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1.
Mycoplasma arthritidis causes arthritis in rodents that resembles human rheumatoid arthritis. It produces a superantigen (MAM) that stimulates production of cytokines by making a bridge between lymphocyte T-cell receptor with the appropriate Vbeta chain, and H-2 1-Ealpha MHC class II molecules. Here we studied MAM-induced nitric oxide (NO) production in mouse peritoneal macrophages and found that it was: (1) time and concentration dependent, (2) possibly derived from inducible NOS synthase since it was reduced significantly by amino guanidine pretreatment, (3) restricted to H-2(K) (C3H/HePas and C3H/HeJ) and H-2(d) strains (BALB/c), (4) independent of TLR4 signaling since the coisogenic strains C3H/HePas and C3H/HeJ (TLR4 deficient) produced similar levels of NO following MAM stimulation, (5) potentiated by lipopolysaccharide, and (6) dependent on the presence of nonadherent peritoneal cells. Neutralization of interferon-gamma (IFNgamma in the peritoneal cell cultures with monoclonal antibodies abolished MAM-induced NO production. Addition of rIFNgamma to the adherent cells substituted the nonadherent cells for MAM-induced NO production. A macrophage cell line, J774A.1 (H-2(d)), also produced NO upon MAM stimulation but only when BALB/c spleen lymphocytes were added. Thus, in murine macrophages, MAM induces NO production that is dependent on signaling through MHC class II molecules and IFNgamma but independent of TLR4 expression.  相似文献   

2.
3.
The cultured cells prepared from the spleens and peritoneal exudate cells of the C3H/HeJ strain of mice produce very little or no interferon (IFN) by stimulation of bacterial lipopolysaccharide (LPS). However, the cells taken from LPS-non-responder C3H/HeJ mice which had been infected with Mycobacterium bovis bacillus Calmette-Guérin (BCG) prior to the experiment were capable of producing IFN in culture in the presence of LPS. The peritoneal exudate cells of BCG-primed C3H/HeJ mice were separated into adherent cell and nonadherent cell populations by their adhesiveness to plastic culture dishes. IFN production required the presence of both these cell populations in the same culture, and the IFN activities produced were mainly IFN-gamma. The cultures with nonadherent cells and fixed adherent cells still produced IFN, but the cell cultures reconstituted with the BCG-primed cell population and unprimed cell population produce little if any IFN-gamma. Moreover, when both of the populations were cultured in Marbrook culture vessels separated by a membrane filter, the cultures produced very little or no IFN-gamma. These results indicate that there is a mechanism of IFN-gamma induction by LPS which requires the direct contact between adherent cells and nonadherent cells without the participation of any soluble factor(s) from the adherent cells. The producer cells were mainly in the nonadherent cell population. Previous treatment of nonadherent cells with anti-Thy-1.2 antibody, anti-Lyt-1.1 antibody, anti-L3T4 antibody, or anti-asialo-GM1 antibody and complement diminished the ability of the cells for LPS-induced IFN production with the help of adherent cells. Therefore, it is concluded that both T cells (presumably L3T4+T cells) and asialo-GM1+ natural killer cells in the BCG-primed C3H/HeJ cell cultures produced IFN-gamma in the presence of LPS, and the production was supported by the function of macrophages.  相似文献   

4.
ABSTRACT Nitrite production by rat alveolar macrophages was studied to determine the role of L-arginine oxidation in the interaction between these cells and Pneumocystis carinii. Alveolar macrophages from rats obtained from two different breeders were used: rats from Janvier breeder had latent P. carinii infection, while those from Charles River breeder were bred in a germ-free environment. Pneumocystis carinii increased in vitro nitrite generation by unstimulated alveolar macrophages from Janvier rats only, and this was blocked by NG-monomethyl-L-arginine. Incubation of cells from Janvier and Charles River rats with lipopolysaccharide and/or interferon-gamma increased nitrite production to a similar extent. Pneumocystis carinii partially decreased nitrite release by activated alveolar macrophages, and this was still inhibited by NG-monomethyl-L-arginine. In the presence of P. carinii, superoxide dismutase used as a superoxide anion scavenger had no effect on nitrite production by activated cells. These results show that prior exposure to P. carinii leads to nitric oxide production by rat alveolar macrophages. Although the magnitude of this production seems to be moderate, it is of biological significance since cells of P. curinii-naive rats do not generate nitrite whereas those of latently infected rats do.  相似文献   

5.
Lymphocyte-activating factor (LAF), a mitogen for thymocytes and T lymphocytes, is released into the culture medium by human mononuclear cells and mouse peritoneal exudate cells following treatment with various macrophage stimulants. Experiments were performed to determine if recently described mouse macrophage cell lines released LAF in response to stimulation with bacterial lipopolysaccharide. Four continuous cell lines (P388-D1, J774, WEHI 3, and PU5-1.8) were found to release LAF in serum-free medium following endotoxin stimulation. The results of partial purification indicated that LAF obtained from cell lines had a higher molecular weight and lower isoelectric point than LAF from human mononuclear cells.  相似文献   

