IL-6 and IL-10. RNA sequencing analysis indicated that the N protein changed the expression of certain genes involved in immune-related functions and four important signaling pathways, including JAK-STAT, TNF, NF-κB and MAPK signaling pathways, which suggested that the N protein may not only regulate the expression of genes involved in the process of resisting viral infection in macrophages of the immune system, but also change cellular signal processing.
... 28,34 Furthermore, IL-10 and CCL17 produced by M2-like macrophages was shown to inhibit the generation of conventional inflammatory macrophages. 68 These data, together with the observed differences in MDM phenotypes at baseline ( Figure 2) and upon Mtb infection or lysate treatment (Figure 6), reinforce the concept that macrophages in the lung are likely phenotypically heterogeneous as a result of differences in their exposure history to M-CSF and GM-CSF in particular. Therefore, the predisposition of individual pulmonary macrophages to a pro-or anti-inflammatory response to Mtb infection likely depends on the integration of multiple signaling pathways. ...
Direct recognition of Mycobacterium tuberculosis (Mtb)-infected cells is required for protection by CD4+ T cells. While impaired T cell recognition of Mtb-infected macrophages was demonstrated in mice, data are lacking for humans. Using T cells and monocyte-derived macrophages (MDMs) from individuals with latent Mtb infection (LTBI), we quantified the frequency of memory CD4+ T cell activation in response to autologous MDMs infected with virulent Mtb. We observed robust T cell activation in response to Mtb infection of M1-like macrophages differentiated using GM-CSF, while M2-like macrophages differentiated using M-CSF were poorly recognized. However, non-infected GM-CSF and M-CSF MDMs loaded with exogenous antigens elicited similar CD4+ T cell activation. IL-10 was preferentially secreted by infected M-CSF MDMs, and neutralization improved T cell activation. These results suggest that preferential infection of macrophages with an M2-like phenotype limits T cell-mediated protection against Mtb. Vaccine development should focus on T cell recognition of Mtb-infected macrophages.
... Endstage Dpf2 D/D mice had elevated plasma levels of pro-inflammatory cytokines and chemokines, including TNFRSF11B, CCL22/MDC, CCL17/TARC, CXCL13/BLC, IL-1alpha, CCL11 and BAFF/TNFSF13B ( Figure 1L). Many of these cytokines are secreted by dendritic cells and macrophages (37)(38)(39). In addition, aminotransferase and alkaline phosphatase levels were elevated in plasma from Dpf2 D/D mice, reflecting liver damage ( Figure 1M). ...
During emergency hematopoiesis, hematopoietic stem cells (HSCs) rapidly proliferate to produce myeloid and lymphoid effector cells, a response that is critical against infection or tissue injury. If unresolved, this process leads to sustained inflammation which can cause life-threatening diseases and cancer. We have identified a novel role of Dpf2 in inflammation. Dpf2 is a defining subunit of the hematopoietic-specific BAF (SWI/SNF) chromatin-remodeling complex, and it is mutated in multiple cancers and neurological disorders. We uncover that hematopoietic-specific Dpf2 knock-out mice develop leukopenia, severe anemia and lethal systemic inflammation characterized by histiocytic and fibrotic tissue infiltration, resembling a clinical hyper-inflammatory state. Dpf2 loss impairs the polarization of macrophages responsible for tissue repair, induces unrestrained activation of T helper cells, and generates an emergency-like state of HSC hyperproliferation and myeloid-biased differentiation. Mechanistically, Dpf2 deficiency results in the loss of the BAF catalytic subunit Brg1 from Nrf2-controlled enhancers, impairing the anti-oxidant and anti-inflammatory transcriptional response needed to modulate inflammation. Finally, pharmacological reactivation of Nrf2 can suppress the inflammation-mediated phenotypes and lethality of Dpf2Δ/Δ mice. Our work establishes the essential role of the Dpf2/BAF complex in licensing Nrf2-dependent gene expression in HSCs and immune effector cells to prevent chronic inflammation.
... Both MDC and TARC are specific ligands for C-C chemokine receptor type 4, a receptor expressed on myeloid cells and lymphocytes but predominately on Th2 cells 26 . In addition to activating the Th2 response, TARC inhibits generation of classically activated macrophages resulting in impaired innate immunity 27 . Our findings suggest a potentially novel role for RIC in the modulation of T cell mediated innate immunity, as post traumatic shift from Th1 towards an anti-inflammatory Th2 phenotype contributes to susceptibility to nosocomial infections 6 and development of sepsis and poorer outcomes. ...
Abstract Resuscitation induced ischemia/reperfusion predisposes trauma patients to systemic inflammation and organ dysfunction. We investigated the effect of remote ischemic conditioning (RIC), a treatment shown to prevent ischemia/reperfusion injury in experimental models of hemorrhagic shock/resuscitation, on the systemic immune-inflammatory profile in trauma patients in a randomized trial. We conducted a prospective, single-centre, double-blind, randomized, controlled trial involving trauma patients sustaining blunt or penetrating trauma in hemorrhagic shock admitted to a Level 1 trauma centre. Patients were randomized to receive RIC (four cycles of 5-min pressure cuff inflation at 250 mmHg and deflation on the thigh) or a Sham intervention. The primary outcomes were neutrophil oxidative burst activity, cellular adhesion molecule expression, and plasma levels of myeloperoxidase, cytokines and chemokines in peripheral blood samples, drawn at admission (pre-intervention), 1 h, 3 h, and 24 h post-admission. Secondary outcomes included ventilator, ICU and hospital free days, incidence of nosocomial infections, 24 h and 28 day mortality. 50 eligible patients were randomized; of which 21 in the Sham group and 18 in the RIC group were included in the full analysis. No treatment effect was observed between Sham and RIC groups for neutrophil oxidative burst activity, adhesion molecule expression, and plasma levels of myeloperoxidase and cytokines. RIC prevented significant increases in Th2 chemokines TARC/CCL17 (P
... 28,34 Furthermore, IL-10 and CCL17 produced by M2-like macrophages was shown to inhibit the generation of conventional inflammatory macrophages. 68 These data, together with the observed differences in MDM phenotypes at baseline ( Figure 2) and upon Mtb infection or lysate treatment (Figure 6), reinforce the concept that macrophages in the lung are likely phenotypically heterogeneous as a result of differences in their exposure history to M-CSF and GM-CSF in particular. Therefore, the predisposition of individual pulmonary macrophages to a pro-or anti-inflammatory response to Mtb infection likely depends on the integration of multiple signaling pathways. ...
... 7,8) CCL17, a specific ligand of CC chemokine receptor (CCR4), was originally implicated in the preferential attraction of Th2 lymphocytes. [9][10][11][12][13] There has been considerable evidence that CCL17 is expressed at elevated levels in many inflammatory conditions. [14][15][16] Studies have found that the application of CCL17 drives inflammatory pain in a COX-dependent manner, while treatment with anti-CCL17 blocks the effects. ...
CCL17 (C-C motif chemokine ligand 17), an important chemokine, plays a vital role in regulating immune balance in the central nervous system. In this study, we explored the potential roles of CCL17 in a rat postoperative pain model and that of blocking CCL17 in the prevention of postoperative pain in rats. A right plantar incision in rat was used as a model of postoperative pain. A behavioral change was measured preoperatively and postoperatively using mechanical withdrawal thresholds and thermal withdrawal latency. CCL17 and its upstream Jmjd-3 mRNA levels in the spinal cord were detected using real-time PCR, CCL17 levels in the serum were measured using ELISA, and the expression of IRF4, which interacts with Jmjd-3, was detected by immunohistochemistry staining. After that, rats were intraperitoneally injected with either anti-CCL17mAb or GSK-J4 (the Jmjd3 inhibitor) to evaluate the protective effects of blocking CCL17 on postoperative pain. We found that CCL17 and Jmjd-3 were significantly increased in the spinal cords of the postoperative pain rat, consistent with changes in hyperalgesia. In addition, our results showed that the mechanical and thermal allodynia was significantly ameliorated using anti-CCL17mAb or GSK-J4. Moreover, we found that anti-CCL17mAb or GSK-J4 treatment decreased c-fos expression in response to peripheral stimulation. Finally, our preliminary exploration found that anti-CCL17mAb or GSK-J4 had a protective effect on tissue damage. These findings indicated that high expression of CCL17 played a critical role in postoperative pain induced by plantar incision and that CCL17 blockade may serve as an effective approach to managing postoperative pain.
... For instance, TGF-β inhibits T-cell activation and suppresses cytotoxic T lymphocytes (CTL) and Th1 and Th2 lymphocyte differentiation [34]. IL-10 negatively regulates the innate and protective Th1 antifungal activity [31], and macrophage-derived IL-10 can inhibit the differentiation of M1 macrophages [35]. By limiting the excessive inflammatory response, IL-10 can protect tissue from damage. ...
: Talaromyces marneffei, a dimorphic fungus, exhibits temperature-dependent growth, existing in a filamentous form at 25 °C and as a yeast at 37 °C. Several studies have highlighted the important roles of macrophages in defense against T. marneffei infection. However, the immune responses to the interaction of macrophages with T. marneffei cells during phase transition require further investigation. This study reports the expression of cytokine profiles in human THP-1 cells during infection by T. marneffei. THP-1 cells were infected with T. marneffei conidia at different multiplicity of infections (MOIs). Surviving conidia transformed into yeasts after phagocytosis by macrophages, and the number of yeasts gradually increased over 36 h. The transcription and secretion levels of pro- and anti-inflammatory cytokines were examined at different times by qRT-PCR and ELISA. Transcription levels of IL-8, IL-12, IL-1β, and TNF-α increased significantly at 12 or 24 h and then slightly decreased at 36 h. In contrast, the transcription levels of IL-6, IL-10, and TGF-β gradually increased at all MOIs. The levels of IL-6 and IL-10 secretion corresponded to their levels of transcription. These results indicated that as the number of intracellular yeasts increased, the infected macrophages first underwent slight M1 polarization before shifting to M2 polarization. This polarization transition was confirmed by the fungicidal ability and the expression of macrophage surface markers. By inducing the M2-type polarization of macrophages, the intracellular T. marneffei cells can successfully evade the immune response. Our study provides a novel insight into the immune characterization during the transition of T. marneffei infection and could further contribute to possible diagnostic and therapeutic interventions for this infection.
... Among these, thymus and activation regulated chemokine (TARC)/CCL17 and smallinducible cytokine B10 (IP-10)/CXCL10) cytokines were the only additional factors enriched in centenarians, thus indicating them as additional key mediators of successful ageing [21]. The two cytokines are mainly released from myeloid cells: IP-10 is an effector cytokine enhancing host performance against infectious disease [22]; TARC is implicated in the T cell recruitment and produced from M2 macrophages as effector molecule that inhibit M1 inflammatory macrophages generation [23]. However only TARC seems to be responsible for M2 polarizing action of LLIs plasma as TARC-, but not IP-10-blocking antibodies were able to rescue the M2 enrichment in monocyte-derived macrophage compartment [21]. ...
Recent discoveries have shed light on the participation of the immune system in the physio pathology of the cardiovascular system underpinning the importance of keeping the balance of the first to preserve the latter. Aging, along with other risk factors, can challenge such balance triggering the onset of cardiovascular diseases.
Among several mediators ensuring the proper cross-talk between the two systems, bactericidal/permeability-increasing fold-containing family B member 4 (BPIFB4) has been shown to have a pivotal role, also by sustaining important signals such as eNOS and PKC-alpha.
In addition, the Longevity-associated variant (LAV), which is an haplotype allele in BPIFB4 characterized by 4 missense polymorphisms, enriched in homozygosity in Long Living Individuals (LLIs), has been shown to be efficient, if administered systemically through gene therapy, in improving many aspects of cardiovascular diseases (CVDs). This occurs mainly through a fine immune system remodeling across: 1) a M2 macrophage polarizing effect, 2) a favorable redistribution of the circulating monocyte cell subsets and 3) the reduction of T-cell activation. Furthermore, LAV-BPIFB4 treatment induced a desirable recovery of the inflammatory balance by mitigating the pro-inflammatory factor levels and enhancing the anti-inflammatory boost through a mechanism that is partially dependent on SDF-1/CXCR4 axis.
Importantly, the remarkable effects of LAV-BPIFB4 treatment, which translates in increased BPIFB4 circulating levels, mirror what occurs in long-living individuals (LLIs) in whom the high circulating levels of BPIFB4 are protective from age-related and CVDs and emphasize the reason why LLIs are considered a model of successful aging. Here, we review the mechanisms by which LAV-BPIFB4 exerts its immunomodulatory activity in improving the cardiovascular-immune system dialogue that might strengthen its role as a key mediator in CVDs.
... Recent studies showed that LPS-tolerant monocytes/macrophages express low levels of M1-associated genes and high levels of M2-associated genes such as IL-10 (20). IL-10 produced by macrophages prevents adjacent cells from becoming classically activated M1 macrophages, allowing cells to self-regulate (23). IL-10 plays a role in limiting the host immune response to pathogens and in maintaining tissue homeostasis by reducing the immunological response of the host to infection, thereby preventing damage to the host. ...
Activation of macrophages by Leptospira spp. infection is thought to be involved in the pathogenesis of leptospirosis. To evaluate the innate macrophage responses to Leptospira spp., specifically pathogenic versus nonpathogenic Leptospira spp., we characterized the entire transcriptome-wide alterations in infected macrophages.
... However, the inclusion of cellular covariates in the assessment of genetic associations led to the identification of three novel proteins with pQTLs, while abrogated the signal for 10 proteins which would have otherwise led to positive hits. This could potentially be explained by the fact that all 13 proteins are expressed by specific circulating immune cell populations [73][74][75][76][77][78][79][80][81][82]. However, the interactions between these proteins, bloodcell populations, and genetic variants are less obvious to interpret, as both direct and indirect effects or co-occurring mechanisms could be involved. ...
Background
Blood plasma proteins play an important role in immune defense against pathogens, including cytokine signaling, the complement system, and the acute-phase response. Recent large-scale studies have reported genetic (i.e., protein quantitative trait loci, pQTLs) and non-genetic factors, such as age and sex, as major determinants to inter-individual variability in immune response variation. However, the contribution of blood-cell composition to plasma protein heterogeneity has not been fully characterized and may act as a mediating factor in association studies.
Methods
Here, we evaluated plasma protein levels from 400 unrelated healthy individuals of western European ancestry, who were stratified by sex and two decades of life (20–29 and 60–69 years), from the Milieu Intérieur cohort. We quantified 229 proteins by Luminex in a clinically certified laboratory and their levels of variation were analyzed together with 5.2 million single-nucleotide polymorphisms. With respect to non-genetic variables, we included 254 lifestyle and biochemical factors, as well as counts of seven circulating immune cell populations measured by hemogram and standardized flow cytometry.
Results
Collectively, we found 152 significant associations involving 49 proteins and 20 non-genetic variables. Consistent with previous studies, age and sex showed a global, pervasive impact on plasma protein heterogeneity, while body mass index and other health status variables were among the non-genetic factors with the highest number of associations. After controlling for these covariates, we identified 100 and 12 pQTLs acting in cis and trans , respectively, collectively associated with 87 plasma proteins and including 19 novel genetic associations. Genetic factors explained the largest fraction of the variability of plasma protein levels, as compared to non-genetic factors. In addition, blood-cell fractions, including leukocytes, lymphocytes, monocytes, neutrophils, eosinophils, basophils, and platelets, had a larger contribution to inter-individual variability than age and sex and appeared as confounders of specific genetic associations. Finally, we identified new genetic associations with plasma protein levels of five monogenic Mendelian disease genes including two primary immunodeficiency genes (Ficolin-3 and FAS).
