槲寄生凝集素腹腔内给药对腹腔巨噬细胞功能的影响

目的：探讨小鼠腹腔内注射槲寄生凝集素对腹腔巨噬细胞功能的影响。方法：16只小鼠分为腹腔内给药组和空白组,分别取腹腔内巨噬细胞并计数,采用ELISA法测定TNF-α表达情况,Griess法测定NO表达,MTT法检测腹腔内巨噬细胞对肠癌HCT116细胞的杀伤作用。结果：腹腔内给槲寄生凝集素后可明显增加腹腔内巨噬细胞数,并增加TNF—α和NO的表达水平;激活的巨噬细胞对HCT116细胞的杀伤作用增强,其杀伤作用与TNF—α和NO表达时间一致。结论：腹腔内注射槲寄生凝集素可激活腹腔内巨噬细胞,激活的巨噬细胞对HCT116细胞有杀伤作用。     

Interaction Between Polyalkylcyanoacrylate Nanoparticles and Peritoneal Macrophages: MTT Metabolism,NET Reduction,and NO Production

Purpose. The nature of interactions between macrophages and drug carriers is of primordial importance either in the design of more effective therapeutic strategies for macrophage-associated pathogenesis or in establishing new approaches for pharmacological action avoiding macrophages. Methods. Polyalkylcyanoacrylate nanoparticles (PMCA, PECA, PBCA and PIBCA nanoparticles) were assayed for their toxicity on peritoneal resident and thioglycolate-elicited macrophages. Cellular viability was assessed by MTT tetrazolium salt assay, oxidative burst by NBT reduction and NO production by nitrite evaluation. Results. The nanoparticles tested led to cellular morphological modifications and induced toxicity in both types of macrophages in culture. The polyalkylcyanoacrylate nanoparticles uptake by peritoneal macrophages caused an increase in respiratory burst, as assessed by the NBT reduction assay, and induced the release of soluble toxic factors to the culture medium. The association of LPS with the PMCA nanoparticles significantly stimulated the production of nitric oxide (NO) by resident macrophages. In contrast, the association of PBCA nanoparticles with LPS does not increase the nitrite production as compared with LPS alone, which may be due to a different physico-chemical interaction between LPS and the two types of polymers. Conclusions. In cultured mice peritoneal macrophages, nanoparticles of PACA induce the production of oxygen reactive products, which cause changes in the cell metabolism of both resident and elicited macrophages. PMCA nanoparticles in association with LPS significantly increase the expression of the inducible isoform of nitric oxide synthase, leading to the release of large amount of NO, which may be highly cytotoxic to the cultured cells in the presence of peroxide generated from the oxidative burst.     

海带多糖对小鼠腹腔巨噬细胞的激活作用

本文研究了海带多糖对Ｃ５７ＢＬ／６小鼠腹腔巨噬细胞的激活作用,结果表明,腹腔注射海带多糖能够明显激活小鼠腹腔巨噬细胞,增强其细胞溶解作用。海带多糖激活小鼠腹腔巨噬细胞,在有ＬＰＳ存在的条件下,能在体外分泌肿瘤坏死因子。     

Extract from <Emphasis Type="Italic">Calotropis procera</Emphasis> latex activates murine macrophages

Calotropis procera latex has long been used in traditional medicines. Extracts from C. procera latex have been reported to have various pharmacological actions, including protection from myocardial infarction, hepatoprotective action, antitumor activity, antinociceptive, and pro- and anti-inflammatory actions. To evaluate the immunomodulatory functions of the water-soluble C. procera extract (CPE), we investigated its ability to activate macrophages—effector cells in inflammatory and immune responses. Intraperitoneal injection of CPE in mice (2 mg/mouse) induced migration of macrophages to the intraperitoneal cavity, confirming the proinflammatory effects of water-soluble CPE. The direct effects of CPE on macrophages were then assessed by measuring the production of nitric oxide (NO) as an indicator for macrophage activation. Addition of CPE (1–10 μg/ml) to the culture medium of the murine monocyte/macrophage cell line RAW264.7 caused an increase in NO production in a time- and dose-dependent manner. CPE-elicited NO production was blocked by application of an inhibitor of inducible nitric oxide synthase (iNOS). Expression of iNOS mRNA was induced by treatment of cultured macrophages with CPE. Injection of CPE in mice also resulted in an increase in plasma NO level. The results suggest that CPE activates macrophages and facilitates NO production via up-regulation of iNOS gene expression. A. l. S. Seddek and M. Elsayed Mahmoud contributed equally to this work.     

