Modulation of Mycobacterium avium growth in murine macrophages: reversal of unresponsiveness to interferon-gamma by indomethacin or interleukin-4

The ability of soluble factors to modulate the growth of a virulent strain of Mycobacterium avium in murine peritoneal macrophages was studied. The virulent strain, TMC 702, grew progressively in the organs of susceptible BALB/C mice. In addition, this strain of M. avium grew progressively in untreated peritoneal macrophages. Treatment of macrophage monolayers with interferon-gamma (IFN-gamma) did not change significantly the intracellular growth of M. avium. Addition of indomethacin to IFN-gamma-treated macrophage monolayers rendered them significantly more bacteriostatic than macrophages treated with interferon alone, suggesting a role for prostaglandins in inducing unresponsiveness to IFN-gamma in infected cells. Additionally, treatment with tumour necrosis factor-alpha led to a modest increase in bacteriostasis, as compared to untreated monolayers. Further experiments with recombinant interleukins showed that interleukin-4 (IL-4), on its own, could increase bacteriostatic activity against M. avium in a reproducible fashion. Experiments with interleukin combinations showed that IFN-gamma and IL-4 treatment of macrophages rendered these cells almost fully bacteriostatic against M. avium, inclusion of scavengers of reactive oxygen species did not modify the beneficial effect of IFN-gamma and IL-4. Overall, our results suggest an important role for interleukins in modulating the interaction between virulent mycobacteria and murine macrophages.     

Interleukin-12-stimulated natural killer cells can activate human macrophages to inhibit growth of Mycobacterium avium.

Interleukin-12 (IL-12) is a critical cytokine that affects many of the biological functions of NK cells and T cells. We have previously shown that both human and murine NK cells are important in host defense against Mycobacterium avium complex and act by secreting cytokines that induce macrophages to inhibit the growth of intracellular M. avium. To define the role of IL-12 in M. avium complex infection, we stimulated human NK cells with recombinant human IL-12 at 0.01 to 1 ng/ml for 24 h and used the tissue culture supernatant to treat human monocyte-derived macrophage monolayers infected with M. avium. IL-12 had no direct effect on M. avium-infected macrophages, but culture supernatant from IL-12-treated NK cells activated macrophages to inhibit the growth of intracellular M. avium in a dose-dependent manner. Stimulation of NK cells with IL-12 in combination with tumor necrosis factor alpha (TNF-alpha) or IL-1 increased the ability of supernatant from NK-cell culture to limit M. avium growth within macrophages, compared with that of culture supernatant from IL-12-treated NK cells. Results with supernatant from nonstimulated NK cells were similar to those with supernatant from untreated controls. Treatment of supernatant from IL-12-stimulated NK cells with anti-TNF-alpha, anti-granulocyte-macrophage colony-stimulating factor, but not anti-gamma interferon antibodies decreased the ability of NK-cell supernatant to induce anti-M. avium activity in infected macrophages. Treatment of macrophage monolayers with anti-transforming growth factor beta antibody before adding supernatant from IL-12-stimulated NK cells was associated with an increase of anti-M. avium activity compared with that of supernatant from IL-12-treated NK cells. These results suggest that IL-12 has a role in host defense against M. avium and that the effect of IL-12 is dependent chiefly on TNF-alpha and granulocyte-macrophage colony-stimulating factor.     

Human peritoneal macrophage activity is increased by tuftsin

Peritonitis caused by Candida albicans is a major complication of continuous ambulatory peritoneal dialysis (CAPD). Increasing the activity of the peritoneal macrophages--the predominant cell type found in the peritoneal cavity--may be of great importance in the prevention and therapy of peritonitis. Therefore, the activating effect of tuftsin was studied on human peritoneal macrophages from CAPD patients. Tuftsin induced a biphasic effect on macrophage activity within a range of 2 X 10(-9)-2 X 10(-6) M, with a maximal activity of 2 X 10(-7) M. At this concentration, tuftsin enhanced by twofold cell association with radiolabelled candida (from 2 +/- 0.2 to 4 +/- 0.2 candida per macrophage) and superoxide anion production in response to exposure to candida (from 150 +/- 20 to 300 +/- 20 nmoles/mg). These results suggest the potential use of tuftsin as a therapeutic drug.     