Conclusions
Our study identified novel genetic and non-genetic factors associated to plasma protein levels which may inform health status and disease management.
... Among these, thymus and activation regulated chemokine (TARC)/CCL17 and smallinducible cytokine B10 (IP-10)/CXCL10) cytokines were the only additional factors enriched in centenarians, thus indicating them as additional key mediators of successful ageing [21]. The two cytokines are mainly released from myeloid cells: IP-10 is an effector cytokine enhancing host performance against infectious disease [22]; TARC is implicated in the T cell recruitment and produced from M2 macrophages as effector molecule that inhibit M1 inflammatory macrophages generation [23]. However only TARC seems to be responsible for M2 polarizing action of LLIs plasma as TARC-, but not IP-10-blocking antibodies were able to rescue the M2 enrichment in monocyte-derived macrophage compartment [21]. ...
... It is activated by such cytokines as TNF, IL-4, and IL-13 and interacts with its receptor CCR4. CCL17 induces chemotaxis of T helper 2 cells, regulatory T cells, basophils, and macrophages and inhibits TLR2 and TLR4 [215]. CCL20 is an adipochemokine whose expression is modulated by an anatomic arrangement of adipose tissue and by obesity severity. ...
The main functions of adipose tissue are thought to be storage and mobilization of the body’s energy reserves, active and passive thermoregulation, participation in the spatial organization of internal organs, protection of the body from lipotoxicity, and ectopic lipid deposition. After the discovery of adipokines, the endocrine function was added to the above list, and after the identification of crosstalk between adipocytes and immune cells, an immune function was suggested. Nonetheless, it turned out that the mechanisms underlying mutual regulatory relations of adipocytes, preadipocytes, immune cells, and their microenvironment are complex and redundant at many levels. One possible way to elucidate the picture of adipose-tissue regulation is to determine genetic variants correlating with obesity. In this review, we examine various aspects of adipose-tissue involvement in innate immune responses as well as variants of immune-response genes associated with obesity.
... These promising, yet less-than-satisfactory, results support the continued pursuit of immune-based technologies to treat these cancers. TAMs can function as gatekeepers within the TNBC immune microenvironment by suppressing CTL function and number through ICBresistant mechanisms (63,64), and our findings raise the possibility that anti-Notch therapies can reduce the load of immunosuppressive TAMs in TNBC, promoting immunoconversion to ICB sensitivity. Dysregulated Notch signaling has long been recognized as an important driver in multiple cancer types (3). ...
Significance
There is clear and irrefutable evidence that the Notch developmental signaling pathway is an important driver in many cancers, specifically in breast cancer. However, because of the ubiquity of this pathway in normal tissues, therapeutics targeting Notch have been associated with significant deleterious side effects. Herein, we provide evidence that USP9x selectively drives Notch in breast cancer and that therapeutic USP9x inhibition prevents tumor progression without collateral damage to normal tissues.
... M2 macrophages promote wound healing, angiogenesis and tissue remodelling (Lievense et al., 2013). They produce anti-inflammatory cytokines like IL10, chemokines like CCL17 (Katakura et al., 2004), CCL22 and CCL24 (Owen and Mohamadzadeh, 2013), produce MMPs, express the mannose (CD206), and scavenger (CD163) receptors and are poor at antigen presentation (Lievense et al., 2013). Hypoxia in the TME induces M2 macrophages to produce pro-angiogenic factors like VEGF-A (Lievense et al., 2013, White et al., 2004. ...
Mesothelioma is a rare cancer usually affecting the pleura. It is characteristically associated with inhalation of asbestos fibres and accounts for 1% of cancers in the United Kingdom. Median survival remains poor at 4-18 months despite treatment.
Immunotherapy has established itself as an important treatment option in many solid tumours where survival benefit has been shown to be associated with CD8 infiltration. In mesothelioma, there are 3 small studies that suggest that CD8 infiltration may confer survival benefit.
Here, a systematic assessment was undertaken of the prognostic and predictive value of infiltrating adaptive and innate immune cells in a large cohort of patients with advanced mesothelioma. A tissue microarray from 302 samples was constructed. Markers of adaptive immune response CD4+ T helper and CD8+ Cytotoxic T cells, FOXP3+Tregs, CD45RO+Memory T cells and B-cells (CD20+), and innate immune response; macrophages (CD68+), natural killer cells (CD56+) and neutrophils (NP57+) were evaluated.
Surprisingly, CD8+ tumour infiltrating lymphocytes (TILs) did not predict for outcome. On multivariate analysis a high CD4+, high CD20+ and low NP57+ count were linked to better outcome in the epithelioid tumours. A low FOXP3+ predicted for good outcome in both epithelioid and non-epithelioid tumours.
Next, multiplex immunohistochemistry was utilised to further evaluate CD4+ and CD8+ T cell subtypes. This established the presence of MHC class II expression on epithelioid mesothelioma tumour cells and confirmed that some CD4+ T cell subsets (Tissue resident memory cells and T follicular helper cells), were associated with better outcome in epithelioid mesothelioma. The intriguing question of why CD4 + T cells function as the outcome determining immune effectors in mesothelioma, remains to be determined.
Mesothelioma-associated pleural fluid was evaluated to determine its utility as a surrogate for immune events in the solid tumour by transcriptomic analysis. T cells in the pleural fluid exhibited a phenotype characteristic of quiescent/naive cells.
... While the focus of the current work was on T cell populations and associated cytokines, these play a crucial regulatory role in the activation of antigen-presenting cells such as infiltrating macrophages and resident microglia. Th 1 -derived cytokines activate M1-type microglia [67], while Th 2 -derived cytokines activate M2-type macrophages and microglia [68,69]. M1 activated macrophages and microglia have been found in early and active demyelinating lesions, suggesting their direct role in tissue destruction [70][71][72]. ...
Human adipose-derived stem cells (ASCs) show immense promise for treating inflammatory diseases, attributed primarily to their potent paracrine signaling. Previous investigations demonstrated that short-term Rapamycin preconditioning of bone marrow-derived stem cells (BMSCs) elevated secretion of prostaglandin E2, a pleiotropic molecule with therapeutic effects in the experimental autoimmune encephalomyelitis (EAE) model of multiple sclerosis (MS), and enhanced immunosuppressive capacity in vitro. However, this has yet to be examined in ASCs. The present study examined the therapeutic potential of short-term Rapamycin-preconditioned ASCs in the EAE model. Animals were treated at peak disease with control ASCs (EAE-ASCs), Rapa-preconditioned ASCs (EAE-Rapa-ASCs), or vehicle control (EAE). Results show that EAE-ASCs improved clinical disease scores and elevated intact myelin compared to both EAE and EAE-Rapa-ASC animals. These results correlated with augmented CD4+ T helper (Th) and T regulatory (Treg) cell populations in the spinal cord, and increased gene expression of interleukin-10 (IL-10), an anti-inflammatory cytokine. Conversely, EAE-Rapa-ASC mice showed no improvement in clinical disease scores, reduced myelin levels, and significantly less Th and Treg cells in the spinal cord. These findings suggest that short-term Rapamycin preconditioning reduces the therapeutic efficacy of ASCs when applied to late-stage EAE.
... Classically activated Mφ (M1Mφ), which are IL-12-producing and IL-10-non-producing Mφ, are major effector cells in innate immune responses 25,26 . In contrast, alternatively activated Mφ (M2Mφ) produce IL-10 but not IL-12 and have a reduced capacity to kill tumor cells 27,28 . These latter cells suppressed the tumoricidal activities of other type 1 tumor-killing cells (M1Mφ, CTLs, NK cells and DCs) 29 . ...
Anti-programmed cell death-1 (PD-1) antibodies has been approved to treat HCC. Some PD-1 ligands (PD–L1 and PD–L2) negative tumors respond to treatment of anti-PD-1 antibodies, and this fact may be caused by the expression of PD-1 ligands on non-tumor cells. PD–L1 was recently found to be expressed on CD14⁺ cells from cancer patients. We investigate PD-1 ligands expression on CD14⁺ cells of patients with HCC and the role of CD14⁺ cells in an antitumor response. In this study, 87 patients diagnosed with HCC were enrolled. CD14⁺ cells from patients with HCC expressed PD–L1 (4.5–95.5%) and PD–L2 (0.2–95.0%). According to cut-off values, we classified patients as those either with PD–L1⁺PD–L2⁺CD14⁺ cells or other types of CD14⁺ cells. The overall survival of patients with PD–L1⁺PD–L2⁺CD14⁺ cells was shorter than that of patients with other types of CD14⁺ cells (p = 0.0023). PD–L1⁺PD–L2⁺CD14⁺ cells produced IL-10 and CCL1, and showed little tumoricidal activity against HepG2 cells. The tumoricidal activity of CD8⁺ cells from patients with PD–L1⁺PD–L2⁺CD14⁺ cells were suppressed by co-cultivation with CD14⁺ cells from the syngeneic patient. Furthermore, anti-PD-1 antibody restored their tumoricidal activity of CD8⁺ cells. In conclusion, some patients with HCC have PD–L1⁺PD–L2⁺CD14⁺ cells that suppress their antitumor response. These inhibitory functions of CD14⁺ cells may be associated with a poor prognosis in these patients.
... Conversely, we found that CD4 ϩ T cells from persistently infected livers were unable to control bacterial replication. IL-10 can alternatively activate and induce M2-like macrophages, a phenotype we ascribe to the liver macrophage subset, potentially leading to increased bacterial numbers (23,(63)(64)(65). Notably, while we didn't observe control of intracellular bacteria, neither did we find that intracellular bacterial burdens increased. ...
The intracellular bacterial pathogen Salmonella is able to evade the immune system and persist within the host. In some cases, these persistent infections are asymptomatic for long periods and represent a significant public health hazard as potential chronic carriers, yet the mechanisms that control persistence are incompletely understood. Using a mouse model of chronic typhoid fever combined with MHC class II tetramers to interrogate endogenous, Salmonella -specific CD4+ helper T cells, we show certain host microenvironments may favorably contribute to a pathogen's ability to persist in vivo . We demonstrate that the environment in the hepatobiliary system may contribute to the persistence of Salmonella Typhimurium through liver resident immunoregulatory CD4+ helper T cells, alternatively activated macrophages, and impaired bactericidal activity. This contrasts with lymphoid organs, such as the spleen and mesenteric lymph nodes, where these same cells appear to have a greater capacity for bacterial killing, which may contribute to control of bacteria in these organs. We also find that, following an extended period of infection of greater than two years, the liver appeared to be the only site that harbored Salmonella . This work establishes a potential role for non-lymphoid organ immunity in regulating chronic bacterial infections and provides further evidence for the hepatobiliary system as the site of chronic Salmonella infection.
... Additionally, the identified GO processes (Fig. 4B) heavily favored the tissuereparative, pro-fibrotic, pro-angiogenic, and phagocytic functions [1,[25][26][27]. Additionally, the most upregulated genes, CCL17, CCl22 and CCL18, support the generally accepted role that M2a MΦs play in cellular proliferation, ECM remodeling, and tissue repair [66,67]. ...
Macrophages (MΦs) display remarkable plasticity and the ability to activate diverse responses to a host of intracellular and external stimuli. Despite extensive characterization of M1 MΦs and a broad set of M2 MΦs, comprehensive characterization of metabolic shifts driving MΦ activation remains. Herein, we utilized an ex vivo model to produce six MΦ functional phenotypes. Isolated CD14+ PBMCs were differentiated into resting M0 MΦs, and then polarized into M1 (IFN-γ/LPS), M2a (IL-4/IL-13), M2b (IC/LPS), M2c (IL-10), and M2d (IL-6/LIF) MΦs. The MΦs were profiled using a bioanalyte matrix of four cell surface markers, ∼50 secreted proteins, ∼800 expressed myeloid genes, and ∼450 identified metabolites relative to M0 MΦs. Signal protein and expressed gene profiles grouped the MΦs into inflammatory (M1 and M2b) and wound resolution (M2a, M2c, and M2d) phenotypes; however, each had a unique metabolic profile. While both M1 and M2b MΦs shared metabotype profiles consistent with an inflammatory signature; key differences were observed in the TCA cycle, FAO and OXPHOS. Additionally, M2a, M2c and M2d MΦs all profiled as tissue repair MΦs; however, metabotype differences were observed in multiple pathways including hexosamine, polyamine, and fatty acid metabolism. These metabolic and other key functional distinctions suggest phagocytic and proliferative functions for M2a MΦs, and angiogenesis and ECM assembly capabilities for M2b, M2c and M2d MΦs. By integrating metabolomics into a systems analysis of MΦ phenotypes, we provide the most comprehensive map of MΦ diversity to date, along with the global metabolic shifts driving MΦ functional plasticity in these phenotypes.
Summary Sentence
Macrophage functional plasticity of six macrophage phenotypes correlates with unique distinctions in cell-surface marker expression, signal protein secretion, transcriptomics profiles, and metabolic processes.
... These two cells are sufficiently proximate to each other so that both cells compete on IL-4 signaling/uptake. CCL24 can be produced by IL-13R1 + hemato-poietic cells such as macrophages, which express both the type 1 and type 2 IL-4Rs (45,46). We hypothesize that in the absence of Il13ra1, macrophages produce CCL24 in response to IL-4 via the type 1 IL-4R. ...
IL-13 and IL-4 are potent mediators of type 2–associated inflammation such as those found in atopic dermatitis (AD). IL-4 shares overlapping biological functions with IL-13, a finding that is mainly explained by their ability to signal via the type 2 IL-4 receptor (R), which is composed of IL-4Rα in association with IL-13Rα1. Nonetheless, the role of the type 2 IL-4R in AD remains to be clearly defined. Induction of two distinct models of experimental AD in Il13ra1−/− mice, which lack the type 2 IL-4R, revealed that dermatitis, including ear and epidermal thickening, was dependent on type 2 IL-4R signaling. Expression of TNF-α was dependent on the type 2 IL-4R, whereas induction of IL-4, IgE, CCL24, and skin eosinophilia was dependent on the type 1 IL-4R. Neutralization of IL-4, IL-13, and TNF-α as well as studies in bone marrow–chimeric mice revealed that dermatitis, TNF-α, CXCL1, and CCL11 expression were exclusively mediated by IL-13 signaling via the type 2 IL-4R expressed by nonhematopoietic cells. Conversely, induction of IL-4, CCL24, and eosinophilia was dependent on IL-4 signaling via the type 1 IL-4R expressed by hematopoietic cells. Last, we pharmacologically targeted IL-13Rα1 and established a proof of concept for therapeutic targeting of this pathway in AD. Our data provide mechanistic insight into the differential roles of IL-4, IL-13, and their receptor components in allergic skin and highlight type 2 IL-4R as a potential therapeutic target in AD and other allergic diseases such as asthma and eosinophilic esophagitis.