Regulation of macrophage function by the antioxidant N-acetylcysteine in mouse-oxidative stress by endotoxin

Changes in several functions of peritoneal macrophages from mice with oxidative stress caused by intraperitoneal injection of endotoxin (Escherichia coli lipopolysaccharide, LPS) (100 mg/kg), and associated with a high production of reactive oxygen species (ROS), have been observed in our previous studies. Antioxidants such as N-acetylcysteine (NAC) are free radical scavengers that improve and modulate the immune response, especially in oxidative stress situations. Therefore, in the present work, we have studied the effects of the administration of NAC (150 mg/kg i.p.) on different functions of peritoneal macrophages from Swiss mice suffering that oxidative stress, caused by LPS (100 mg/kg). NAC was injected 30 min after LPS injection, and the peritoneal macrophages were obtained at 2, 4, 12, and 24 h after endotoxin injection. The following functions, key stages of the phagocytic process, were studied: adherence to substrate, chemotaxis, ingestion of particles, and production of ROS (reactive oxygen species), as well as tumor necrosis factor (TNFalpha) release. The decrease in chemotaxis and the increase in adherence, ingestion, superoxide anion production, and TNFalpha release shown by macrophages from animals with oxidative stress were counteracted by NAC injection. These data suggest that NAC administration may be useful for the treatment of oxidative stress-linked endotoxic shock, modulating the function of macrophages, specifically in decreasing the production of ROS and of inflammatory cytokines such as TNFalpha.     

Liposomes as immunomodulator--inhibitory effect of liposomes on NO production from macrophages

Liposomes have been artificially made into membranous vesicles composed essentially of naturally occurring phospholipids and have been found to serve as a carrier of drugs and an immunological adjuvant. After being intravenously injected, they are quickly removed from the blood circulation and trapped by Kupffer cells of the liver and macrophages of spleen. However, the changes liposomes exert in these cells with which liposomes interact remain unresolved. To clarify this point is very important to assure the safe use of liposomes as drug carriers. Macrophages have many unique functions, and nitric oxide (NO) produced by NO synthase (NOS) which is induced in response to some cytokines and bacterial products such as lipopolysaccharide (LPS) is responsible for the bactericidal, tumoricidal and immune regulatory activities. On the other hand, overexpressed NO is implicated in the development of atherosclerosis, DNA injury, and hypotension associated with septic shock. This article focuses on the effects of liposomes on NO production from LPS-stimulated mice peritoneal macrophages in vitro; we found that liposomes composed of phosphatidylserine inhibit NO production. We also discuss the mechanism of the inhibitory activity of liposomes.     

灵芝多糖肽对小鼠腹腔巨噬细胞一氧化氮产生的影响

目的　研究灵芝多糖肽 (GLPP)对小鼠腹腔巨噬细胞一氧化氮产生的影响并探讨其作用机制。方法　以Griess法 ,观察GLPP对LPS诱导小鼠腹腔巨噬细胞一氧化氮(NO)产生的影响 ;以免疫组化法检测诱导型一氧化氮合成酶 (iNOS)的表达 ,观察GLPP对iNOS的影响。结果　GLPP(2 5～ 2 0 0mg·kg-1)灌胃给药 5d或体外给药 (3 12 5～ 2 0 0mg·L-1)均可促进巨噬细胞NO释放 ,但对LPS刺激NO的释放影响不大 ;GLPP(10 0mg·kg-1)灌胃给药 5d或体外给药 (10mg·L-1)均可使巨噬细胞iNOS含量增加。结论　GLPP可增加小鼠腹腔巨噬细胞NO产生 ,其机制可能与其促进巨噬细胞iNOS合成有关。     

Contrasting effects of an aminobisphosphonate,a potent inhibitor of bone resorption,on lipopolysaccharide-induced production of interleukin-1 and tumour necrosis factor α in mice