Cytokine modulation of Mycobacterium tuberculosis growth in human macrophages

This study was concerned with the handling of ingested tubercle bacilli by normal human macrophages. Intracellular growth was determined after exposure of macrophages to viable bacilli in vitro and the effect of various cytokines, alone or in combination, on bacilli growth/survival was determined. It was found that Mycobacterium tuberculosis (M.tb) grew quite readily in untreated cultured human macrophages. Treatment with soluble factors showed that a crude lymphokine containing supernatant elicited with Concanavalin A (Con A) was ineffective at reducing growth of M.tb in vitro; similarly a crude lymphokine preparation from M.tb lysate-stimulated mononuclear cells failed to induce any mycobacteriostatic activity in human monocyte-derived macrophages. Recombinant cytokines were then evaluated for their ability to modulate growth of the tubercle bacilli in human macrophages. Recombinant interferon-gamma (IFN-gamma), interleukin-2 (IL-2) and recombinant interleukin 4 (IL-4) were all ineffective at modifying M. tuberculosis growth in human macrophages. Recombinant tumour necrosis-alpha (TNF-alpha) curbed the growth of the bacilli in human macrophages in a reproducible fashion. No cytokine combination was more efficient than TNF-alpha alone. These studies thus highlight the resistance of virulent mycobacteria against different mechanisms of cytokine-induced macrophage bactericidal activity.     

Macrophage polarization in response to wear particles in vitro

Total joint replacement is a highly successful surgical procedure for treatment of patients with disabling arthritis and joint dysfunction. However, over time, with high levels of activity and usage of the joint, implant wear particles are generated from the articulating surfaces. These wear particles can lead to activation of an inflammatory reaction, and subsequent bone resorption around the implant （periprosthetic osteolysis）. Cells of the monocyte/macrophage lineage orchestrate this chronic inflammatory response, which is dominated by a pro-inflammatory （M 1） macrophage phenotype rather than an anti-inflammatory pro-tissue healing （M2） macrophage phenotype. While it has been shown that interleukin-4 （IL-4） selectively polarizes macrophages towards an M2 anti-inflammatory phenotype which promotes bone healing, rather than inflammation, little is known about the time course in which this occurs or conditions in which repolarization through I L-4 is most effective. The goal of this work was to study the time course of murine macrophage polarization and cytokine release in response to challenge with combinations of polymethyl methacrylate （PMMA） particles, lipopolysaccharide （LPS） and IL-4 in vitro. Treatment of particle-challenged monocyte/macrophages with IL-4 led to an initial suppression of pro-inflammatory cytokines and inducible nitric oxide synthase （iNOS） production and subsequent polarization into an M2 anti-inflammatory phenotype. This result was optimized when IL-4 was delivered before PMMA particle challenge, to an M 1 phenotype rather than to uncommitted （MO） macrophages. The effects of this polarization were sustained over a 5-day time course. Polarization of M1 macrophages into an M2 phenotype may be a strategy to mitigate wear particle associated periprosthetic osteolysis.     

Modulation of human lepromatous monocyte-macrophage functions in vitro by Tuftsin

Human peripheral blood monocytes/macrophages derived from normal donors, patients of tuberculoid leprosy (BT/TT) and lepromatous leprosy (BL/LL) were assayed for stimulated phagocytic responses to the potent macrophage stimulator “Tuftsin” (NH₂ThrLysProArgOH) after varying periods (6 h to 14 days) of culture in vitro. The assaysconsisted of visual scoring of ingested Mycobacterium leprae and radiometric measurement of ingested ¹⁴C-acetate labelled Staphylococcus aureus and Mycobacterium tuberculosis (H₃₇R_a). While normal and BT/TT macrophages showed a progressively increasing ability for tuftsin-stimulated phagocytosis with increasing age of culture in vitro, BL/LL macrophages showed the opposite response so that 14-day cultures were refractory to a stimulatory dose of up to 7.0 μM (10 to 20 times the optimal dose for normal and BT/TT macrophages). The 14-day BL/LL macrophage cultures were, however, responsive to 35 μM tuftsin (100 times the optimal dose for normal macrophages). Analysis of the dose-response curves also indicates that BT/TT cultures despite exhibiting an apparent similarity to normal macrophages demonstrate a rightward shift for a maximal stimulated phagocytosis. Finally SEM photo-micrographs of 14-day macrophage cultures of the three groups revealed that while normal and BT/TT cultures demonstrated an increase in membrane ruffling and filopodia on stimulation with 0.8 μM tuftsin, BL/LL cultures exhibited none of the features associated with stimulation. From the above findings, we conclude that lepromatous macrophages may display an aberrant differentiation profile leading to a terminal state of unresponsiveness and that the defect may possibly lie at the level of tuftsin receptor expression or transmembrane signal transduction.     