... Increasing data from several studies brought up the evidence that in inflammatory Th2 responses, like allergic conditions or inflammation in helminth infection with pathology, IL-10 has a down-regulatory role and counteracts Th2 responses (39 -41). IL-10 is also produced by Tr-1 cells (8,13,42,43), by Th3 cells (44,45), and by macrophages (46). In helminth infections, including schistosomiasis (47), onchocerciasis, and lymphatic filariasis, IL-10 is associated with several regulatory mechanisms that favor high parasite loads but mild disease. ...
... 11 In the literature, IL-10 has been reported to play a critical function in both the classical and alternative macrophage activation processes, although its roles vary and are sometimes opposing. 12 However, given the high degree of macrophage plasticity and propensity for phenotype switching in response to changing microenvironmental conditions, this contradictory evidence should not be interpreted as inconsistent or conflicting. 10 More recently, new metabolomic evidence for IL-10's role in macrophage phenotype switching has Wound healing and fibrogenesis. ...
Significance: Fibrosis is the endpoint of chronic disease in multiple organs, including the skin, heart, lungs, intestine, liver, and kidneys. Pathologic accumulation of fibrotic tissue results in a loss of structural integrity and function, with resultant increases in morbidity and mortality. Understanding the pathways governing fibrosis and identifying therapeutic targets within those pathways is necessary to develop novel antifibrotic therapies for fibrotic disease.
Recent Advances: Given the connection between inflammation and fibrogenesis, Interleukin-10 (IL-10) has been a focus of potential antifibrotic therapies because of its well-known role as an anti-inflammatory mediator. Despite the apparent dissimilarity of diseases associated with fibrotic progression, pathways involving IL-10 appear to be a conserved molecular theme. More recently, many groups have worked to develop novel delivery tools for recombinant IL-10, such as hydrogels, and cell-based therapies, such as ex vivo activated macrophages, to directly or indirectly modulate IL-10 signaling.
Critical Issues: Some efforts in this area, however, have been stymied by IL-10's pleiotropic and sometimes conflicting effects. A deeper, contextual understanding of IL-10 signaling and its interaction with effector cells, particularly immune cells, will be critical to future studies in the field.
Future Directions: IL-10 is clearly a gatekeeper of fibrotic/antifibrotic signaling. The development of novel therapeutics and cell-based therapies that capitalize on targets within the IL-10 signaling pathway could have far-reaching implications for patients suffering from the consequences of organ fibrosis.
... In addition to its effects on neutrophil hypoxic survival, IL-4 suppressed LPS-induced expression of the proinflammatory genes CCL2 , CCL3 , TNF, and IL1B, previously linked to HIF-1a activity ( Figure 2J) (18). Conversely, IL-4 induced CCL17 ( Figure 2K), a chemokine associated with Treg recruitment, M2polarized macrophages, and tumorassociated neutrophils (19)(20)(21)(22). IL-4 did not influence baseline reactive oxidative species production or neutrophil respiratory burst after stimulation with N-formyl-met-leuphe under normoxic or hypoxic conditions ( Figure 2L). ...
Rationale: Acute respiratory distress syndrome is defined by the presence of systemic hypoxia and consequent on disordered neutrophilic inflammation. Local mechanisms limiting the duration and magnitude of this neutrophilic response remain poorly understood.
Objectives: To test the hypothesis that during acute lung inflammation tissue production of proresolution type 2 cytokines (IL-4 and IL-13) dampens the proinflammatory effects of hypoxia through suppression of HIF-1α (hypoxia-inducible factor-1α)-mediated neutrophil adaptation, resulting in resolution of lung injury.
Methods: Neutrophil activation of IL4Ra (IL-4 receptor α) signaling pathways was explored ex vivo in human acute respiratory distress syndrome patient samples, in vitro after the culture of human peripheral blood neutrophils with recombinant IL-4 under conditions of hypoxia, and in vivo through the study of IL4Ra-deficient neutrophils in competitive chimera models and wild-type mice treated with IL-4.
Measurements and Main Results: IL-4 was elevated in human BAL from patients with acute respiratory distress syndrome, and its receptor was identified on patient blood neutrophils. Treatment of human neutrophils with IL-4 suppressed HIF-1α–dependent hypoxic survival and limited proinflammatory transcriptional responses. Increased neutrophil apoptosis in hypoxia, also observed with IL-13, required active STAT signaling, and was dependent on expression of the oxygen-sensing prolyl hydroxylase PHD2. In vivo, IL-4Ra–deficient neutrophils had a survival advantage within a hypoxic inflamed niche; in contrast, inflamed lung treatment with IL-4 accelerated resolution through increased neutrophil apoptosis.
Conclusions: We describe an important interaction whereby IL4Rα-dependent type 2 cytokine signaling can directly inhibit hypoxic neutrophil survival in tissues and promote resolution of neutrophil-mediated acute lung injury.
... The resulting adipocyte hypertrophy and concurrent release of chemokines, like MCP-1, by adipocytes, facilitates the recruitment of C-C motif chemokine receptor (CCR)2positive monocytes to the expanding fat. Upon arrival, these monocytes differentiate into macrophages, and become activated cells that produce proinflammatory factors such as TNFα, IL-6, and in mice, iNOS (225,484). Lumeng and colleagues proposed that these proinflammatory macrophages may be classically activated or M1 macrophages (280). While this was a plausible hypothesis at the time, it is now clear that the proinflammatory ATM in obese adipose tissue of mice and humans are not similar to M1 macrophages. ...
The objective of this comprehensive review is to summarize and discuss the available evidence of how adipose tissue inflammation affects insulin sensitivity and glucose tolerance. Low‐grade, chronic adipose tissue inflammation is characterized by infiltration of macrophages and other immune cell populations into adipose tissue, and a shift toward more proinflammatory subtypes of leukocytes. The infiltration of proinflammatory cells in adipose tissue is associated with an increased production of key chemokines such as C‐C motif chemokine ligand 2, proinflammatory cytokines including tumor necrosis factor α and interleukins 1β and 6 as well as reduced expression of the key insulin‐sensitizing adipokine, adiponectin. In both rodent models and humans, adipose tissue inflammation is consistently associated with excess fat mass and insulin resistance. In humans, associations with insulin resistance are stronger and more consistent for inflammation in visceral as opposed to subcutaneous fat. Further, genetic alterations in mouse models of obesity that reduce adipose tissue inflammation are—almost without exception—associated with improved insulin sensitivity. However, a dissociation between adipose tissue inflammation and insulin resistance can be observed in very few rodent models of obesity as well as in humans following bariatric surgery‐ or low‐calorie‐diet‐induced weight loss, illustrating that the etiology of insulin resistance is multifactorial. Taken together, adipose tissue inflammation is a key factor in the development of insulin resistance and type 2 diabetes in obesity, along with other factors that likely include inflammation and fat accumulation in other metabolically active tissues. © 2019 American Physiological Society. Compr Physiol 9:1‐58, 2019.
... It is activated by cytokines such as TNF, IL-4 and IL-13 and contacts with chemokine C-C of receptor 4 (CCR4). CCL17 performs a number of functions, including Th2-and regulatory T-cells migration, TLR2 and TLR4 receptors inhibition [59]. General impact of the aforesaid gene in allergic diseases is shown. ...
Recent studies have shown that chemokines play an important role in the development of chronic inflammation in adipose tissue, obesity pathogenesis, glucose intolerance and type 2 diabetes. It has also been revealed that some SNPs in chemokine genes are associated with obesity, insulin resistance, type 2 diabetes and diabetes complications in different ethnic groups. The aim of this study was to determine the associations between SNPs in chemokine genes and type 2 diabetes in participants of Tatar ethnic group, living in Bashkortostan. Case–control and cross-sectional study were included in our study design. Five SNPs were genotyped in 440 type 2 diabetes (160 men and 280 women), 58.8 ± 9.2 years old (mean ± SD), BMI 29.3 ± 3.9 kg/m² (mean ± SD) patients of Tatar ethnicity, and a control group of 500 Tatars (180 men and 320 women), 55.2 ± 11.6 years old (mean ± SD), BMI 25.9 ± 4.3 kg/m² (mean ± SD). The SNPs rs6749704 in CCL20 [odds ratio (OR) = 2.77 (95% CI 1.81–4.25), р = 0.0001], rs2107538 in CCL5 [odds ratio (OR) = 1.80 (95% CI 1.46–2.22), p = 0.0001] were significantly associated with type 2 diabetes. Regression analysis revealed that rs1696941 in CCL11 was associated with the onset age and duration of type 2 diabetes as well as with HbA1c level (p = 0.034, p = 0.036 and p = 0.0054, respectively). The SNPs rs223828 in CCL17 and rs6749704 in CCL20 were correlated with obesity as estimated by BMI (p = 0.0004, p = 0.029, respectively). Rs223828 in CCL17 revealed the association with postprandial glucose level (p = 0.024) and HbA1c (p = 0.008). These data demonstrate that variants of chemokine genes are associated with type 2 diabetes and obesity of Tatar ethnic group inhabiting Bashkortostan Republic. Novel associations of the polymorphic loci in CCL20 (rs6749704) and CCL5 (rs2107538) genes with type 2 diabetes had been identified as a result of the conducted research.
... Additionally, although the difference was not statistically significant, this combination treatment tended to elevate the frequency of tetramer + cells to a greater extent than the IFN-α gene transfer alone (Fig. 4c). Since CCL17 has immune-suppressive activities such as not only Treg recruitment but also the induction of Th2 response [24] and inhibition of M1 macrophages [25], the inhibition of CCL17 may accentuate the antitumor immunity of IFN-α. These results demonstrate that this combination therapy could induce an efficient cellular immunity, leading to a strong antitumor effect, as shown in Fig. 3d. ...
The effect of IFN-α on the immunosuppressive tumor microenvironment is not fully understood. We previously reported that intratumoral IFN-α gene transduction decreased the frequency of regulatory T cells (Tregs) in the tumor by inducing the secretion of IL-6 from dendritic cells. In this study, we examined whether IFN-α affects the trafficking of Tregs to the tumor. Since CT26 cells expressed CCL17 among Treg-attracting chemokines, we focused on its role in IFN-α-mediated Treg suppression. IFN-α directly suppressed CCL17 production from CT26 cells in vitro, and IFN-α transduction reduced CCL17 expression in tumors in vivo. Next, to investigate whether CCL17 downregulation is related to the suppression of Treg trafficking, CCL17-downregulated CT26 cells produced using short hairpin RNA (CT26-shCCL17) were inoculated into mice. The frequency of Tregs in CT26-shCCL17 tumors was reduced and tumor growth was suppressed. Finally, to examine the combinatorial effect of IFN-α expression with CCL17 downregulation, IFN-α was transduced into CT26-shCCL17 tumors. This resulted in an elevation of CT26-specific CD8+ T cells and the complete eradication of tumors. This study shows a novel mechanism of IFN-α-mediated Treg suppression, and combining IFN-α gene therapy with strong CCL17 downregulation could offer a promising strategy for the treatment of cancer.
... vitro by exposure to IL-4 and IL-13, and show high expression of anti-inflammatory agents: cytokines such as IL-10, TGF-and Arginase-1(Kamei, 2006;Mosser, 2008;Lumeng, 2007;Fujisaka, 2009). M2 macrophages have been described to partially prevent the activation and accumulation of M1 cells (van Ginderachter, 2006;Katakura, 2004) by the secretion of the anti-inflammatory cytokine IL-10. During progression of DIO, M1-polarized CCR2 + monocytes are getting recruited and the increasing accumulation of CD11c + ATMs eventually overwhelms the protective effects of M2-like macrophages. ...
The metabolic syndrome is defined as the simultaneous clinical apparence of
obesity, insulin resistance, hepatic steatosis, and dyslipidemia. It represents a major
life-threatening risk factor for cardiovascular disease, such as coronary artery
disease. Chronic, low-grade inflammation and leukocyte infiltration of the adipose
tissue is a prerequisite of the metabolic syndrome. β2-integrins, such as Mac-1, are
classic adhesive receptors required for efficient inflammatory cell recruitment. In this
present study, we tested the contribution of CD40L/Mac-1 interaction, the elsewhere
proven pro-inflammatory dyad, to adipose tissue inflammation and its metabolic
outcome in mice by the genetic abrogation of Mac-1 and specific blockade of CD40L.
In a genetic knock out study, WT and Mac-1-/- mice consumed either a standard lowfat
diet (LFD) or a high fat diet (HFD) for 20 week. Contrasting our initial hypothesis,
Mac-1-/- mice gained significantly more weight on HFD compared with respective
control animals, while animals fed with LFD did not show differences in weight. While
intraperitoneal glucose- and insulin tolerance testing showed no alteration, Mac-1
deficiency potentiated accumulation of CD11c+ M1-like macrophages and proinflammatory
CD8+ cells. Furthermore, Mac-1-/- mice presented histological features
of non-alcoholic liver steatosis (NASH). To block Mac-1’s interaction to the costimulatory
molecule CD40L, we employed the highly specific peptide inhibitor, cM7,
or its unspecific control peptide, scM7, in a mouse model of diet-induced obesity for
20 weeks. cM7-treated animals showed no alterations in metabolic parameters, such
as weight, glucose tolerance, or insulin resistance. In contrast to the genetic loss of
Mac-1, adipose tissue from cM7-treated animals contained fewer CD8+ T-cells. In
particular, CD8+ T-effector-memory cell counts were reduced by 37 ± 9.6 %, while
numbers of macrophages and B-cells remained unaffected, which suggest a specific
Mac-1-dependent pro-inflammatory role of CD40L.
Taken together, we present the surprising finding that genetic absence of the
leukocyte integrin Mac-1 critically does not protect from, but enhances adipose tissue
inflammation during diet-induced obesity. These effects could not be attributed to the
interaction with CD40L since its specific inhibition by the small peptide inhibitor cM7
ameliorated adipose tissue inflammation by protecting from CD8+ T-cell immune cell
accumulation. These findings demonstrate: while unselective modulation of Mac-1 is
not favorable, selective blockade of CD40L’s inflammatory interaction with Mac-1
might describe a new therapeutic tool against the Metabolic Syndrome.
... CCL17 was originally considered to be a M2 cytokine due to its preferential attraction of T H 2 lymphocytes [25,40]. It can be produced by certain macrophage/dendritic cell populations [18,23,41,42] and is elevated in many inflammatory conditions [18,[43][44][45] and in synovial fluid in OA [46]. CiOA synovitis in the absence of CCL17 suggests that CCL17 has other functions, apart from a chemotactic role [18]. ...
Background:
Granulocyte macrophage-colony stimulating factor (GM-CSF) has been implicated in the pathogenesis of a number of inflammatory diseases and in osteoarthritis (OA). We identified previously a new GM-CSF→Jmjd3→interferon regulatory factor 4 (IRF4)→chemokine (c-c motif) ligand 17 (CCL17) pathway, which is important for the development of inflammatory arthritis pain and disease. Tumour necrosis factor (TNF) can also be linked with this pathway. Here we investigated the involvement of the pathway in OA pain and disease development using the GM-CSF-dependent collagenase-induced OA (CiOA) model.