1. Aminobisphosphonates (aminoBPs), potent inhibitors of bone resorption, have been reported to induce inflammatory reactions such as fever and an increase in acute phase proteins in human patients, and to induce the histamine-forming enzyme, histidine decarboxylase, in mice. In the present study, we examined the effect of aminoBP, 4-amino-1-hydroxybutylidene-1,1-bisphosphonic acid (AHBuBP), on the production of the pro-inflammatory cytokines, IL-1 and TNFα, in mice.
2. Intraperitoneal injection of AHBuBP did not itself produce detectable levels of IL-1 (α and β) and TNFα in the serum. However, the elevation of serum IL-1 induced by lipopolysaccharide (LPS) was greatly augmented in mice injected with AHBuBP 3 days before the LPS injection, whereas the LPS-induced elevation of serum TNFα was almost completely abolished.
3. Spleen and bone marrow cells taken from mice injected with AHBuBP produced IL-1β in vitro spontaneously, and the production was augmented following the addition of LPS. Cells that accumulated in the peritoneal cavity in response to AHBuBP produced a particularly large amount of IL-1β. However, AHBuBP treatment of mice did not lead to an impairment of the in vitro production of TNFα by these three types of cells.
4. Liposomes encapsulating dichloromethylene bisphosphonate (a non-amino BP) selectively deplete phagocytic macrophages. When an intraperitoneal injection of these liposomes was given 2 days after an injection of AHBuBP, there was a marked decrease in the LPS-induced elevation of serum IL-1 (α and β) (LPS being injected 3 days after the injection of AHBuBP).
5. These results indicate that AHBuBP has contrasting effects on the in vivo LPS-induced production of IL-1 and TNFα in mice, enhancing the production of IL-1 by phagocytic macrophages and suppressing the production of TNFα, although underling mechanisms remain to be clarified.

     

Modulation of rat macrophage function by the Mangifera indica L. extracts Vimang and mangiferin

Vimang is an aqueous extract of Mangiferia indica L., traditionally used in Cuba as an anti-inflammatory, analgesic and antioxidant. In the present study, we investigated the effects of Vimang and of mangiferin (a C-glucosylxanthone present in the extract) on rat macrophage functions including phagocytic activity and the respiratory burst. Both Vimang and mangiferin showed inhibitory effects on macrophage activity: (a) intraperitoneal doses of only 50-250 mg/kg markedly reduced the number of macrophages in peritoneal exudate following intraperitoneal injection of thioglycollate 5 days previously (though there was no significant effect on the proportion of macrophages in the peritoneal-exudate cell population); (b) in vitro concentrations of 0.1-100 microg/ml reduced the phagocytosis of yeasts cells by resident peritoneal and thioglycollate-elicited macrophages; (c) in vitro concentrations of 1-50 microg/ml reduced nitric oxide (NO) production by thioglycollate-elicited macrophages stimulated in vitro with lipopolysaccharide (LPS) and IFNgamma; and (d) in vitro concentrations of 1-50 microg/ml reduced the extracellular production of reactive oxygen species (ROS) by resident and thioglycollate-elicited macrophages stimulated in vitro with phorbol myristate acetate (PMA). These results suggest that components of Vimang, including the polyphenol mangiferin, have depressor effects on the phagocytic and ROS production activities of rat macrophages and, thus, that they may be of value in the treatment of diseases of immunopathological origin characterized by the hyperactivation of phagocytic cells such as certain autoimmune disorders.     

Manganese chloride enhances natural cell-mediated immune effector cell function: effects on macrophages

A single intramuscular injection of MnC12 in mice caused an increase in macrophage functional activity. Spleen cell antibody-dependent cell-mediated cytotoxicity against both chicken erythrocytes and P815 tumor cell targets was enhanced 24 h following a single injection of MnC12. Enhanced antibody-dependent cell-mediated cytotoxicity activity following MnC12 treatment was not associated with a change in spleen cellularities compared with saline-injected mice. Resident peritoneal macrophages from mice injected intramuscularly with MnC12 displayed enhanced phagocytic activity for chicken erythrocytes in the presence or absence of opsonizing antibody. Enhanced cytolytic activity against P815 mastocytoma target cells and enhanced cytostatic activity against MBL-2 lymphoma target cells was also observed for nonelicited resident peritoneal macrophages from mice injected intramuscularly with MnC12. There were no differences in the cellularity or relative number of adherent cells obtained from the peritoneal cavity of saline or MnC12-injected mice. These enhanced macrophage functions were associated with the induction of increased interferon levels in mice injected with MnC12.     