Alternative Reprogramming of M1/M2 Phenotype of Mouse Peritoneal Macrophages In Vitro with Interferon-γ and Interleukin-4

An important role in the development of the immune response is played by macrophages that acquire either anti-inflammatory M1 or anti-inflammatory M2 phenotype depending on their microenvironment. The possibility of targeted reprogramming of the initial M2 macrophage phenotype towards M1 phenotype and vice versa using macrophage reprogramming factors IFN-γ and IL-4, respectively, was demonstrated. We showed that macrophages of genetically different mouse strains did not practically differ by their reprogramming capacity. Our findings suggest that macrophage programming not only participates in the triggering of the immune response, but also can ensure plasticity of functional activity during the developing response.     

Claudin-1, claudin-2 and claudin-11 genes differentially associate with distinct types of anti-inflammatory macrophages in vitro and with parasite- and tumour-elicited macrophages in vivo

Macrophages altered by various Th2-associated and anti-inflammatory mediators--including IL-4 and IL-13 [inducing alternatively activated macrophages (AAMs)], IL-10 and TGF-β--were generically termed M2. However, markers that discriminate between AAMs and other M2 remain scarce. We previously described E-cadherin as a marker for AAMs, permitting these macrophages to fuse upon IL-4 stimulation. To identify novel potential contributors to macrophage fusion, we assessed the effect of IL-4 on other adherens and tight junction-associated components. We observed an induction of claudin-1 (Cldn1), Cldn2 and Cldn11 genes by IL-4 in different mouse macrophage populations. Extending our findings to other stimuli revealed Cldn1 as a mainly TGF-β-induced gene and showed that Cldn11 is predominantly associated with IL-4-induced AAMs. Cldn2 is upregulated by diverse stimuli and is not associated with a specific macrophage activation state in vitro. Interestingly, different claudin genes preferentially associate with M2 from distinct diseases. While Cldn11 is predominantly expressed in AAMs from helminth-infected mice, Cldn1 is the major macrophage claudin during chronic trypanosomiasis and Cldn2 dominates in tumour-associated macrophages. Overall, we identified Cldn1, Cldn2 and Cldn11 as genes that discriminate between diverse types of M2.     

Immunological Effects of d-Penicillamine during Experimentally Induced Inflammation in Rats

Administration of d -penicillamine (50 mg/kg/day orally) to rats with adjuvant arthritis for up to 42 days significantly modified the incorporation of ³H-thymidine (³H-TdR) in concanavalin A (Con A)-stimulated lymph node cells. Treatment with d -penicillamine abolished the ability of macrophages from arthritic rats to inhibit lymphocyte responsiveness to Con A and lipopolysaccharide (LPS) 14 days after the induction of the disease. Increased T-cell responsiveness to Con A was found from day 14 to day 35 in cultures of unseparated and adherent-cell-depleted lymph node cells from d -penicillamine-treated arthritic rats. B-cell responsiveness to LPS was not affected. Experiments with bovine serum albumin gradient-separated lymph node cells confirmed these findings and indicated that treatment with d -penicillamine may specifically enhance T-helper cell responsiveness to Con A. It is suggested that administration of d -penicillamine may interfere with macrophage function during the course of an immunologically induced chronic inflammation, leading to an increased response of T-helper cells. The theoretical implications of these findings are discussed.     