Methods:
CiOA was induced in C57BL/6 wild-type (WT), Irf4-/-, Ccl17E/E, Ccr4-/-, Tnf-/-and GM-CSF-/-mice. Additionally, therapeutic targeting of CCL17, Jmjd3 and cyclooxygenase 2 (COX-2) was evaluated. Development of pain (assessment of weight distribution) and OA disease (histologic scoring of synovitis, cartilage destruction and osteophyte size) were assessed. Synovial joint cells, including neutrophils, macrophages, fibroblasts and endothelial cells, were isolated (cell sorting) and gene expression analyzed (quantitative PCR).
Results:
Studies in the gene-deficient mice indicated that IRF4, CCL17 and the CCL17 receptor, CCR4, but not TNF, were required for CiOA pain and optimal cartilage destruction and osteophyte size. Therapeutic neutralization of CCL17 and Jmjd3 ameliorated both pain and disease, whereas the COX-2 inhibitor only ameliorated pain. In the synovium Ccl17 mRNA was expressed only in the macrophages in a GM-CSF-dependent and IRF4-dependent manner.
Conclusions:
The GM-CSF→Jmjd3→IRF4→CCL17 pathway is important for the development of CiOA, with CCL17 thus being a potential therapeutic target for the treatment of both OA pain and disease.
... The particle-uptaking AM population was CD38 Int early J ALLERGY CLIN IMMUNOL VOLUME 142, NUMBER 4 growth response 2 (Egr2) Hi (M2-like), whereas the IM population was CD38 Hi Egr2 Int (M1-like; see Fig E3, A and B). 43 In agreement with their M2-like phenotype, in vitro treatment of cell-sorted AM with UFP sharply upregulated their production of the cytokines IL-10, CCL17, IL-6 and TNF-a but induced little change in their baseline production of IL-12 (see Fig E3, C). 44,45 Of the remaining beads (15%), about two thirds were picked up by CD11c hi CD11b int CD8a 1 CD103 1 B220 2 PDCA1 2 classical DC 46 and the rest by Gr-1 1 Siglec-F low neutrophils (Fig 1, B and C, and see Fig E1). These results indicate that AM and IM were the major cell subsets responsible for clearance of inhaled UFP in the context of allergic airway inflammation. ...
Background:
Exposure to traffic-related particulate matter (PM) promotes asthma and allergic diseases. However, the precise cellular and molecular mechanisms by which PM exposure acts to mediate these effects remain unclear.
Objective:
We sought to elucidate the cellular targets and signaling pathways critical for the augmentation of allergic airway inflammation induced by ambient ultra fine particles (UFP).
Methods:
We employed in vitro cell culture assays using lung-derived antigen presenting cells and allergen-specific T cells, and in vivo mouse models of allergic airway inflammation that employed myeloid lineage-specific gene deletions, cellular reconstitution approaches and antibody inhibition studies.
Results:
We identified lung alveolar macrophage (AM) as the key cellular target of UFP in promoting airway inflammation. Aryl hydrocarbon receptor (AhR)-dependent induction of Jagged 1 (Jag1) expression in AM was necessary and sufficient for the augmentation of allergic airway inflammation by UFP. UFP promoted Th2 and Th17 cell differentiation of allergen-specific T cells in a Jag1- and Notch4-dependent manner. Treatment of mice with an anti-Notch 4 antibody abrogated the exacerbation of allergic airway inflammation induced by UFP.
Conclusion:
UFP exacerbate allergic airway inflammation by promoting a Jag1-Notch4-dependent interaction between Alveolar Macrophages and Allergen-Specific T cells, leading to augmented Th cell differentiation.
... M2 macrophages generally have high levels of mannose receptors and scavenger receptors, and play important roles in polarized Th2 reactions. For example, M2 macrophages promote the encapsulation and killing of parasites and have immunoregulatory and anti-inflammatory functions [48]. This macrophage population is also thought to play a critical role in negative regulation of host protective immunity against microbial infections. ...
Background
Entamoeba nuttalli is an intestinal protozoan with pathogenic potential that can cause amoebic liver abscess. It is highly prevalent in wild and captive macaques. Recently, cysts were detected in a caretaker of nonhuman primates in a zoo, indicating that E. nuttalli may be a zoonotic pathogen. Therefore, it is important to evaluate the pathogenicity of E. nuttalli in detail and in comparison with that of E. histolytica.
Methodology/Principal findings
Trophozoites of E. nuttalli GY4 and E. histolytica SAW755 strains were inoculated into liver of hamsters. Expression levels of proinflammatory factors of hamsters and virulence factors from E. histolytica and E. nuttalli were compared between the two parasites. Inoculations with trophozoites of E. nuttalli resulted in an average necrotic area of 24% in liver tissue in 7 days, whereas this area produced by E. histolytica was nearly 50%. Along with the mild liver tissue damage induced by E. nuttalli, expression levels of proinflammatory factors (TNF-α, IL-6 and IL-1β) and amebic virulence protein genes (lectins, cysteine proteases and amoeba pores) in local tissues were lower with E. nuttalli in comparison with E. histolytica. In addition, M2 type macrophages were increased in E. nuttalli-induced amebic liver abscesses in the late stage of disease progression and lysate of E. nuttalli trophozoites induced higher arginase expression than E. histolytica in vitro.
Conclusions/Significance
The results show that differential secretion of amebic virulence proteins during E. nuttalli infection triggered lower levels of secretion of various cytokines and had an impact on polarization of macrophages towards a M1/M2 balance. However, regardless of the degree of macrophage polarization, there is unambiguous evidence of an intense acute inflammatory reaction in liver of hamsters after infection by both Entamoeba species.
Triple negative breast cancer (TNBC) has poor prognosis when compared to other breast cancer subtypes. Despite pre-clinical data supporting an immune targeted approach for TNBCs, immunotherapy has failed to demonstrate the impressive responses seen in other solid tumor malignancies. Additional strategies to modify the tumor immune microenvironment and potentiate response to immunotherapy are needed. In this review, we summarise phase III data supporting the use of immunotherapy for TNBC. We discuss the role of IL-1β in tumorigenesis and summarize pre-clinical data supporting IL-1β inhibition as a potential therapeutic strategy in TNBC. Finally, we present current trials evaluating IL-1β in breast cancer and other solid tumor malignancies and discuss future studies that may provide a strong scientific rationale for the combination of IL-1β and immunotherapy in the neoadjuvant and metastatic setting for people with TNBC.
Effective regeneration after peripheral nerve injury requires macrophage recruitment. We investigated the activation of remodeling pathways within the macrophage population when repair is delayed and identified alteration of key upstream regulators of the inflammatory response. We then targeted one of these regulators, using exogenous IL10 to manipulate the response to injury at the repair site. We demonstrate that this approach alters macrophage polarization, promotes macrophage recruitment, axon extension, neuromuscular junction formation, and increases the number of regenerating motor units reaching their target. We also demonstrate that this approach can rescue the effects of delayed nerve graft.
Ischemia-reperfusion injury (IRI) amplifies T cell alloimmune responses after transplantation with thrombin playing a key pro-inflammatory role. To explore the influence of thrombin on regulatory T cell recruitment and efficacy we used a well-established model of IRI in the native murine kidney. Administration of the cytotopic thrombin inhibitor PTL060 inhibited IRI, and by skewing expression of chemokines (reducing CCL2 and CCL3 but increasing CCL17 and CCL22) increased the infiltration of M2 macrophages and Tregs. When PTL060 was combined with infusion of additional Tregs, these effects were further amplified. To test the benefits of thrombin inhibition in a transplant model, BALB/c hearts were transplanted into B6 mice with or without perfusion with PTL060 in combination with Tregs. Thrombin inhibition or Treg infusion alone led to small increments in allograft survival. However, the combined therapy led to modest graft prolongation by the same mechanisms as in renal IRI; graft survival was accompanied by increased numbers of Tregs and anti-inflammatory macrophages, and reduced expression of pro-inflammatory cytokines. While the grafts succumbed to rejection associated with the emergence of alloantibody, these data suggest that thrombin inhibition within the transplant vasculature enhances the efficacy of Treg infusion, a therapy that is currently entering the clinic to promote transplant tolerance.
As burn injuries are often followed by a derailed immune response and excessive inflammation, a thorough understanding of the occurring reactions is key to prevent secondary complications. This systematic review, that includes 247 animal studies, shows the post-burn response of 14 different immune cell types involved in immediate and long-term effects, in both wound tissue and circulation. Peripheral blood neutrophil and monocyte numbers increased directly after burns, whereas thrombocyte numbers increased near the end of the first week. Lymphocyte numbers, however, were decreased for at least two weeks. In burn wound tissue, neutrophil and macrophage numbers accumulated during the first three weeks. Burns also altered cellular functions as we found increased migratory potential of leukocytes, impaired antibacterial activity of neutrophils and enhanced inflammatory mediator production by macrophages. Neutrophil surges were positively associated with burn size and were highest in rats. Altogether, this comprehensive overview of the temporal immune cell dynamics shows that unlike normal wound healing, burn injury induces a long-lasting inflammatory response. It provides a fundamental research basis to improve experimental set-ups, burn care and outcome.
Cytokines are critical for intercellular communication in human health and disease, but the investigation of cytokine signaling activity has remained challenging due to the short half-lives of cytokines and the complexity/redundancy of cytokine functions. To address these challenges, we developed the Cytokine Signaling Analyzer (CytoSig; https://cytosig.ccr.cancer.gov/), providing both a database of target genes modulated by cytokines and a predictive model of cytokine signaling cascades from transcriptomic profiles. We collected 20,591 transcriptome profiles for human cytokine, chemokine and growth factor responses. This atlas of transcriptional patterns induced by cytokines enabled the reliable prediction of signaling activities in distinct cell populations in infectious diseases, chronic inflammation and cancer using bulk and single-cell transcriptomic data. CytoSig revealed previously unidentified roles of many cytokines, such as BMP6 as an anti-inflammatory factor, and identified candidate therapeutic targets in human inflammatory diseases, such as CXCL8 for severe coronavirus disease 2019.
Blood plasma proteins play an important role in immune defense against pathogens, including cytokine signaling, the complement system and the acute-phase response. Recent large-scale studies have reported genetic (i.e. quantitative trait loci, pQTLs) and non-genetic factors, such as age and sex, as major determinants to inter-individual variability in immune response variation. However, the contribution of blood cell composition to plasma protein heterogeneity has not been fully characterized and may act as a confounding factor in association studies. Here, we evaluated plasma protein levels from 400 unrelated healthy individuals of western European ancestry, who were stratified by sex and two decades of life (20-29 and 60-69 years), from the Milieu Interieur cohort. We quantified 297 proteins by Luminex in a clinically certified laboratory and their levels of variation were analysed together with 5.2M single-nucleotide polymorphisms. With respect to non-genetic variables, we included more than 700 lifestyle and biochemical factors, as well as counts of seven circulating immune cell populations measured by hemogram and standardized flow cytometry. Collectively, we found 152 significant associations involving 49 proteins and 20 non-genetic variables. Consistent with previous studies, age and sex showed a global, pervasive impact on plasma protein heterogeneity, while body mass index and other health status variables were among the non-genetic factors with the highest number of associations. After controlling for these covariates, we identified 100 and 12 pQTLs acting in cis and trans, respectively, collectively associated with 87 plasma proteins and including 30 novel genetic associations. Genetic factors explained the largest fraction of the variability of plasma protein levels, as compared to non-genetic factors. In addition, blood cell fractions, including leukocytes, lymphocytes and three types of polymorphonuclear cells, had a larger contribution to inter-individual variability than age and sex, and appeared as confounders of specific genetic associations. Finally, we identified new genetic associations with plasma protein levels of eight monogenic Mendelian disease genes including three primary immunodeficiency genes (Ficolin-3, Interleukine-2 Receptor alpha and FAS). Our study identified novel genetic and non-genetic factors associated to plasma protein levels which may inform health status and disease management.
A way to delay aging and the related low-grade chronic inflammatory state is to study the model of positive physiology such as the Long-Living Individuals (LLIs). Our recent studies have shown higher levels of the host defense BPI Fold-Containing Family B Member 4 (BPIFB4) protein in the LLIs’ blood. Notably, BPIFB4 has been shown to influence monocytes typesetting and M2 anti-inflammatory phenotype (CD206+CD163++) macrophages skewing.
According to the role of a complex cytokine milieu in guiding the macrophage polarization, here we found that circulating concentrations of thymus and activation regulated chemokine (TARC)/CCL17 and small-inducible cytokine B10 (IP-10)/CXCL10) cytokines, were additionally associated with the LLIs’ state.
In a differentiation process in vitro, the addition of LLIs’ plasma to the cell culture medium, enhanced the ability of monocytes, either from LLIs or controls, to acquire a M2 phenotype. Interestingly, a neutralizing antibody against TARC blunted the M2 skewing effect of the LLIs’ plasma. Collectively, these data indicate that exceptional longevity may associate with a peculiar anti-inflammatory myeloid profile responsible for improved reparative processes and reduced inflammatory status mediated in part by TARC and M2 generation.
In the previous chapters, we have reviewed the definition and major functions of M2 macrophages of mammals, the evolutionary origin of the M2 macrophages, and possible M2 macrophage equivalents in invertebrates and nonmammalian vertebrates. This chapter is dedicated to the molecular mechanisms that drive M2 macrophage activation, or as often called, M2 macrophage polarization. There are prototypical M2 activating signals: Th2 cytokines, and the uptake of apoptotic cells. Th2 cytokines act through signal transducer and activator of transcription 6 (STAT6) by inducing its phosphorylation, nuclear translocation, and promoting transcription of STAT6-responsive genes. Apoptotic cells are metabolized within the macrophages, eventually activating lipid-sensing nuclear receptors. Excess M2 macrophage number and unwanted M2 macrophage activation cause disease; hence, mechanisms that inactivate STAT6 signaling are important to balance macrophage activation. Inactivation of STAT6 signaling can occur via STAT6 ubiquitination by the ubiquitin ligase ring finger protein 128 [RNF128, also known as gene related to anergy in lymphocytes (GRAIL)]. Ubiquitination of STAT6 leads to its proteasomal degradation, hence allowing an M2 activation state to be switched off. There are noncanonical inducers of M2 genes, such as certain pathogen products, inflammatory mediators, lipid metabolites, hormones, and neurotransmitters. Epigenetic modifications also affect M2 activation, and there is an autocrine cytokine loop that sustains M2 activation.
Mechanisms of immune activation in effector cells during Haemonchus contortus infection in sheep are currently unknown. Microarray experiments have been performed on tissues of H. contortus infected sheep of varying parasite resistance during early and late points of infection, but not in immune effector cells. The purpose of this study was to compare early gene activation in peripheral blood mononuclear cells (PBMC) from primed parasite susceptible (Suffolk) and resistant (St. Croix) sheep in response to H. contortus larval antigen (HcLA). Peripheral Blood Mononuclear Cells were cultured for 6 hours with HcLA and RNA‐Sequencing was performed. St. Croix PBMC upregulated 499 unique genes in response to HcLA while Suffolk PBMC upregulated 130 unique genes and 25 genes were shared between the two breeds. St. Croix PBMC had increased expression of genes associated with immune function, signal transduction, response to stress and others. In addition, while mechanisms of innate recognition of H. contortus are unknown, multiple pattern recognition receptors were found to be upregulated in St. Croix PBMC cultured with HcLA and none were found to be upregulated in Suffolk PBMC. These patterns of immune gene activation may contribute to St. Croix’s rapid response and ability to resist H. contortus infection.