Biological and Inflammatory Effects of Antigen 5 from Polybia paulista (Hymenoptera,Vespidae) Venom in Mouse Intraperitoneal Macrophages

The social wasp Polybia paulista (Hymenoptera, Vespidae) is highly aggressive, being responsible for many medical occurrences. One of the most allergenic components of this venom is Antigen 5 (Poly p 5). The possible modulation of the in vitro immune response induced by antigen 5 from P. paulista venom, expressed recombinantly (rPoly p 5), on BALB/c mice peritoneal macrophages, activated or not with LPS, was assessed. Here, we analyzed cell viability changes, expression of the phosphorylated form of p65 NF-κB subunit, nitric oxide (NO), proinflammatory cytokines production, and co-stimulatory molecules (CD80, CD86). The results suggest that rPoly p 5 does not affect NO production nor the expression of co-stimulatory molecules in mouse peritoneal macrophages. On the other hand, rPoly p 5 induced an increase in IL-1β production in non-activated macrophages and a reduction in the production of TNF-α and MCP-1 cytokines in activated macrophages. rPoly p 5 decreased the in vitro production of the phosphorylated p65 NF-κB subunit in non-activated macrophages. These findings suggest an essential role of this allergen in the polarization of functional M2 macrophage phenotypes, when analyzed in previously activated macrophages. Further investigations, mainly in in vivo studies, should be conducted to elucidate Polybia paulista Ag5 biological role in the macrophage functional profile modulation.     

Saquinavir‐NO Inhibits IL‐6 Production in Macrophages

Covalent attachment of the nitric oxide (NO) moiety to the HIV protease inhibitor Saquinavir (Saq) produced a new chemical entity, named Saquinavir‐NO, (Saq‐NO) with reduced toxicity and potent immunoregulatory influence on T lymphocytes. In this study, we have compared head‐to‐head the effects of Saq‐NO and Saq on mouse and rat peritoneal macrophage cytokine secretion and NO production upon in vitro, ex vivo and in vivo conditions. The results demonstrate that Saq‐NO, but not Saq, potently decreased interleukin (IL)‐10, IL‐6 and nitrite accumulation and increased the levels of IL‐1β and tumour necrosis factor (TNF) in supernatants of mouse and rat macrophage cultures in vitro. Treatment of mice with Saq‐NO, but not Saq, inhibited ex vivo secretion of IL‐6 from macrophages. Consistent with these findings, Saq‐NO also reduced blood levels of IL‐6 in lipopolysaccharide‐treated mice. The observed inhibitory influence of Saq‐NO on IL‐6 generation in macrophages may be involved in the observed antitumour and immunomodulatory effects of the drug.     

Salvianic acid A inhibits induction of inflammatory mediators by blocking Nuclear Factor-kB activation in macrophages

Objective To investigate the anti-inflammation effect and possible mechanism of Salvianic acid A (SAA) in mouse peritoneal macrophages. Methods Peritoneal macrophages were obtained from BALB/c mice. LPS induced nitric oxide (NO), tumor necrosis factor-alpha (TNF-α) and interleukin-6 (IL-6) in supernatant, protein expression of inducible nitric oxide synthase (iNOS), matrix metalloproteinase-9 (MMP-9) and activation of nuclear factor-kappa B (NF-kB) in the extract were measured. Results SAA strongly inhibited the excessive production of NO, TNF-α and IL-6 in LPS-induced peritoneal macrophages in a concentration-dependent manner and blocked the expression of iNOS and MMP-9. Treatment with LPS alone increased the translocation of NF-kB (1065) from cytosol to the nucleus, but the SAA inhibited the translocation of NF-kB (p65). Conclusions The results showed that SAA had strong anti-inflammatory effects in LPS-stimulated peritoneal macrophages. The important mechanism is due to its inhibition of NF-kB activation.     