慢性华支睾吸虫感染对脂多糖刺激巨噬细胞反应的影响

 目的：了解慢性华支睾吸虫（Cs）感染的免疫状态及其对巨噬细胞表型和炎症反应的影响。方法：选择Sprague-Dawley雄性大鼠,按随机数字表法分组进行2部分实验。(1) 经口华支睾吸虫虫卵灌胃感染70 d建立慢性Cs感染模型,分为正常对照组和慢性Cs感染组,每组10只,用酶联免疫吸附法（ELISA）检测大鼠血清白细胞介素 4（IL-4)、IL-10、肿瘤坏死因子α（TNF-α）和干扰素γ（IFN-γ）水平,腹腔灌洗获取巨噬细胞,流式细胞术检测腹腔巨噬细胞亚型;(2)腹腔灌洗获取巨噬细胞,按照腹腔巨噬细胞的来源,分为空白对照组、正常组和慢性Cs感染组,以脂多糖（LPS,10 μg/L）刺激孵育,用ELISA法动态监测LPS刺激后0、2、12和24 h细胞培养上清液的TNF-α和IL-10的变化。结果：(1) 慢性Cs感染组大鼠血清促炎因子TNF-α［(77.64±3.25) ng/L］和IFN-γ［(212.69±12.60)  ng/L］水平较正常组TNF-α［(55.13±3.25) ng/L］和IFN-γ［(108.15±10.49)  ng/L］升高（均P＜0.05）,而抗炎因子IL-4［(248.24±8.99) ng/L］和IL-10［(223.56±5.60) ng/L］水平较正常组血清IL-4［(52.40±3.97) ng/L］和IL-10［(60.18±5.36) ng/L］也显著升高（均P＜0.01）,并可使腹腔巨噬细胞向替代活化型（M2型）巨噬细胞分化;(2) 2组腹腔巨噬细胞培养上清液TNF-α水平在LPS刺激后2 h即开始上升,至24 h达到高峰,慢性Cs感染组巨噬细胞在LPS刺激后2、12和24 h上清液的TNF-α水平均显著低于正常大鼠组［2 h： （88.20±1.91） ng/L vs（123.45±2.98） ng/L;12 h：（123.66±4.31） ng/L vs（161.11±7.38） ng/L;24 h：（154.52±4.83） ng/L vs（227.85±10.67） ng/L,均P＜0.05］,而IL-10的水平较正常组升高［2 h：（105.46±12.28） ng/L vs（80.86±2.28） ng/L;12 h：（194.19±8.62） ng/L vs（117.56±8.02） ng/L;24 h：（280.02±11.31） ng/L vs（168.14±8.97） ng/L,均P＜0.05］。结论：慢性Cs感染时宿主血清的促炎因子升高,而抗炎因子升高得更明显,使巨噬细胞向M2型极化;体外实验表明慢性Cs感染宿主的巨噬细胞对LPS刺激产生耐受。     

The enhanced immune response of hepatitis B virus DNA vaccine using SiO2@LDH nanoparticles as an adjuvant

Various approaches have been used to improve systemic immune response to infectious disease or virus, and DNA vaccination has been demonstrated to be one of these effective ways to elicit protective immunity against pathogens. Our previous studies showed that layered double hydroxides (LDH) nanoparticles could be efficiently taken up by the MDDCs and had an adjuvant activity for DC maturation. To further enhance the immune adjuvant activity of LDH, core–shell structure SiO₂@LDH nanoparticles were synthesized with an average diameter of about 210 nm. And its high transfection efficiency in vitro was demonstrated by using GFP expression plasmid as model DNA. Exposing SiO₂@LDH nanoparticles to macrophages caused a higher dose-dependent expression of IFN-γ, IL-6, CD86 and MHC II, compared with SiO₂ and LDH respectively. Furthermore, in vivo immunization of BALB/c mice indicated that, DNA vaccine loaded-SiO₂@LDH nanoparticles not only induced much higher serum antibody response than naked DNA vaccine and plain nanoparticles, but also obviously promoted T-cell proliferation and skewed T helper to Th1 polarization. Additionally, it was proved that the caveolae-mediated uptake of SiO₂@LDH nanoparticles by macrophage lead to macrophages activation via NF-κB signaling pathway. Our results indicate that SiO₂@LDH nanoparticles could serve as a potential non-viral gene delivery system.     