Patients with ulcerative colitis have an increased risk of developing colitis-associated colon cancer (CACC). Changes in glycosylation of the oncoprotein MUC1 commonly occur in chronic inflammation, including ulcerative colitis, and this abnormally glycosylated MUC1 promotes cancer development and progression. It is not known what causes changes in glycosylation of MUC1. Gene expression profiling of myeloid cells in inflamed and malignant colon tissues showed increased expression levels of inflammatory macrophage–associated cytokines compared with normal tissues. We analyzed the involvement of macrophage-associated cytokines in the induction of aberrant MUC1 glycoforms. A coculture system was used to examine the effects of M1 and M2 macrophages on glycosylation-related enzymes in colon cancer cells. M2-like macrophages induced the expression of the glycosyltransferase ST6GALNAC1, an enzyme that adds sialic acid to O-linked GalNAc residues, promoting the formation of tumor-associated sialyl-Tn (sTn) O-glycans. Immunostaining of ulcerative colitis and CACC tissue samples confirmed the elevated number of M2-like macrophages as well as high expression of ST6GALNAC1 and the altered MUC1-sTn glycoform on colon cells. Cytokine arrays and blocking antibody experiments indicated that the macrophage-dependent ST6GALNAC1 activation was mediated by IL13 and CCL17. We demonstrated that IL13 promoted phosphorylation of STAT6 to activate transcription of ST6GALNAC1. A computational model of signaling pathways was assembled and used to test IL13 inhibition as a possible therapy. Our findings revealed a novel cellular cross-talk between colon cells and macrophages within the inflamed and malignant colon that contributes to the pathogenesis of ulcerative colitis and CACC.
See related Spotlight on p. 160
Type 2 immunity participates in the pathogeneses of helminth infection and allergic diseases. Emerging evidence indicates that the components of type 2 immunity are also involved in maintaining metabolic hemostasis and facilitating the healing process after tissue injury. Numerous preclinical studies have suggested regulation of type 2 immunity-related cytokines, such as interleukin-4, -13, and -33, and cell types, such as M2 macrophages, mast cells, and eosinophils, affects cardiac functions after myocardial infarction (MI), providing new insights into the importance of immune modulation in the infarcted heart. This review provides an overview of the functions of these cytokines and cells in the setting of MI as well as their potential to predict the severity and prognosis of MI.
Lung cancer continues to be the leading cause of cancer deaths and more than one million lung cancer patients will die every year worldwide. Radiotherapy (RT) plays an important role in lung cancer treatment, but the side effects of RT are pneumonitis and pulmonary fibrosis. RT-induced lung injury causes damage to alveolar-epithelial cells and vascular endothelial cells. Macrophages play an important role in the development of pulmonary fibrosis despite its role in immune response. These injury activated macrophages develop into classically activated M1 macrophage or alternative activated M2 macrophage. It secretes cytokines, interleukins, interferons, and nitric oxide. Several pro-inflammatory lipids and pro-apoptotic proteins cause lipotoxicity such as LDL, FC, DAG, and FFA. The overall findings in this review conclude the importance of macrophages in inducing toxic/inflammatory effects during RT of lung cancer, which is clinically vital to treat the radiation-induced fibrosis.
Purpose:
Tumor-associated macrophages (TAMs) and the hyperactivation of the PI3K/AKT pathway are involved in the pathogenesis of Hodgkin lymphoma and affect disease outcome. Because the δ and γ isoforms of PI3K are overexpressed in Hodgkin/Reed-Sternberg (HRS) cells and the tumor microenvironment (TME), we propose that the PI3Kδ/γ inhibitor RP6530 might affect both HRS cells and TME, ultimately leading to an enhanced antitumor response.
Experimental design:
Hodgkin lymphoma cell lines (L-540, KM-H2, and L-428) and primary human macrophages were used to investigate the activity of RP6530 in vitro and in vivo in Hodgkin lymphoma cell line xenografts.
Results:
In vitro, RP6530 besides killing and inhibiting the proliferation of Hodgkin lymphoma cells, downregulated lactic acid metabolism, switching the activation of macrophages from an immunosuppressive M2-like phenotype to a more inflammatory M1-like state. By RNA sequencing, we define tumor glycolysis as a specific PI3Kδ/γ-dependent pathway implicated in the metabolic reprogramming of cancer cells. We identify the metabolic regulator pyruvate kinase M2 as the main mediator of tumor-induced immunosuppressive phenotype of macrophages. Furthermore, we show in human tumor xenografts that RP6530 repolarizes TAMs into proinflammatory macrophages and inhibits tumor vasculature, leading to tumor regression. Interestingly, patients with Hodgkin lymphoma experiencing objective responses (complete response and partial response) in a phase I trial using RP6530 showed a significant inhibition of circulating myeloid-derived suppressor cells and an average mean reduction in serum thymus and activation-regulated chemokine levels of 40% (range, 4%-76%).
Conclusions:
Our results support PI3Kδ/γ inhibition as a novel therapeutic strategy that targets both malignant cells and the TME to treat patients with Hodgkin lymphoma.
Pancreatic ductal adenocarcinoma (PDAC) is a highly lethal malignancy remaining incurable to date. The tumor cells and the relevant oncogenic signaling have been extensively investigated, still, the exact molecular and cellular pathology of the disease is not well known due to its fibrous stroma. The tumor microenvironment of PDAC is characterized by a dense desmoplastic stroma consisting of fibroblast, pancreatic stellate cells (PSC), immune cells and the extracellular matrix components. PSCs and tumor associated macrophages (TAM) are the most abundant and important cells in the tumor microenvironment of PDAC and their cellular communication play a significant role in PDAC progression and immune escape. The molecular mechanisms of PSCs activation and the role of their secreted proteins in macrophage polarization as well as the possibility of defining biomarkers from the secretome of pancreatic cancer cells were studied in this thesis. The secretome of tumor necrosis factor-alpha (TNF-α) activated PSCs were analyzed and validated using different techniques. Activation of PSCs resulted in regulating the secretome profile of PSCs and increased ROS production. The study also demonstrated that activated PSCs secrete numerous proteins which are associated with the cancer cell proliferation and immune cells activation. Co-culturing PDAC cells with the secretome of activated PSCs resulted in a significant increase of cells proliferation. Moreover, the secretome of PDAC cells and activated PSCs promoted the expression of HLA-DR on monocytes and induced the polarization of macrophages into the immunosuppressive M2 phenotype. The M2 polarization was confirmed by their ability to express CD163 and CD206 surface markers, higher arginase activity and interestingly, their conditioned medium increased Panc-1 cell proliferation. Additionally, interleukin-6 (IL-6) differentially regulated the expression of M1 and M2 surface markers pre- and post-polarization. Moreover, the secreted molecules by pancreatic cancer cells, sera of PDAC patients, chronic pancreatitis (CP) and sera from healthy donors were studied for the establishment of a set of defined biomarkers. The study indicated that eight markers which were similarly regulated in the PDAC cell secretome and serum of PDAC patients could increase the accuracy of PDAC diagnosis up to 10%. Abstract 8 Taken together, these results will provide insight into the complex and dynamic microenvironment of pancreatic cancer and show the critical role of tumor and stromal cell’s secretome in PDAC development and cellular communications in the tumor microenvironment.
The glucocorticoid (GC) receptor (GR) suppresses inflammation by activating anti-inflammatory and repressing pro-inflammatory genes. GR-interacting protein-1 (GRIP1) is a GR corepressor in macrophages, however, whether GRIP1 mediates GR-activated transcription , and what dictates its coactivator versus corepressor properties is unknown. Here we report that GRIP1 loss in macrophages attenuates glucocorticoid induction of several anti-inflammatory targets, and that GC treatment of quiescent macrophages globally directs GRIP1 toward GR binding sites dominated by palindromic GC response elements (GRE), suggesting a non-redundant GRIP1 function as a GR coactivator. Interestingly, GRIP1 is phosphorylated at an N-terminal serine cluster by cyclin-dependent kinase-9 (CDK9), which is recruited into GC-induced GR:GRIP1:CDK9 hetero-complexes, producing distinct GRE-specific GRIP1 phospho-isoforms. Phosphorylation potentiates GRIP1 coactivator but, remarkably, not its corepressor properties. Consistently, phospho-GRIP1 and CDK9 are not detected at GR transrepression sites near pro-inflammatory genes. Thus, GR restricts actions of its own coregulator via CDK9-mediated phosphorylation to a subset of anti-inflammatory genes.
Activated mouse peritoneal macrophages produce nitric oxide (NO) via a nitric oxide synthase that is inducible by interferon gamma (IFN-gamma): iNOS. We have studied the mechanisms by which transforming growth factor beta 1 (TGF-beta) suppresses IFN-gamma-stimulated NO production. TGF-beta treatment reduced iNOS specific activity and iNOS protein in both cytosolic and particulate fractions as assessed by Western blot with monospecific anti-iNOS immunoglobulin G. TGF-beta reduced iNOS mRNA without affecting the transcription of iNOS by decreasing iNOS mRNA stability. Even after iNOS was already expressed, TGF-beta reduced the amount of iNOS protein. This was due to reduction of iNOS mRNA translation and increased degradation of iNOS protein. The potency of TGF-beta as a deactivator of NO production (50% inhibitory concentration, 5.6 +/- 2 pM) may reflect its ability to suppress iNOS expression by three distinct mechanisms: decreased stability and translation of iNOS mRNA, and increased degradation of iNOS protein. This is the first evidence that iNOS is subject to other than transcriptional regulation.
The initiation of an immune response is critically dependent on the activation of dendritic cells (DCs). This process is triggered by surface receptors specific for inflammatory cytokines or for conserved patterns characteristic of infectious agents. Here we show that human DCs are activated by influenza virus infection and by double-stranded (ds)RNA. This activation results not only in increased antigen presentation and T cell stimulatory capacity, but also in resistance to the cytopathic effect of the virus, mediated by the production of type I interferon, and upregulation of MxA. Because dsRNA stimulates both maturation and resistance, DCs can serve as altruistic antigen-presenting cells capable of sustaining viral antigen production while acquiring the capacity to trigger naive T cells and drive polarized T helper cell type 1 responses.
Expression of the macrophage mannose receptor is inhibited by interferon gamma (IFN-gamma), a T helper type 1 (Th-1)-derived lymphokine. Interleukin 4 (IL-4), a Th-2 lymphocyte product, upregulates major histocompatibility class II antigen expression but inhibits inflammatory cytokine production by macrophages. We have studied the effect of IL-4 on expression of the macrophage mannose receptor (MMR) by elicited peritoneal macrophages. We found that recombinant murine IL-4 enhances MMR surface expression (10-fold) and activity (15-fold), as measured by the respective binding and degradation of 125I-mannose-bovine serum albumin. Polymerase chain reaction analysis of cDNAs from purified primary macrophage populations revealed that MMR, but not lysozyme or tumor necrosis factor alpha, mRNA levels were markedly increased by IL-4. The above effects were associated with morphologic changes. These data establish IL-4 as a potent and selective enhancer of murine MMR activity in vitro. IL-4 induces inflammatory macrophages to adopt an alternative activation phenotype, distinct from that induced by IFN-gamma, characterized by a high capacity for endocytic clearance of mannosylated ligands, enhanced (albeit restricted) MHC class II antigen expression, and reduced proinflammatory cytokine secretion.
Activated mouse peritoneal macrophages produce nitric oxide (NO) via a nitric oxide synthase that is inducible by interferon gamma (IFN-gamma): iNOS. We have studied the mechanisms by which transforming growth factor beta 1 (TGF-beta) suppresses IFN-gamma-stimulated NO production. TGF-beta treatment reduced iNOS specific activity and iNOS protein in both cytosolic and particulate fractions as assessed by Western blot with monospecific anti-iNOS immunoglobulin G. TGF-beta reduced iNOS mRNA without affecting the transcription of iNOS by decreasing iNOS mRNA stability. Even after iNOS was already expressed, TGF-beta reduced the amount of iNOS protein. This was due to reduction of iNOS mRNA translation and increased degradation of iNOS protein. The potency of TGF-beta as a deactivator of NO production (50% inhibitory concentration, 5.6 +/- 2 pM) may reflect its ability to suppress iNOS expression by three distinct mechanisms: decreased stability and translation of iNOS mRNA, and increased degradation of iNOS protein. This is the first evidence that iNOS is subject to other than transcriptional regulation.
T helper cells type 1 (Th1s) that produce interferon-γ predominantly mediate cellular immune responses and are involved in
the development of chronic inflammatory conditions, whereas Th2s which produce large amounts of IL-4 and IL-5 upregulate
IgE production and are prominent in the pathogenesis of allergic diseases. The precise factors determining whether Th1- or
Th2-mediated immune responses preferentially occur at a peripheral site of antigen exposure are largely unknown. Chemokines,
a superfamily of polypeptide mediators, are a key component of the leukocyte recruitment process. Here we report that among
four CXC (CXCR1-4) and five CC (CCR1-5) chemokine receptors analyzed, CXCR3 and CCR5 are preferentially expressed in human
Th1s. In contrast, Th2s preferentially express CCR4 and, to a lesser extent, CCR3. In agreement with the differential chemokine
receptor expression, Th1s and Th2s selectively migrate in response to the corresponding chemokines. The differential expression
of chemokine receptors may dictate, to a large extent, the migration and tissue homing of Th1s and Th2s. It may also determine
different susceptibility of Th1s and Th2s to human immunodeficiency virus strains using different fusion coreceptors.
The mannose receptor is a macrophage surface receptor that mediates both endocytosis and phagocytosis. Previous work has demonstrated that the prototypical Th2 cytokine, interleukin-4 (IL-4), increases both cell-surface receptor expression and mannose receptor-mediated endocytosis, whereas the prototypical Th1 cytokine, interferon-gamma (IFN-gamma), decreases both surface expression and endocytosis. In many aspects of the immune response, Th1 and Th2 cytokines oppose each others' actions. We demonstrate that IL-4 and IFN-gamma alone and together enhance mannose receptor-mediated phagocytosis, despite opposing effects on cell-surface mannose receptor expression and endocytosis. Thus these usually antagonistic cytokines cooperate in increasing mannose receptor phagocytic function. The cooperative effect of these cytokines is not observed for Fc receptor-mediated phagocytosis. The Th2 cytokine IL-13 exerts similar effects to IL-4. Our results suggest that Th1 and Th2 cytokines may act in concert at sites of inflammation to enhance mannose receptor-mediated phagocytosis of microorganisms.
Helper T cells are classified into Th1 and Th2 subsets based on their profiles of cytokine production. Th1 cells are involved in cell-mediated immunity, whereas Th2 cells induce humoral responses. Selective recruitment of these two subsets depends on specific adhesion molecules and specific chemoattractants. Here, we demonstrate that the T cell-directed CC chemokine thymus and activation-regulated chemokine (TARC) was abundantly produced by monocytes treated with granulocyte macrophage colony stimulating factor (GM-CSF) or IL-3, especially in the presence of IL-4 and by dendritic cells derived from monocytes cultured with GM-CSF + IL-4. The receptor for TARC and another macrophage/dendritic cell-derived CC chemokine macrophage-derived chemokine (MDC) is CCR4, a G protein-coupled receptor. CCR4 was found to be expressed on approximately 20% of adult peripheral blood effector/memory CD4+ T cells. T cells attracted by TARC and MDC generated cell lines predominantly producing Th2-type cytokines, IL-4 and IL-5. Fractionated CCR4+ cells but not CCR4- cells also selectively gave rise to Th2-type cell lines. When naive CD4+ T cells from adult peripheral blood were polarized in vitro, Th2-type cells selectively expressed CCR4 and vigorously migrated toward TARC and MDC. Taken together, CCR4 is selectively expressed on Th2-type T cells and antigen-presenting cells may recruit Th2 cells expressing CCR4 by producing TARC and MDC in Th2-dominant conditions.