Effects of mancozeb on the activities of murine peritoneal macrophagesin vitro andex vivo

Mancozeb (MCZ) is known to have detrimental effects on the reproductive system, but the toxicity of MCZ on immune responses has not been systematically investigated. We investigated the effects of MCZ exposure on the activities of murine peritoneal macrophages through evaluation of MCZ-induced alteration of nitric oxide (NO) production and tumor necrosis factor-alpha (TNF-alpha) synthesis. Macrophages were examined ex vivo from mice orally treated with various doses of MCZ for 5 consecutive days per week for 4 weeks (subacute exposure, 250, 1000, 1500 mg/kg/day) followed by culture for 2 (TNF-alpha) or 3 days (NO) in the presence of LPS plus IFN-gamma. Macrophages from naive mice were also cultured with various concentrations of MCZ (0.05, 0.25, 0.5, 1 and 2 microg/mIL in the presence of LPS plus IFN-gamma for 2 (TNF-alpha) or 3 days (NO) in vitro. NO production was decreased with the in vitro exposure to all concentrations of MCZ. However, the amount of NO production by peritoneal macrophages from MCZ-subacutely exposed mice was increased in comparision with that of control group. In vitro, MCZ suppressed TNF-alpha secretion with significant reduction at 2 microg/mL MCZ. Conversely, TNF-alpha release was enhanced ex vivo. This study provides the substantial evidence on MCZ-induced alternation in macrophage activity. In order to clearly understand the contrasting effect of MCZ on peritoneal macrophage activity, it is necessary to further investigate the influence of major metabolite of MCZ (ETU) exposure on the NO production and TNF-alpha synthesis.     

Macrophage activation by polysaccharide biological response modifier isolated from Aloe vera L. var. chinensis (Haw.) Berg

A mannose-rich polysaccharide biological response modifier (BRM), derived from Aloe vera L. var. chinensis (Haw.) Berg., was demonstrated to be a potent murine B- and T-cell stimulator in our previous study. We here report the stimulatory activity of PAC-I on murine peritoneal macrophage. The polysaccharide when injected into mice enhanced the migration of macrophages to the peritoneal cavity. Peritoneal macrophage when treated by PAC-I in vitro had increased expression of MHC-II and FcgammaR, and enhanced endocytosis, phagocytosis, nitric oxide production, TNF-alpha secretion and tumor cell cytotoxicity. The administration of PAC-I into allogeneic ICR mice stimulated systemic TNF-alpha production in a dose-dependent manner and prolonged the survival of tumor-bearing mice. PAC-I is thus a potent stimulator of murine macrophage and the in vitro observed tumoricidal properties of activated macrophage might account for the in vivo antitumor properties of PAC-I. Our research findings may have therapeutic implications in tumor immunotherapy.     

Stimulation of various functions in murine peritoneal macrophages by high mannuronic acid-containing alginate (HMA) exposure in vivo

High mannuronic acid-containing alginate (HMA) was tested to affect murine peritoneal macrophages. In the present study, we measured various functions of murine peritoneal macrophages that were isolated 20 h after intraperitoneal injection with HMA (25 and 100 mg/kg). HMA increased the number of peritoneal macrophages and phagocytosis. Macrophages from HMA-treated mice significantly inhibited growth of tumor cells compared to macrophages from control mice. In addition, supernatants from macrophages of HMA-treated mice contained nitric oxide (NO), hydrogen peroxide (H2O2) and TNF-alpha. The increased production of these cytotoxic molecules induced by HMA is consistent with tumoricidal activity of activated macrophages. Furthermore, HMA-induced tumoricidal activity was partially abrogated by anti-TNF-alpha, inhibitors of NO and the scavenger of reactive oxygen. Thus, the tumoricidal activity induced by HMA appeared to be mediated by the production of TNF-alpha, NO and H2O2. Taken together, these results suggest that HMA has the immunostimulating effect on macrophages after in vivo exposure of it.     

The effects of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) on tumor necrosis factor (TNF) production by peritoneal cells

Recent studies in mice have demonstrated that TNF plays a critical role in mediating the TCDD-induced enhanced inflammatory response to intraperitoneal (i.p.) sheep red blood cells. The current studies were designed to evaluate the effects of TCDD on TNF production by ex-vivo peritoneal cells and a peritoneal macrophage cell line (IC-21) stimulated with LPS. In support of the hypothesis that TCDD can act directly on the peritoneal macrophage to increase TNF production, following pretreatment with TCDD, both ex-vivo peritoneal cells and IC-21 cells produced increased levels of bioactive TNF when stimulated with LPS. Flow cytometric analyses of IC-21 cells indicate that TCDD exposure increases intracellular production and secretion of TNF but does not alter levels of membrane associated TNF.     