Recombinant tumour necrosis factor-alpha decreases whereas recombinant interleukin-6 increases growth of a virulent strain of Mycobacterium avium in human macrophages.

The ability of a virulent strain of Mycobacterium avium to infect and replicate within human monocyte-derived macrophages of normal donors was assessed. Moreover, the ability of selected cytokines to modulate the intracellular growth of M. avium was investigated. Our virulent strain of M. avium grew progressively in human macrophages. Treatment of macrophage monolayers with interferon-gamma (IFN-gamma) did not lead to any significant change in the infection pattern. Conversely, treatment with tumour necrosis factor-alpha (TNF-alpha) led to a significant reduction in the growth of M. avium in the macrophages. In contrast, treatment of macrophages with interleukin-6 (IL-6) enhanced their susceptibility to M. avium significantly. This finding was substantiated by other results which showed that IL-6 increased the growth of M. avium in tissue culture medium. These results suggest that cytokines may influence the M. avium-macrophage interaction, in a positive or negative manner.     

MiR-146a modulates macrophage polarization in systemic juvenile idiopathic arthritis by targeting INHBA

Monocytes from patients with systemic juvenile idiopathic arthritis (SJIA) have both features of classical activated M1 and alternatively activated M2 macrophages. An increasing number of studies have indicated that microRNAs (miRNAs) are critical regulators of monocyte polarization. Here, we focused on miR-146a expression in SJIA and investigated the function of miR-146a in monocyte polarization. We found that miR-146a expression was highly up-regulated in SJIA monocytes and correlated with the systemic features. miR-146a was expressed at a higher level in monocytes polarized with M2 conditions than those polarized with M1 conditions. miR-146a overexpression significantly decreased the production of M1 phenotype markers such as IL-6, IL-12, TNF-α, CD86 and iNOS in M1 macrophages, but increased the production of M2 marker genes such as Arg1, CCL17, CCL22 and CD206 in M2 macrophages. Conversely, knockdown of miR-146a promoted M1 macrophage polarization but diminished M2 macrophage polarization. We subsequently demonstrated that miR-146a targeted the 3′-untranslated region (UTR) of INHBA to inhibit its expression. Additionally, INHBA overexpression rescued the reduced IL-6, IL-12, and TNF-α levels induced by miR-146a overexpression in M1 macrophages, and rescued the increased Arg1, CCL17, and CCL22 levels induced by miR-146a overexpression in M2 macrophages. Similarly, the effects of miR-146a inhibition in monocyte polarization were all partly reversed by INHBA inhibition. Taken together, the data suggest that miR-146a serves as a molecular regulator in monocyte polarization and might play an important role in monocytes from patients with SJIA.     

Heme Oxygenase-1 expression in M-CSF-polarized M2 macrophages contributes to LPS-induced IL-10 release

The shift between pro-inflammatory (M1) and anti-inflammatory (M2) states of macrophage polarization allows the resolution of inflammatory processes as well as the maintenance of a basal anti-inflammatory environment in tissues continuously exposed to harmless antigens (e.g., lung and gut). To identify markers for the anti-inflammatory state of macrophages, expression profiling was performed on human macrophages polarized by either GM-CSF or M-CSF, which lead to the generation of TNF-α and IL-12p40-producing pro-inflammatory macrophages [M1 (GM-CSF)] or IL-10-producing anti-inflammatory macrophages [M2 (M-CSF)] upon exposure to LPS, respectively. A different iron metabolism gene signature was detected in both macrophage types, with the heme regulatory molecules CD163 and Heme Oxygenase-1 (HO-1) being preferentially expressed by M2 (M-CSF) macrophages. M1-polarizing cytokines (GM-CSF, IFNγ) inhibited, while IL-4 enhanced, the M-CSF-driven HO-1 expression. In agreement with this in vitro data, HO-1 expression in metastatic melanoma was primarily detected in CD163⁺ tumor-associated macrophages, which are known to exhibit an M2-skewed polarization phenotype. In contrast to the HO-1 inhibitor tin protoporphyrin (SnPP), the administration of cobalt protoporphyrin (CoPP), a potent inducer of HO-1 resulted in increased LPS-triggered IL-10 release from M2 (M-CSF) macrophages. The data suggests that HO-1 is important for the anti-inflammatory activities of M-CSF-polarized M2 macrophages. Moreover, since M2 (M-CSF) macrophages also express higher levels of the CD163 scavenger receptor, the CD163/HO-1/IL-10 axis appears to contribute to the generation of an immunosuppressive environment within the tumor stroma.     