Evidence is provided that macrophages can make M-1 or M-2 responses. The concept of M-1/M-2 fomented from observations that macrophages from prototypical Th1 strains (C57BL/6, B10D2) are more easily activated to produce NO with either IFN-gamma or LPS than macrophages from Th2 strains (BALB/c, DBA/2). In marked contrast, LPS stimulates Th2, but not Th1, macrophages to increase arginine metabolism to ornithine. Thus, M-1/M-2 does not simply describe activated or unactivated macrophages, but cells expressing distinct metabolic programs. Because NO inhibits cell division, while ornithine can stimulate cell division (via polyamines), these results also indicate that M-1 and M-2 responses can influence inflammatory reactions in opposite ways. Macrophage TGF-beta1, which inhibits inducible NO synthase and stimulates arginase, appears to play an important role in regulating the balance between M-1 and M-2. M-1/M-2 phenotypes are independent of T or B lymphocytes because C57BL/6 and BALB/c NUDE or SCID macrophages also exhibit M-1/M-2. Indeed, M-1/M-2 proclivities are magnified in NUDE and SCID mice. Finally, C57BL/6 SCID macrophages cause CB6F1 lymphocytes to increase IFN-gamma production, while BALB/c SCID macrophages increase TGF-beta production. Together, the results indicate that M-1- or M-2-dominant macrophage responses can influence whether Th1/Th2 or other types of inflammatory responses occur.
Thymus- and activation-regulated chemokine (TARC; CCL17) is a lymphocyte-directed CC chemokine that specifically chemoattracts CC chemokine receptor 4-positive (CCR4(+)) Th2 cells. To establish the pathophysiological roles of TARC in vivo, we investigated here whether an mAb against TARC could inhibit the induction of asthmatic reaction in mice elicited by OVA. TARC was constitutively expressed in the lung and was up-regulated in allergic inflammation. The specific Ab against TARC attenuated OVA-induced airway eosinophilia and diminished the degree of airway hyperresponsiveness with a concomitant decrease in Th2 cytokine levels. Our results for the first time indicate that TARC is a pivotal chemokine for the development of Th2-dominated experimental allergen-induced asthma with eosinophilia and AHR. This study also represents the first success in controlling Th2 cytokine production in vivo by targeting a chemokine.
Monocyte chemoattractant protein (MCP)-1 has a pathogenic role in herpesvirus-induced encephalomyelitis (HSM). Anti-MCP-1 antibody greatly decreased HSM severity in mice infected with herpes simplex virus type 2 (HSM mice), compared with its effect in control HSM mice treated with rabbit immunoglobulin. HSM severity was markedly enhanced in mice previously treated with a mixture of interleukin (IL) 4 and -10. In response to stimulation with antigen, HSM mouse cells isolated from cerebrospinal fluids (CSF cells) produced IL-4 in culture fluids; however, IL-4 production decreased in CSF cells derived from HSM mice previously treated with anti-MCP-1 antibody. A macrophage population isolated in CSF cells from HSM mice (CSF-Mphi) produced MCP-1 in culture fluids. In response to stimulation with herpesvirus antigen, a population of T cells isolated from CSF cells from HSM mice (CSF-T cells) produced IL-4 into their culture fluids, although MCP-1 was not produced by CSF-T cells stimulated by this antigen. IL-4 production by CSF-T cells was markedly enhanced when they were stimulated with viral antigen in the presence of murine recombinant MCP-1 (rMCP-1). Furthermore, IL-4 was produced in naive splenic T cells cocultured with CSF-Mphi. These results indicate that the severity of HSM is influenced by MCP-1, which stimulates Th2 responses.
Several reports have indicated that cell lineages apart from NK and T cells can also express IFN-gamma. However, the biological relevance of this finding is uncertain. We show in this study that bone marrow-derived macrophages (BMMs) express IFN-gamma at the mRNA and protein level early after infection with Chlamydia pneumoniae. Increased IFN-gamma mRNA accumulation by infected BMMs is early, transient, and requires both bacterial and host protein synthesis. The induction of IFN-gamma mRNA levels is independent of IL-12 and was dramatically enhanced in IL-10(-/-) BMMs. Such IL-10(-/-) BMMs contained less bacteria than the wild-type controls, whereas IFN-gammaR(-/-) BMMs showed increased C. pneumoniae load. Inducible NO synthase (iNOS) also participates in the control of bacterial load, as shown by the enhanced numbers of C. pneumoniae in iNOS(-/-) BMMs. However, the increased accumulation of iNOS mRNA and NO in C. pneumoniae-infected BMMs depended on the presence of IFN-alphabeta, but was independent of IFN-gamma. Interestingly, IFN-alphabeta are also required for increased IFN-gamma mRNA accumulation in C. pneumoniae-infected BMMs. Accordingly, IFN-alphabetaR(-/-) BMMs showed higher levels of C. pneumoniae than wild-type BMMs. Our findings unravel an autocrine/paracrine macrophage activation pathway by showing an IFN-alphabeta-dependent IFN-gamma and iNOS induction in response to infection, which protects macrophages against intracellular bacterial growth.
Unmethylated CpG motifs are prevalent in bacterial but not vertebrate genomic DNAs. Oligodeoxynucleotides (ODN) containing CpG motifs activate host defense mechanisms leading to innate and acquired immune responses. The recognition of CpG motifs requires Toll-like receptor (TLR) 9, which triggers alterations in cellular redox balance and the induction of cell signaling pathways including the mitogen activated protein kinases (MAPKs) and NF kappa B. Cells that express TLR-9, which include plasmacytoid dendritic cells (PDCs) and B cells, produce Th1-like proinflammatory cytokines, interferons, and chemokines. Certain CpG motifs (CpG-A) are especially potent at activating NK cells and inducing IFN-alpha production by PDCs, while other motifs (CpG-B) are especially potent B cell activators. CpG-induced activation of innate immunity protects against lethal challenge with a wide variety of pathogens, and has therapeutic activity in murine models of cancer and allergy. CpG ODN also enhance the development of acquired immune responses for prophylactic and therapeutic vaccination.
Previous studies have shown that IL-10 can induce the expression of the suppressor of cytokine signaling 3 (SOCS-3) mRNA in human monocytes and neutrophils, suggesting that the capacity of IL-10 to inhibit the expression of LPS-inducible proinflammatory genes may depend on SOCS-3 induction. However, no direct experimental evidence has been provided to support such hypothesis. Herein, we show that stable transfection of SOCS-3 into the mouse macrophage cell line J774 resulted in an inhibition of NO, TNF-alpha, IL-6, and GM-CSF secretion in response to LPS at levels similar to those exerted by IL-10 in LPS-stimulated wild-type J774. Constitutive SOCS-3 expression also down-regulated the mRNA expression of inducible NO synthase and IL-6 and impaired the production of TNF-alpha, mainly at a post-transcriptional level. In addition, SOCS-3-transfected cells displayed a constitutive expression of the IL-1R antagonist gene, consistent with the observation that IL-10 enhances IL-1R antagonist mRNA in LPS-stimulated wild-type cells. Furthermore, in peritoneal macrophages harvested from mice carrying heterozygous disruption of the SOCS-3 gene, IL-10 was less effective in repressing LPS-stimulated TNF-alpha and NO production. Taken together, our data show that SOCS-3 inhibits LPS-induced macrophage activation, strongly supporting the idea that it plays a role in the molecular mechanism by which IL-10 down-modulates the effector functions of LPS-activated macrophages. Finally, we show that forced expression of SOCS-3 significantly suppresses the ability of IL-10 to trigger tyrosine phosphorylation of STAT3. Therefore, SOCS-3 functions both as an LPS signal inhibitor and as a negative feedback regulator of IL-10/STAT3 signaling.
On human macrophages IL-10 acts as a more potent anti-inflammatory cytokine than IL-6, although both cytokines signal mainly via activation of the transcription factor STAT3. In this study we compare IL-10 and IL-6 signaling in primary human macrophages derived from blood monocytes. Pretreatment of macrophages with PMA or the proinflammatory mediators LPS and TNF-alpha blocks IL-6-induced STAT3 activation, whereas IL-10-induced activation of STAT3 remains largely unaffected. Although LPS induces the feedback inhibitor suppressor of cytokine signaling 3 (SOCS3) in macrophages, inhibition of IL-6 signal transduction by LPS occurs rapidly and does not depend on gene transcription. We also found that pretreatment of macrophages with IL-10 inhibits subsequent STAT3 activation by IL-6, whereas IL-10-induced STAT3 activation is not affected by preincubation with IL-6. This cross-inhibition is dependent on active transcription and might therefore be explained by different sensitivities of IL-10 and IL-6 signaling toward the feedback inhibitor SOCS3, which is induced by both cytokines. In contrast to the IL-6 signal transducer gp130, which has been previously shown to recruit SOCS3 to one of its phosphotyrosine residues (Y759), peptide precipitation experiments suggest that SOCS3 does not interact with phosphorylated tyrosine motifs of the IL-10R. Taken together, different sensitivities of IL-10 and IL-6 signaling toward mechanisms that inhibit the Janus kinase/STAT pathway define an important mechanism that contributes to the different anti-inflammatory potencies of these two cytokines.
The use of bacteria and bacterial extracts for immunotherapy has a checkered past. Recent developments in immunology reveal that these nonspecific immune activators actually work by triggering specific receptors that are expressed by subsets of immune cells. Identification of these receptors and the molecular signaling pathways that they activate has enabled a new era of specific targeted immunotherapy using chemically synthesized mimics of pathogen molecules.
Expression of the macrophage mannose receptor is inhibited by interferon gamma (IFN-gamma), a T helper type 1 (Th-1)-derived lymphokine. Interleukin 4 (IL-4), a Th-2 lymphocyte product, upregulates major histocompatibility class II antigen expression but inhibits inflammatory cytokine production by macrophages. We have studied the effect of IL-4 on expression of the macrophage mannose receptor (MMR) by elicited peritoneal macrophages. We found that recombinant murine IL-4 enhances MMR surface expression (10-fold) and activity (15-fold), as measured by the respective binding and degradation of 125I-mannose-bovine serum albumin. Polymerase chain reaction analysis of cDNAs from purified primary macrophage populations revealed that MMR, but not lysozyme or tumor necrosis factor alpha, mRNA levels were markedly increased by IL-4. The above effects were associated with morphologic changes. These data establish IL-4 as a potent and selective enhancer of murine MMR activity in vitro. IL-4 induces inflammatory macrophages to adopt an alternative activation phenotype, distinct from that induced by IFN-gamma, characterized by a high capacity for endocytic clearance of mannosylated ligands, enhanced (albeit restricted) MHC class II antigen expression, and reduced proinflammatory cytokine secretion.
T cells depend on costimulation by accessory cells for an immune response, Costimulatory macrophage activity involves the expression of B7 molecules whose expression is upregulated by interferon-gamma (IFN-gamma) and downregulated by interleukin-10 (IL-10). The expression of low-affinity Fc gamma IIIR (CD16), in contrast, is upregulated in the presence of IL-10 and downregulated in the presence of IFN-gamma, In human immunodeficiency virus-1 (HIV-1) infection, the balance between IFN-gamma and IL-10 expression shifts toward IL-10 predominance, Herein, we compare B7 and CD16 macrophage phenotypes from healthy and from HIV-1-infected patients, Patient macrophages express B7 molecules in lower density than macrophages from healthy donors and are resistant to the upregulation of costimulatory molecule expression, B7 expression can be normalized in patient macrophages by treating them with anti IL-10 monoclonal antibodies (mAb) and IFN-gamma together but not by treatment with either anti-IL-10 mAb or IFN-gamma alone, This finding suggests an excess of IL-10 in HIV-1 infection and an IFN-gamma deficiency, consistent with previous cytokine assessments in HIV-1-infected subjects, The upregulation of CD16 expression was readily induced in patient macrophages by treatment with IL-10 and was inhibited by treatment with IFN-gamma. CD16 expression identifies the subset of cytotoxic macrophages that has been shown to destroy CD4 T cells, which they target through CD4-reactive immune-complexed HIV-1 envelope molecules.
Bacterial DNA (bDNA) activates B cells and macrophages and can augment inflammatory responses by inducing release of proinflammatory cytokines. We found that bDNA stimulation of mouse spleen cells induced NK cell IFN-γ production that was dependent upon the presence of unmethylated CpG motifs, and oligonucleotides with internal CpG motifs could also induce splenocytes to secrete IFN-γ. The bDNA-induced IFN-γ response was strictly macrophage dependent. While splenocytes from SCID mice secreted IFN-γ in response to bDNA, depletion of macrophages eliminated this response. Additionally, purified NK cells did not respond to bDNA; however, addition of macrophages restored the NK cell IFN-γ response. Coculture of NK cells with preactivated macrophages further increased bDNA-induced NK cell IFN-γ production. Anti-IL-12 or IL-10 inhibited bDNA-induced IFN-γ response. Treatment of purified macrophages with bDNA resulted in IL-12 secretion accompanied by an increase in IL-12 p40 mRNA level. Although isolated NK cells did not make IFN-γ in response to bDNA, NK cells costimulated with IL-12 gained the ability to respond to bDNA. These experiments show that bDNA induces macrophage IL-12 production which, in turn, stimulates NK cell IFN-γ production. Macrophage-derived IL-12 renders NK cells responsive to bDNA permitting an even greater IFN-γ response to bDNA.
Growth hormone (GH) has been shown to promote wound healing and to improve protein metabolism in burned patients. Through immunomodulation, GH has also protected rats infected with Salmonella typhimurium and mice infected with Escherichia coli. In spite of advances in the management of patient care for those with thermal injuries, high mortality rates of burned patients as a result of infections are of special concern. An improvement in the resistance of burned patients to certain infections will make the beneficial role of GH very clear. In this study, therefore, the immunomodulating effects of recombinant human GH (rhGH) in thermally injured mice exposed to opportunistic herpesvirus infections were investigated.
(1) Burned mice, exposed to herpes simplex virus type 1 (HSV-1), were treated subcutaneously with rhGH (4 mg/kg) and observed for 21 days to determine the protective antiviral effect of rhGH. (2) Because of reports describing a lack of interferon-gamma (IFN-gamma) responsiveness in burned mice, the IFN-gamma-producing ability of the splenic mononuclear cells (SMNC) from burned mice treated with rhGH was examined. (3) Because the generation of burn-associated suppressor macrophages that can inhibit the IFN-gamma production by SMNC has been previously described, the suppressor cell activities of macrophages from burned mice treated with rhGH were examined.