Ternatin, an anti-inflammatory flavonoid, inhibits thioglycolate-elicited rat peritoneal neutrophil accumulation and LPS-activated nitric oxide production in murine macrophages

Ternatin, an anti-inflammatory flavonoid from Egletes viscosa Less., was examined for its possible influence on thioglycolate-elicited neutrophil influx into the rat peritoneal cavity in vivo and nitric oxide production in lipopolysaccharide (LPS)-activated mouse peritoneal macrophages ex vivo. The neutrophil influx induced by thioglycolate was found to be significantly lower in ternatin (25 and 50 mg/kg, s. c.) pre-treated rats with a similar magnitude of inhibition produced by dexamethasone (1 mg/kg, s. c.), a known anti-inflammatory agent. Also, peritoneal macrophages from ternatin (25 mg/kg)-treated mice that were exposed to LPS demonstrated significantly less production of nitric oxide (NO). These results suggest that ternatin exerts its anti-inflammatory action, at least in part, through inhibition of neutrophil migration and modulation of macrophage function.     

Hematein inhibits atherosclerosis by inhibition of reactive oxygen generation and NF-kappaB-dependent inflammatory mediators in hyperlipidemic mice

Hematein, a natural compound, is a known anti-inflammatory and antiatherogenic agent in the rabbit model. The authors investigated the effects of this compound on atherogenesis and possible mechanisms of the actions in the hyperlipidemic mice. Low-density lipoprotein receptor-deficient (Ldlr-/-) mice fed a high-cholesterol diet alone for 8 weeks developed the fatty streak lesion in the aortic sinus, whereas this lesion was significantly reduced by hematein treatment without a change in plasma lipid levels compared with control mice. Hematein treatment reduced plasma levels of lipid peroxide and superoxide generation in LPS-stimulated peritoneal macrophage. Hematein treatment inhibited NF-kappaB-DNA binding activity in peritoneal macrophages from Ldlr-/- mice and the activation of NF-kappaB in RAW264.7 macrophages. This compound suppressed plasma nitrite/nitrate levels in Ldlr-/- mice and NO production and iNOS expression in LPS+IFNgamma-stimulated peritoneal macrophages. Hematein treatment also suppressed the activity of iNOS promoters in RAW264.7 macrophages, and reduced the plasma levels of TNF-alpha and IL-1beta and the production of these cytokines in LPS+IFNgamma-stimulated peritoneal macrophages. These results suggest that hematein inhibits atherosclerotic lesion formation, possibly by reducing proinflammatory mediators through a decrease in reactive oxygen species generation and NF-kappaB activation.     

Comparison of effects of CH50 on macrophage activation and its anti-tumor activity with those of lipopolysaccharides

AIM: To investigate the characterization of pharmacological action & of CH50, a recombinant polypeptide of human fibronectin, by comparing the effects of CH50 on macrophage activation and its anti-tumor activity with those of lipopolysaccharides (LPS). METHODS: The production of nitric oxide (NO) as an index macrophage activation was determined by colorimetric assay. The interferon-gamma (IFN-gamma) transfection was performed with coprecipitation of calcium phosphate and DNA. The melanoma B16 cells were inoculated into abdominal cavity of mice and the number of tumor nodes was recorded. RESULTS: At lower concentrations or when given alone in vitro, CH50 produced ten times less NO than LPS (P < 0.01). But at concentrations higher than 1 mg.L-1, CH50 activated the IFN-gamma-primed macrophages to produce NO to the same extent as LPS (P > 0.05). There was no synergism between CH50 and LPS. Both CH50 and LPS alone could reduce the number of tumor nodes in abdominal cavity of mice but CH50 had a stronger inhibitory effect on the growth of tumor in vivo as compared to LPS (P < 0.01). CH50/IFN-gamma had also a better inhibitory effect on tumor growth in vivo than LPS/IFN-gamma did. CONCLUSION: In the presence of IFN-gamma, the ability of CH50 to activate macrophages is the same as that of LPS. But CH50 has better antitumorogenic effects in vivo against mouse melanoma as compared to LPS.