Inflammatory response under zinc deficiency is exacerbated by dysfunction of the T helper type 2 lymphocyte–M2 macrophage pathway

Nutritional zinc deficiency leads to immune dysfunction and aggravates inflammation. However, the underlying mechanism remains unknown. In this study, the relationship between macrophage subtypes (M1 and M2) and helper T lymphocytes (Th1 and Th2) was investigated using the spleen from rats fed zinc-deficient or standard diet. In experiment I, 5-week-old male Sprague-Dawley rats were fed a zinc-deficient diet (without zinc additives) or a standard diet (containing 0·01% zinc) for 6 weeks. In experiment II, the rats were divided into four groups: one group was fed a standard diet for 6 weeks; two groups were fed zinc-deficient diets and were injected three times a week with either saline or interleukin-4 (IL-4) (zinc-deficient/IL-4 i.p.); a fourth group (zinc-deficient/standard) was fed a zinc-deficient diet for 6 weeks followed by a standard diet for 4 weeks. In experiment I; GATA-binding protein 3 (GATA-3) protein level, M2 macrophage, CD3⁺ CD8⁺ cells, and IL-4/IL-13-positive cells significantly decreased in the spleens of the zinc-deficient group. Additionally, IL-1β and macrophage inflammatory protein-1α (MIP-1α) mRNA levels significantly increased in the splenic macrophages of the zinc-deficient group. In experiment II; M2 macrophages, CD3⁺ CD8⁺ cells, IL-4/IL-13-positive cells, and GATA-3 protein levels significantly increased in the spleens of the zinc-deficient/IL-4 i.p. and zinc-deficient/standard groups. Furthermore, IL-1β and MIP-1α mRNA levels decreased in the splenic macrophages of the zinc-deficient/IL-4 i.p. and zinc-deficient/standard groups. Zinc deficiency-induced aggravated inflammation is related to Th2 lymphocytes and followed by the association with loss of GATA-3, IL-4 and anti-inflammatory M2 macrophages. Importantly, IL-4 injection or zinc supplementation can reverse the effects of zinc deficiency on immune function.     

Interleukin-10 reduces the pathology of mdx muscular dystrophy by deactivating M1 macrophages and modulating macrophage phenotype

M1 macrophages play a major role in worsening muscle injury in the mdx mouse model of Duchenne muscular dystrophy. However, mdx muscle also contains M2c macrophages that can promote tissue repair, indicating that factors regulating the balance between M1 and M2c phenotypes could influence the severity of the disease. Because interleukin-10 (IL-10) modulates macrophage activation in vitro and its expression is elevated in mdx muscles, we tested whether IL-10 influenced the macrophage phenotype in mdx muscle and whether changes in IL-10 expression affected the pathology of muscular dystrophy. Ablation of IL-10 expression in mdx mice increased muscle damage in vivo and reduced mouse strength. Treating mdx muscle macrophages with IL-10 reduced activation of the M1 phenotype, assessed by iNOS expression, and macrophages from IL-10 null mutant mice were more cytolytic than macrophages isolated from wild-type mice. Our data also showed that muscle cells in mdx muscle expressed the IL-10 receptor, suggesting that IL-10 could have direct effects on muscle cells. We assayed whether ablation of IL-10 in mdx mice affected satellite cell numbers, using Pax7 expression as an index, but found no effect. However, IL-10 mutation significantly increased myogenin expression in vivo during the acute and the regenerative phase of mdx pathology. Together, the results show that IL-10 plays a significant regulatory role in muscular dystrophy that may be caused by reducing M1 macrophage activation and cytotoxicity, increasing M2c macrophage activation and modulating muscle differentiation.     