After exposure to lethal amounts of HSV-1, mice treated with rhGH displayed a reduced mortality rate compared with control mice treated with saline. SMNC from burned mice treated with rhGH produced IFN-gamma, whereas this cytokine was not produced by SMNC from burned mice treated with saline. Also, an inhibition of the generation of burn-associated suppressor macrophages was displayed in burned mice treated with rhGH.
Exogenous administration of rhGH caused an improvement in the resistance of burned mice to HSV-1 infection. In burned mice treated with rhGH, the impaired IFN-gamma responsiveness was restored and the generation of burn-associated suppressor macrophages was inhibited. IFN-gamma, a typical antiviral cytokine induced by rhGH through the regulation of the suppressor macrophage generation, may therefore play a role in the protection of burned mice infected with a lethal amount of HSV-1.
We compared the immunological functions of interferon-gamma (IFN-gamma)-induced, classically activated macrophages (caM phi) and of interleukin-4 (IL-4)- and glucocorticoid-induced, alternatively activated macrophages (aaM phi) in a human co-culture system in vitro. Proliferation of peripheral blood leucocytes (PBL) or CD4+ T cells mediated by optimal doses of phytohaemagglutinin (PHA) or concanavalin A (Con A) was only marginally influenced by caM phi, but was strongly inhibited by aaM phi. The degree of lymphocyte proliferation sustained in the presence of caM phi was gradually reduced in a dose-dependent fashion by the addition of aaM phi. Flow cytometric analysis revealed that expression of costimulatory molecules such as CD11a, CD40, CD54, CD58, CD80 and CD86 did not vary significantly between caM phi and aaM phi and was low for CD58, CD80 and CD86. As shown by reverse transcriptase-polymerase chain reaction (RT-PCR) analysis, IL-10 was expressed in caM phi, aaM phi and control macrophages; the level of expression of IL-10 was slightly enhanced in aaM phi. Neither neutralizing anti-IL-10 antibodies, indomethacin nor NG-monomethyl-L-arginine (NMMLA) was able to reverse aaM phi-mediated inhibition of lymphocyte proliferation. Of several agents interfering with various second messenger pathways, cAMP and the Ca(2+)-ionophore A23187 inhibited differentiation of cultured human monocytes into phenotypically mature aaM phi expressing MS-1 high molecular weight protein (MS-1-HMWP) and RM 3/1 antigen, and prevented the suppressive action of aaM phi on lymphocyte proliferation. In conclusion, these results who that aaM phi actively inhibit mitogen-mediated proliferation of PBL and CD4+ T cells independently of the expression of costimulatory molecules and of IL-10, NO or prostaglandin synthesis, and that inhibition of phenotypic differentiation of aaM phi is paralleled by a lack of functional maturation. Thus, fully matured aaM phi may be functional in down-regulating CD4+ T-cell-mediated immune reactions by an as yet unknown mechanism.
Copoly(styrene-maleic acid)-conjugated neocarzinostatin (SMANCS), a lipophilic derivative of the proteinaceous antitumor antibiotic neocarzinostatin, has been reported to stimulate a nonspecific resistance to tumors (NSRT) in solid tumor-bearing mice, in addition to its chemotherapeutic antitumor effect through the arrest of DNA synthesis by direct DNA strand scission. In the present study, splenic or peritoneal effector cells were used to investigate the ability of SMANCS to augment natural killer (NK) cell activity and to generate cytostatic macrophages (A-M phi). Splenic NK cell activity augmented by SMANCS was characterized by cytotoxicity to various target cells, nylon wool nonadherence, and sensitivity to treatment with anti-asialo-GM1 antiserum or monoclonal anti-Thy-1.2 antibody followed by complement. The A-M phi generated by SMANCS stimulation were characterized by their adherence to a plastic surface coated with fetal calf serum and their ability to phagocytize carbonyl-iron. The maximum level of NK cell activity in the spleens of mice was detected 3 days after i.v. injection of SMANCS, and the highest activity of the peritoneal A-M phi was demonstrated in mice 4 days after SMANCS treatment. On the other hand, the NSRT of mice stimulated with SMANCS was not detectable in mice treated with carrageenan or trypan blue, whereas SMANCS-stimulated NSRT was observed in mice treated with anti-asialo-GM1 antiserum. The NSRT that was stimulated with SMANCS was also demonstrated in mice homozygous for the beige mutation and their non-beige littermates, when NK cell-resistant EL-4 thymoma was used as a tumor target. These results suggest that the expression of NSRT of mice stimulated with SMANCS may require the function of A-M phi, although NK cell activity was also augmented in spleens of mice by administration of SMANCS.
Suppressor macrophages (M phi) which can inhibit mitogen-induced lymphocyte proliferation appeared in the spleens of mice bearing transplanted MC-A fibrosarcoma cells. An analysis of the ontogeny of such M phi revealed additional suppressor activity directed against macrophage stem cells. Treatment of spleen cell suspensions with carbonyl iron followed by centrifugation removed suppressor M phi but did not deplete M phi-colony forming cells (M-CFC) which could be demonstrated in soft agar culture in L-cell conditioned medium (LCM). Untreated spleen cells had normal numbers of M-CFC; phagocyte-depleted mononuclear cells showed a threefold increase in M-CFC 14 days after subcutaneous inoculation of 10(6) MC-A cells per mouse. Further increases in M-CFC were also evident in similar preparations on Days 21 and 28 when the M-CFC concentration reached a maximum of eight times the normal level. The M phi which developed from the M-CFC grown in the presence of LCM were later shown to have indomethacin-sensitive suppressor activity suggesting the mediation of this phenomenon by prostaglandins. These observations suggest that locally produced phagocytic suppressor M phi from the spleens of tumor-bearing mice play important roles not only as inhibitors of lymphocyte proliferation as reported earlier, but also as regulators of monocyte-M phi production.
The mouse Chromosome 1 locus Ity regulates the extent to which Salmonella typhimurium replicates within the reticuloendothelial cell system (RES) during the first days of infection. If animals are homozygous for the Itys susceptibility allele, the Gram-negative bacterium undergoes rapid net multiplication, and mice die of a typhoid fever-like disease by day 10 of infection. Animals that are homozygous or heterozygous for the resistance allele, Ityr, control net bacterial replication and survive the first phase of salmonellosis. Indirect studies have implicated the resident macrophage as the effector cell for regulation of early in vivo salmonellae growth. To verify this supposition and to evaluate the phenotypic expression of Ity, we developed an in vitro assay to compare kinetics of S. typhimurium growth within Ityr and Itys macrophages. Resident peritoneal and splenic macrophages were used from inbred Ityr and Itys mice and from Ity congeneic mice. With these mice and through the use of radiolabeled S. typhimurium and an avirulent temperature-sensitive mutant of the bacterium, we found that: phagocytosis of S. typhimurium by Ityr and by Itys macrophages was the same; S. typhimurium grew to a greater extent in Itys peritoneal and splenic macrophages than in Ityr cells; Ityr macrophages killed intracellular salmonellae more efficiently than did Itys macrophages. Thus, we have demonstrated directly that Ity is expressed by the macrophage and have shown for the first time with Ity congeneic mice that the basis for differential net growth of virulent S. typhimurium in Ityr and Itys macrophages is a variation in the degree of bacterial kill.
Macrophage arginine metabolism via nitric oxide (NO) synthase and arginase pathways reduces and enhances tumor cell proliferation, respectively. Transforming growth factor-beta (TGF-beta) has been shown to down-regulate the NO synthase pathway. The present study describes the effect of TGF-beta on the arginase pathway. TGF-beta up-regulated arginase activity in rat peritoneal macrophages as assessed by measuring the generation of [14C]urea from [14C]-L-arginine in the presence of NG-monomethyl-L-arginine (L-NMMA). The stimulation, which reached fivefold after a 48-h exposure of macrophages to 10 ng/ml TGF-beta, was due to reduction in Km value of arginase. TGF-beta-induced up-regulation of arginase activity led to the release of more polyamines, mainly putrescine. The role of this up-regulation on macrophage cytotoxicity toward L-929 tumor cells was analyzed in coculture experiments. Macrophages blunted DNA synthesis by L-929 cells as assessed by measuring the incorporation of [3H]TdR into the cells and the proportion of cells in the G2 phase. Addition of TGF-beta in the presence of L-NMMA permitted L-929 cells cocultured with macrophages to resume DNA synthesis. The mechanism responsible for this restoration was the up-regulation of arginase activity rather than the down-regulation of NO synthase activity since TGF-beta in the presence of L-NMMA failed to further reduce NO synthase activity whereas it still enhanced arginase activity; synthetic putrescine (1-10 microM) also blunted macrophage cytotoxicity toward L-929 cells. This is the first evidence that TGF-beta up-regulates arginase activity in macrophages and, hence, limits macrophage-dependent cytostasis.
T cells depend on costimulation by accessory cells for an immune response. Costimulatory macrophage activity involves the expression of B7 molecules whose expression is upregulated by interferon-gamma (IFN-gamma) and downregulated by interleukin-10 (IL-10). The expression of low-affinity Fc gamma IIIR (CD16), in contrast, is upregulated in the presence of IL-10 and downregulated in the presence of IFN-gamma. In human immunodeficiency virus-1 (HIV-1) infection, the balance between IFN-gamma and IL-10 expression shifts toward IL-10 predominance. Herein, we compare B7 and CD16 macrophage phenotypes from healthy and from HIV-1-infected patients. Patient macrophages express B7 molecules in lower density than macrophages from healthy donors and are resistant to the upregulation of costimulatory molecule expression. B7 expression can be normalized in patient macrophages by treating them with anti IL-10 monoclonal antibodies (mAb) and IFN-gamma together but not by treatment with either anti-IL-10 mAb or IFN-gamma alone. This finding suggests an excess of IL-10 in HIV-1 infection and an IFN-gamma deficiency, consistent with previous cytokine assessments in HIV-1-infected subjects. The upregulation of CD16 expression was readily induced in patient macrophages by treatment with IL-10 and was inhibited by treatment with IFN-gamma. CD16 expression identifies the subset of cytotoxic macrophages that has been shown to destroy CD4T cells, which they target through CD4-reactive immune-complexed HIV-1 envelope molecules.
Bacterial DNA (bDNA) activates B cells and macrophages and can augment inflammatory responses by inducing release of proinflammatory cytokines. We found that bDNA stimulation of mouse spleen cells induced NK cell IFN-gamma production that was dependent upon the presence of unmethylated CpG motifs, and oligonucleotides with internal CpG motifs could also induce splenocytes to secrete IFN-gamma. The bDNA-induced IFN-gamma response was strictly macrophages dependent. While splenocytes from SCID mice secreted IFN-gamma in response to bDNA, depletion of macrophages eliminated this response. Additionally, purified NK cells did not respond to bDNA; however, addition of macrophages restored the NK cell IFN-gamma response. Coculture of NK cells with preactivated macrophages further increased bDNA-induced NK cell IFN-gamma production. Anti-IL-12 or IL-10 inhibited bDNA-induced IFN-gamma response. Treatment of purified macrophages with bDNA resulted in IL-12 secretion accompanied by an increase in IL-12 p40 mRNA level. Although isolated NK cells did not make IFN-gamma in response to bDNA, NK cells costimulated with IL-12 gained the ability to respond to bDNA. These experiments show that bDNA induces macrophage IL-12 production which, in turn, stimulates NK cell IFN-gamma production. Macrophage-derived IL-12 renders NK cells responsive to bDNA permitting an even greater IFN-gamma response to bDNA.
The RM3/1 Ag is a membrane glycoprotein restricted to human monocytes and macrophages that evolve in the late phase of inflammation. Peptide sequence analysis of the RM3/1 protein revealed similarity to CD163, a member of the scavenger receptor cysteine-rich family. Using specific Abs (RM3/1, Ki-M8), we demonstrate an identical cellular regulation for the RM3/1 and the CD163 protein. Most notably, we show for the first time that CD163 is significantly up-regulated by glucocorticoids. In contrast, the protein is down-regulated by the immunosuppressant cyclosporin A and by phorbol esters, while the inflammatory mediator LPS has no significant influence on the expression. We describe the first isolation of a full-length cDNA of CD163 and expression of the corresponding protein. Several splice variants of CD163 exist, and we elucidated the kinetics of induction of three major mRNA splice variants by fluticasone propionate; another splice variant was proved to be unresponsive to this glucocorticoid. Taken together with a previous result showing an involvement of RM3/1 in adhesion of monocytes to the activated endothelium, we discuss that CD163 might play an important role in inflammatory processes.
Phagocytosis of pathogens by macrophages initiates the innate immune response, which in turn orchestrates the adaptive response. In order to discriminate between infectious agents and self, macrophages have evolved a restricted number of phagocytic receptors, like the mannose receptor, that recognize conserved motifs on pathogens. Pathogens are also phagocytosed by complement receptors after relatively nonspecific opsonization with complement and by Fc receptors after specific opsonization with antibodies. All these receptors induce rearrangements in the actin cytoskeleton that lead to the internalization of the particle. However, important differences in the molecular mechanisms underlying phagocytosis by different receptors are now being appreciated. These include differences in the cytoskeletal elements that mediate ingestion, differences in vacuole maturation, and differences in inflammatory responses. Infectious agents, such as M. tuberculosis, Legionella pneumophila, and Salmonella typhimurium, enter macrophages via heterogeneous pathways and modify vacuolar maturation in a manner that favors their survival. Macrophages also play an important role in the recognition and clearance of apoptotic cells; a notable feature of this process is the absence of an inflammatory response.
Murine macrophages are able to distinguish bacterial from mammalian DNA. The response is mimicked by single-stranded oligonucleotides containing unmethylated CG dinucleotides ("CpG" motifs) in specific sequence contexts. The dose-response curve for activation is influenced by variation in the sequence flanking the core CpG motif. CpG or bacterial DNA activates several signaling pathways in common with bacterial lipopolysaccharide (LPS), leading to induction of cytokine genes such as tumor necrosis factor alpha. Pretreatment with LPS causes desensitization to subsequent activation by CpG DNA. Both stimuli also cause cell cycle arrest in macrophages proliferating in response to the macrophage growth factor colony-stimulating factor-1 (CSF-1), but prevent apoptosis caused by growth factor removal. In part, cell cycle arrest by CpG DNA and LPS may be linked to rapid down-modulation of the CSF-1 receptor from the cell surface, a response that occurs in an all-or-nothing manner. The response of macrophages to CpG DNA has aspects in common with the DNA damage response in other cell types, which may provide clues to the underlying mechanism.
Activated macrophages regulate fibrogenesis by providing cytokines and growth factors that modulate the proliferation and collagen synthesis of fibroblasts. However, macrophages can be activated in a classical pathway induced by LPS or IFN-gamma and an alternative pathway induced by IL-4 or glucocorticoid. Differently activated macrophages display distinct biological features. To clarify the difference between these two subsets of macrophages in the regulatory mechanisms controlling fibrogenesis, human peripheral blood monocytes were used as the source of macrophages and cocultivation of differently activated macrophages and a fibroblast cell line, WI-38, was performed. Alternatively activated macrophages increased the proliferation index and collagen synthesis of cocultivated WI-38 cells in comparison to untreated monocytes, while classically activated macrophages markedly reduced collagen production of cocultivated WI-38 cells. Additionally, mRNA expression and protein production of TGF-beta(1), PDGF-AA, and PDGF-BB were elevated in alternatively activated macrophages in parallel to their profibrogenic effects. In contrast, expression and production of TNF-alpha, as well as MMP-7, were enhanced in classically activated macrophages. These findings suggested that alternatively activated macrophages enhance fibrogenesis of fibroblasts by providing profibrogenic factors, while classically activated macrophages inhibit fibrogenesis of fibroblasts by releasing antifibrogenic or fibrolytic factors.