6.
Lacto-N-fucopentaose III (LNFPIII) is found in human milk and on the Th2 driving helminth parasite Schistosoma mansoni. This pentasaccharide drives Th2-type responses in vivo and in vitro when conjugated to a carrier. In an attempt to further understand early events in Th1 to Th2 switching, we examined phenotypic and functional changes in peritoneal cell populations in BALB/c and SCID mice following LNFPIII-dextran injection. We found that i.p. injection with LNFPIII-dextran resulted in rapid (<20 h) expansion of the Gr1(+) subpopulation of F4/80(+)/CD11b(+) peritoneal cells, comprising up to 75% of F4/80(+)/CD11b(+) peritoneal cells compared with 18% in uninjected or dextran-injected mice. Functionally, these cells suppressed anti-CD3- and anti-CD28-induced proliferation of naive CD4(+) T cells. LNFPIII-dextran also expanded functional Gr1(+) suppressor macrophages in SCID mice, demonstrating that expansion and function of suppressor cells did not require T cells. Suppression in both BALB/c and SCID mice was NO and IFN-gamma dependent, as addition of inhibitors of inducible NO synthase (N(G)-monomethyl-L-arginine), as well as anti-IFN-gamma Abs, restored the ability of CD4(+) T cells to proliferate in vitro. Depletion of the F4/80(+) subset of Gr1(+) cells eliminated the suppressive activity of peritoneal exudate cells showing that these cells were macrophages. Thus, LNFPIII-dextran rapidly expands the Gr1(+) suppressor macrophage population in the peritoneal cavities of otherwise naive mice. These Gr1(+) cells suppress proliferation of naive CD4(+) T cells in an NO-dependent mechanism, and may play a regulatory role in the switching of Th1- to Th2-type responses.  相似文献   

7.
Luminol-enhanced chemiluminescence (CL) was used to examine the response of various leukocyte populations following stimulation with a crude extract of Phaseolus vulgaris, namely phytohaemagglutinin (PHA-C). Populations stimulated included a human peripheral mixed leukocyte preparation (MLP), and purified preparations of lymphocytes, monocytes and polymorphonuclear leukocytes (PMNL). Mouse peritoneal exudate cells and the lymphocytic cells lines Molt #4 and Daudi were also stimulated. Following stimulation, a characteristic three-peaked chemiluminescent response was obtained from the MLP population. Little or no response was obtained from the purified lymphocytes. Monocytes produced a sharp peak corresponding to the second peak of the MLP response and PMNL produced a broad peak corresponding to the third peak of the MLP response. Mouse peritoneal exudate cells containing lymphocytes and monocytes/macrophages showed a two-peaked stimulation which corresponded to the first two peaks of the MLP response. Molt #4 and Daudi showed no chemiluminescence if stimulated individually, but if added to a MLP substantial enhancement of the first and second peaks was observed. These results indicate some form of lymphocyte/monocyte interaction leading to enhanced CL following PHA-C stimulation.  相似文献   

8.
Production of prostaglandin D2 by murine macrophage cell lines   总被引:2,自引:0,他引:2  
Several tumor-derived murine macrophage cell lines were evaluated in vitro as cloned prototypes of tissue macrophages for their ability to metabolize arachidonic acid. Unexpectedly, two cell lines, J774A.1 and WR19M.1, rapidly converted exogenous 14C-arachidonic acid (AA) to a single major prostaglandin metabolite. The compound, PGD2, was positively identified by TLC, HPLC, and GC-MS. The enzymatic formation of the PGD2 was shown by inhibition of its formation by indomethacin and reduced formation of 14C-PGD2 from 14C-PGH2 in boiled cells. When J774A.1 cells were prelabeled with 3H-AA, cultured for 24 hours, and stimulated with lipopolysaccharide (LPS), PGD2 was again the predominant product. No other tumor derived cell lines, including several other murine macrophage lines, produced significant amounts of PGD2. Elicited and activated murine peritoneal macrophages produced only small amounts of PGD2, but resident peritoneal macrophages produced modest amounts of PGD2. Exaggerated formation of PGD2 by J774A.1 and WR19M.1 cells may be a consequence of neoplastic transformation or the clonal expansion of a minor subpopulation of normal tissue macrophages.  相似文献   

9.
Injection of adult mouse peritoneal exudate cells into newborn mice results in a premature decrease of splenic suppressor cell activity in the neonates. The effect becomes apparent 4–5 days after ip injection of 10–15 × 106 thioglycollate-induced peritoneal exudate cells into mice on the day of birth. The macrophage in the peritoneal exudate is the responsible cell type. The effect is not H-2 restricted or strain limited. Heat-killed peritoneal exudate cells or peritoneal cells from unstimulated donors can also decrease neonatal suppressor cell activity prematurely. Adult spleen cells, injected into neonatal mice, do not affect suppressor cell activity. The data are discussed in light of the hypothesis that macrophages control suppressor activity in neonatal mice and that an increase in the number and/or function of macrophages shortly after birth results in a decrease in the number and/or function of suppressor cells, allowing for immunological competence to emerge.  相似文献   