Aire 对巨噬细胞极化影响的研究

目的:研究自身免疫调节因子(Aire)对巨噬细胞极化的影响。方法:分别用LPS、IL-4 以及LPS 联合免疫复合物刺激小鼠单核巨噬细胞系RAW264郾7 细胞、稳定表达GFP-Aire 的RAW264.7 细胞(A33-3) 细胞和稳定表达GFP 的RAW264.7 细胞(C1-6),使其向M1(LPS)、M2a(IL-4)和M2b(LPS 联合免疫复合物)型巨噬细胞极化。通过Real-time PCR 检测各组细胞中M1 型巨噬细胞特征分子IL-1、iNOS 和IL-6,M2a 型特征分子Arg-1 和M2b 型特征分子IL-10 的表达水平,研究Aire 对各种类型巨噬细胞极化的影响。结果:LPS 在0.5 g/ ml 浓度时,RAW264.7 细胞中M1 型巨噬细胞产物IL-1 、iNOS和IL-6 基因表达量最高;而IL-4 以及LPS 联合免疫复合物的刺激作用有显著的剂量依赖性,都在浓度最高时RAW264.7 细胞中Arg1(M2a)和IL-10(M2b)基因表达量最高。LPS 刺激后,A33-3 细胞中IL-1 和iNOS 表达水平明显高于C1-6 细胞,IL-6 则相反;IL-4 及LPS 联合免疫复合物刺激后,A33-3 细胞中Arg1 和IL-10 的表达水平明显低于C1-6 细胞。结论:Aire 可能促进巨噬细胞向M1 极化,同时抑制其向M2a 和M2b 极化。     

Vascular endothelial growth factor-induced skin carcinogenesis depends on recruitment and alternative activation of macrophages

Inflammation contributes to tumour growth, invasion and angiogenesis. We investigated the contribution of macrophages and their polarization to tumour progression in a model of VEGF-A-induced skin carcinogenesis. Transfection of the human non-tumourigenic keratinocyte cell line HaCaT with murine VEGF-A leads to malignant tumour growth in vivo. The resulting tumours are characterized by extensive vascularization, invasive growth and high numbers of M2-polarized macrophages that crucially contribute to the establishment of the malignant phenotype. Accordingly, macrophage depletion from tumour-bearing animals resulted in reduced tumour growth, inhibition of invasion, decreased proliferation and reduced angiogenesis. In vitro, VEGF-A exerted a chemo-attracting effect on macrophages, but did not induce M2 polarization. We identified IL-4 and IL-10 as the factors involved in M2 polarization. These factors were produced by tumour cells (IL-10) and macrophages (IL-4) in vivo. Addition of recombinant IL-4 and IL-10 in vitro induced a pro-invasive M2 macrophage phenotype and inhibition of the IL-4 receptor in vivo blocked M2 polarization of macrophages, resulting in a less aggressive tumour phenotype. Thus, we provide evidence that M2 macrophages are crucial for the development of VEGF-A-induced skin tumours and that VEGF-A contributes to malignant tumour growth, not only by enhancing angiogenesis but also by establishing an anti-inflammatory microenvironment. However, VEGF-A alone is not sufficient to create a tumour-promoting microenvironment and requires the presence of IL-4 and IL-10 to induce M2 polarization of macrophages.     

Augmentation of oxidative and arachidonate metabolism in macrophages by tuftsin (Thr-Lys-Pro-Arg)

Conclusions We have obtained evidence for novel effects of the naturally occurring tetrapeptide tuftsin. It induces an oxidative burst inC. parvum-elicited mø and excites the release of the arachidonic acid cyclo-oxygenation product TXB₂ from albumin-elicited mø.Considering the role that has been ascribed to tuftsin in augmenting tumoricidal capacities of mø, tuftsin-evoked release of superoxide anion and hydrogen species generated in the oxidative burst have been implicated as effector molecules in microbial and tumor cell killing by macrophages [4].Arachidonic acid conversion yielding eicosanoids such as TXB₂, another early sequela of macrophage membrane perturbation, is also enhanced in response to tuftsin. Whether this action bears significance in the context of macrophage-platelet interaction in inflammation remains to be elucidated. Equally, it needs to be determined whether other pro-inflammatory compounds derived from arachidonic acid are produced by macrophages upon contact with tuftsin. In any case, tuftsin has to be added to the list of endogenous and exogenous substances capable of activating macrophage.