The cytokine microenvironment is thought to play an important role in the generation of immunoregulatory cells. Nematode infections are commonly associated with Th2 cytokines and hyporesponsive T cells. Here we show that IL-4-dependent macrophages recruited in vivo by the nematode parasite Brugia malayi actively suppress the proliferation of lymphocytes on co-culture in vitro. These alternatively activated macrophages block proliferation by cell-to-cell contact, implicating a receptor-mediated mechanism. Further, the proliferative block is reversible and is not a result of apoptosis. Suppressed cells accumulate in the G1 and G2/M phase of the cell cycle. Interestingly, the G1 and G2/M block correlates with increased levels of Ki-67 protein, suggesting a mechanism that affects degradation of cell cycle proteins. We also show that, in addition to lymphocyte cell lines of murine origin, these suppressive cells can inhibit proliferation of a wide range of transformed human carcinoma lines. Our data reveal a novel mechanism of proliferative suppression induced by a parasitic nematode that acts via IL-4-dependent macrophages. These macrophages may function as important immune regulatory cells in both infectious and noninfectious disease contexts.
Apoptosis is induced in the course of immune responses to infectious agents. The last step of apoptosis is recognition and ingestion of the dying cells by phagocytes. Here, Marcela F. Lopes and colleagues discuss recent studies and argue that phagocytosis of apoptotic cells plays a previously unrecognized role in regulating the nature of immune responses against pathogens.
Intramuscular injection of synthetic oligodeoxynucleotides (ODN) expressing unmethylated CpG motifs trigger the rapid development of a local inflammatory response. In vitro studies demonstrate that macrophages exposed to CpG ODN up-regulate expression of mRNA encoding the chemokines MIP-1alpha, MIP-1beta, MIP-2, RANTES, JE/MCP-1, and IP-10. Within 6 h of in vivo administration, CpG ODN induce a significant increase in chemokine mRNA levels at the site of injection and draining lymph nodes. These chemokines may contribute to the migration and stimulation of inflammatory cells that contribute to the development of CpG ODN-induced immune responses.
Numerous observations have established a crucial role for phagocytic cells in host resistance to Salmonella. Activated macrophages rely on a complex array of oxygen-dependent antimicrobial molecules to inhibit or kill intracellular Salmonella. An initial oxidative bactericidal phase, which is dependent on the respiratory burst phagocyte oxidase (phox) is succeeded by a prolonged nitrosative bacteriostatic phase, which is dependent on inducible nitric oxide synthase (iNOS). The sequential contribution of phox and iNOS to anti-Salmonella innate immunity has been demonstrated both in vitro and in vivo. The temporal progression from the predominant production of reactive oxygen species to the production of nitrogen oxides could optimize the initial reduction in microbial burden while minimizing the immunopathological consequences of the host inflammatory response.
Activation of macrophages plays an important role in the host resistance against intracellular pathogens. Various mechanisms are employed to control the activation processes and limit tissue damage by factors produced by activated macrophages. One of these mechanisms is the production of macrophage-deactivating cytokines, such as tumour growth factor (TGF)-beta. The present study concerns the effects of TGF-beta on interferon (IFN)-gamma-induced activation of murine macrophages with respect to induction of toxoplasmastatic activity, and production of tumour necrosis factor (TNF)-alpha, prostaglandin E2 (PGE2) and reactive nitrogen intermediates (RNI). IFN-gamma activation of macrophages resulted in inhibition of T. gondii proliferation [mean fold increase (FI) = 1.8, control mean FI = 7.0]; polymyxin B had no effect on this activation. The IFN-gamma-induced toxoplasmastatic activity of macrophages was inhibited by TGF-beta (mean FI = 6.3), which was also found for the IFN-gamma-induced production of TNF-alpha, RNI and PGE2 by macrophages. We found that PGE2, which has macrophage deactivating properties, was not involved in the inhibition of macrophage activation by TGF-beta. The deactivating activities of TGF-beta on the IFN-gamma-induced toxoplasmastatic activity and production of RNI are mediated by inhibition of production of TNF-alpha. Addition of exogenous TNF-alpha during the incubation of macrophages with IFN-gamma and TGF-beta abrogated the deactivating activity of TGF-beta. In sum, the results demonstrate that inhibition of TNF-alpha production is a key factor in the TGF-beta-induced suppression of macrophage activation with respect to toxoplasmastatic activity and RNI production.
Chemokines are important mediators of macrophage and T-cell recruitment in a number of inflammatory pathologies, and chemokines expressed in atherosclerotic lesions may play an important role in mononuclear cell recruitment and macrophage differentiation. We have analyzed the expression of the linked chromosome 16q13 genes that encode macrophage-derived chemokine (MDC/CCL22), thymus- and activation-regulated chemokine (TARC/CCL17), and the CX(3)C chemokine fractalkine (CX(3)CL1) in primary macrophages and human atherosclerotic lesions by reverse transcription-polymerase chain reaction and immunohistochemistry. We show that macrophage expression of the chemokines MDC, fractalkine, and TARC is upregulated by treatment with the Th2-type cytokines interleukin-4 and interleukin-13. High levels of MDC, TARC, and fractalkine mRNA expression are seen in some, but not all, human arteries with advanced atherosclerotic lesions. Immunohistochemistry shows that MDC, fractalkine, and TARC are expressed by a subset of macrophages within regions of plaques that contain plaque microvessels. We conclude that MDC, fractalkine, and TARC, which are chromosome 16q13 chemokines, could play a role in mononuclear cell recruitment into atherosclerotic lesions and influence the subsequent inflammatory response. Macrophage-expressed chemokines upregulated by interleukin-4 may be useful surrogate markers for the presence of Th2-type immune responses in human atherosclerotic lesions.
Members of the SOCS (suppressor of cytokine signalling) family of proteins play key roles in the negative regulation of cytokine signal transduction. A series of elegant biochemical and molecular biological studies has revealed that these proteins act in a negative feedback loop, inhibiting the cytokine-activated Janus kinase/signal transducers and activators of transcription (JAK/ STAT) signalling pathway to modulate cellular responses. Although structurally related, the precise mechanisms of SOCS-1, SOCS-3 and cytokine-inducible SH2-containing protein (CIS) action vary. Direct interaction of SOCS SH2 domains with the JAK kinases or cytokine receptors allows their recruitment to the signalling complex, where they inhibit JAK catalytic activity or block access of the STATs to receptor binding sites. The defining feature of the family, the C-terminal SOCS box domain, appears dispensable for these actions but is likely to play a key role in negative regulation of signalling by targeting molecules associated with the SOCS proteins for degradation. The relevance of SOCS-mediated regulation of cytokine responses has been brought into sharp focus by the dramatic phenotypes of mice lacking these regulators. Indispensable roles for members of this family have been identified in the regulation of interferon gamma, growth hormone and erythropoietin, and the absence of SOCS-1 or SOCS-3 is lethal in mice.
An immune response can deviate toward either a Th1- or Th2-like response. In this work we examine the contribution that activated macrophages and IgG Abs make toward this deviation. The use of activated macrophages as APCs resulted in a strong polarized T cell response that was predominated by IFN-gamma. However, when Ag was targeted to FcgammaRs on these macrophages, the T cell response was reversed and biased toward a Th2-like response. This Th2-like phenotype was stable and was retained when the T cells were subsequently restimulated under nonbiasing conditions. The T cell biasing and its reversal via FcgammaR was also observed in vivo. Mice vaccinated with IgG-opsonized OVA made high levels of IgG Ab of the IgG1 isotype. These studies demonstrate that the ligation of FcgammaR on activated macrophages can reverse the Th1 biasing that occurs as a result of innate immune responses to microbial products.
Synthetic CpG containing oligodeoxynucleotide (CpG ODN) is recognized for its ability to activate cells to produce several cytokines, such as IL-12 and TNF-alpha. In the present study we have demonstrated that CpG ODN 1826, known for its immunostimulatory activity in the mouse system could, by itself, induce nitric oxide (NO) and inducible nitric oxide synthase (iNOS) production from mouse macrophage cell line (RAW 264.7). Neutralizing antibody against TNF-alpha was not able to inhibit NO or iNOS production from the CpG ODN 1826-activated macrophages, suggesting that although the TNF-alpha was also produced by CpG ODN-activated macrophages, the production of iNOS was not mediated through TNF-alpha. Although both CpG ODN 1826 and lipopolysaccharide (LPS) were able to stimulate NO and iNOS production, the exposure time required for maximum production of NO and iNOS for the CpG ODN 1826-activated macrophages was significantly longer than those activated with LPS. These results were due probably to a delay of NF-kappaB translocation, as indicated by the delay of IkappaBalpha degradation. Moreover, the fact that chloroquine abolished NO and iNOS production from the cells treated with CpG ODN 1826 but not from those treated with LPS suggested that the induction of NO and iNOS production from the cells stimulated with CpG ODN (1826) also required endosomal maturation/acidification.
The suppressors of cytokine signalling (SOCS) are a family of intracellular proteins, several of which have emerged as key physiological regulators of cytokine responses, including those that regulate the immune system. The SOCS proteins seem to regulate signal transduction by combining direct inhibitory interactions with cytokine receptors and signalling proteins with a generic mechanism of targeting associated proteins for degradation. Evidence is emerging for the involvement of SOCS proteins in diseases of the human immune system, which raises the possibility that therapeutic strategies that are based on the manipulation of SOCS activity might be of clinical benefit.
To determine whether monocyte chemoattractant protein-1 (MCP-1), which initiates subsequent development of burn-associated type 2 T cells, is produced in mice early after thermal injury.
A predominance of type 2 T-cell responses is commonly observed in animals and patients with severe thermal injuries. Burn-associated type 2 T cells have been identified as the cells responsible for the increased susceptibility of thermally injured mice to infections with herpes simplex virus type 1 and Candida albicans. Recently, the necessity of MCP-1 for the generation of type 2 T cells was shown in MCP-1 knockout mice. MCP-1 may have an important role in the increased susceptibility of thermally injured mice to various intracellular opportunistic pathogens.
The production of MCP-1 in sera or in cultures of various cells prepared from thermally injured mice was measured. Dual-chamber transwell cultures were performed to determine the influence of MCP-1-producing cells on the generation of burn-associated type 2 T cells.
Without any stimulation, splenic macrophages from mice (1/2D-M(phi)) produced MCP-1 into their culture fluids 12 hours after thermal injury. Interleukin-4 was detected in culture fluids of splenic T cells from normal mice cultured with 1/2D-M(phi) in a dual-chamber transwell system; however, this cytokine was not produced by normal T cells cultured with normal macrophages in the transwells. Also, normal T cells cultured with 1/2D-M(phi) did not produce interleukin-4 when transwell cultures were performed in the presence of anti-MCP-1 monoclonal antibody. Further, normal T cells directly stimulated with MCP-1 produced interleukin-4 into their culture fluids. Normal T cells, cultured with 1/2D-M(phi) for 24 hours in the transwells and recultured with fresh medium for an additional 7 days, produced interleukin-10 (but not interferon-gamma) and expressed ST2L mRNA (but not interleukin-12 receptor beta2 chain) when they were stimulated with anti-CD3 monoclonal antibody.
Results indicate that MCP-1 is produced in mice within 1 day of thermal injury, and the subsequent development of burn-associated type 2 T-cell responses may be triggered by MCP-1 produced early after thermal injury.
Mononuclear phagocytes are versatile cells that can express different functional programs in response to microenvironmental signals. Fully polarized M1 and M2 (or alternatively activated) macrophages are the extremes of a continuum of functional states. Macrophages that infiltrate tumor tissues are driven by tumor-derived and T cell-derived cytokines to acquire a polarized M2 phenotype. These functionally polarized cells, and similarly oriented or immature dendritic cells present in tumors, have a key role in subversion of adaptive immunity and in inflammatory circuits that promote tumor growth and progression.
The classical pathway of interferon-gamma-dependent activation of macrophages by T helper 1 (T(H)1)-type responses is a well-established feature of cellular immunity to infection with intracellular pathogens, such as Mycobacterium tuberculosis and HIV. The concept of an alternative pathway of macrophage activation by the T(H)2-type cytokines interleukin-4 (IL-4) and IL-13 has gained credence in the past decade, to account for a distinctive macrophage phenotype that is consistent with a different role in humoral immunity and repair. In this review, I assess the evidence in favour of alternative macrophage activation in the light of macrophage heterogeneity, and define its limits and relevance to a range of immune and inflammatory conditions.
The activation of a pro-inflammatory cascade after burn injury appears to be important in the development of subsequent immune dysfunction, susceptibility to sepsis and multiple organ failure. Macrophages are major producers of pro-inflammatory mediators and their productive capacity for these mediators is markedly enhanced following thermal injury. Thus, macrophage hyperactivity (as defined by increased productive capacity for pro-inflammatory mediators) appears to be of critical importance in the development of post-burn immune dysfunction. This review will focus on the current state of knowledge with regards to the role of macrophages in the development of post-burn immune dysfunction. Particular areas of discussion include: nitric oxide synthase (NOS) and cyclooxygenase (COX) enzyme systems, macrophages and the T-helper (Th)-1/Th-2 cytokine responses, alterations in macrophages signal transduction and a potential role for gamma/delta T-cells in the development of macrophage hyperactivity following thermal injury. A more comprehensive understanding of the relationship between macrophage activity and post-burn immune dysfunction will hopefully provide the basis for improved therapeutic regimes in the treatment of burn patients.
Macrophage heterogeneity used to be a research topic on which the careers of many postdoctoral fellows were misspent. The lack of definitive markers and dubious biochemical assays prevented the unequivocal identification of specific cell subsets. There has now been significant progress in establishing the heterogeneity of activated macrophages. There are at least three distinct populations of macrophages, and each cell type appears to have different biological roles. The interplay among these populations of cells may help to shape not only the magnitude but also the character of the immune response. The manipulation of these cells may lead to new approaches to treat or prevent disease.
Th2-specific chemokine thymus and activation-regulated chemokine (TARC)/CC chemokine ligand (CCL)17 is highly implicated in the pathogenesis of Th-2-dominated allergic diseases such as bronchial asthma (BA) and atopic dermatitis (AD). We performed polymorphism screening of the coding and promoter regions of the TARC gene. We found two rare variations in the coding region of exon 3 (2134C>T and 2037G>A) and a single nucleotide polymorphism (SNP) in the 5'-flanking region (-431C>T). Individuals carrying the 431T allele showed significantly increased serum levels of TARC compared with those not carrying the 431T allele, suggesting that this SNP has functional significance. However, when the genotypes at the SNP site were determined for 158 healthy individuals, 105 patients with BA and 148 patients with AD, we observed no significant association of the SNP with susceptibility to BA or AD.
Overexpression studies suggest that SOCS3 is a pleiotropic negative regulator of cytokines. The generation of Socs3(-/-) mice indicates that SOCS3 actually functions in a remarkably specific manner.