10.
When mouse spleen (Ig) cells undergo maximal mitogenic stimulation by optimal concentrations of concanavalin A (conA), the Ig cells form caps of conA very slowly, with 50% of maximum cap formation occurring after about 10 h and maximal capping after about 24 h. Anti-conA antibody added after optimal conA accelerates the rate of cap formation and effectively blocks mitogenic stimulation (< 10%) by optimal conA concentrations when the rate of capping is increased more than about 2-fold. The effect of anti-conA antibody in accelerating cap formation by optimal conA is antagonized by cytochalasin D (CD), which substantially restores the mitogenic action of optimal conA. Thus there is an inverse relationship between rate of cap formation and extent of mitogenic stimulation. Further experiments showed that if anti-conA antibody, α-methyl mannoside or EGTA were added at increasing intervals after the addition of conA, these inhibitors block the stimulation of the cells with very similar time courses. Addition of appropriate concentrations of an inhibitor at the same time as optimal conA blocks mitogenic stimulation completely, but has negligible effects after 24 h. The extent of stimulation which occurs after the addition of inhibitor at intermediate times closely follows the extent of cap formation at the same time. The simplest interpretation of these results is that mitogenic action by optimal conA can be blocked by (i) accelerated capping of uncapped cells; or (ii) by the removal of either conA or calcium before, but not after, cap formation has occurred. These results suggest that the rate of cap formation by conA, and the presence of external calcium (>10−4 M) in the medium for some unspecified period before cap formation occurs are both significant factors in generating the primary mitogenic signals which commit the cells to DNA synthesis.  相似文献   

11.
A cyanine photosensitizing dye, platonin, is a potent macrophage-activating agent. Four days after the administration to mice of small amounts of platonin (20–40 ng/mouse), peritoneal macrophages exhibited greatly enhanced Fc-receptor-mediated phagocytic and superoxide-generating capacities. Much higher doses (more than 3000 ng/mouse) did not have this effect. Photodynamic experiments for macrophage activation were performed by exposing mouse peritoneal cells (mixture of macrophages and B and T lymphocytes) to white fluorenscent light (3 J m–2s–1) in media containing various low concentrations of platonin. A short exposure to white fluorescent light (5 s, 15 J m–2) of peritoneal cells in a medium containing 3 ng platonin/ml produced a maximal level of phagocytic capacity of macrophages. Although platonin absorbs light poorly at wavelengths longer than 630 nm, the region of the spectrum in which the tissues are transparent allows reasonable penetration of light. Thus, we designed experiments in which peritoneal cells were exposed to a red fluorescent light (0.5 J m–2s–1). In a medium containing 10 ng platonin/ml with 15 J m–2 red light, a markedly enhanced ingestion activity of macrophages was observed. Photodynamic treatment of peritoneal macrophages alone did not activate macrophages. Thus, participation of nonadherent cells is required for photodynamic activation of macrophages, implying that a macrophage-activating factor is generated within the nonadherent cells and transmitted to macrophages.  相似文献   

12.
Fucose-binding protein (FBP), the fucolectin from Lotus tetragonolobus, was compared with migration inhibitory factor (MIF) for its ability to stimulate [14C]glucosamine incorporation into trichloroacetic acid (TCA)-insoluble material of guinea pig peritoneal exudate cells (PEC) using a microtiter assay. Both MIF and FBP inhibit macrophage migration and were shown to stimulate glucosamine incorporation in a similar dose response fashion over time. Both unpurified PEC and PEC depleted of nonadherent cells displayed significant levels of glucosamine incorporation when stimulated by MIF or FBP. Tunicamycin and 2-deoxy-d-glucose, known inhibitors of glycosylation, inhibited glucosamine incorporation by control and MIF- or FBP-stimulated PEC. These results confirm the similarities between MIF and FBP in their biological activity for macrophages using a second in vitro correlate of cell-mediated immunity and suggest involvement of enhanced glycoprotein or glycolipid biosynthesis by FBP and lymphokine-activated macrophages.  相似文献   

13.
Previous reports have suggested that repeated alloantigenic challenge increases humoral responses to alloantigens, but may cause decreasing cellular responses. We stimulated BALB/c (H-2d) mice with intraperitoneal EL-4 tumor (H-2b) and serially assessed cytotoxic responses in spleen and peritoneal lymphocytes using 51Cr-labeled EL-4 target cells. We observed that cytotoxicity generated in the spleen of nonimmune BALB/c mice was much greater than that in immunized mice; similar peak responses were generated in peritoneal lymphocytes in normal and immunized hosts. Complement-mediated cytotoxicity was not necessary for diminished splenic responses in hyperimmune hosts, for the same phenomenon was seen in the Hzl anti-BL/6 system which is free of humoral responses. Irradiation (2000 rad) of the EL-4 tumor challenge prevented tumor cell proliferation and markedly reduced splenic responses in nonimmune mice. We suggest that cytotoxic cells suppressed further generation of cyto-toxicity; by effecting an early elimination of tumor inoculum, tumor proliferation was abrogated and dose of cellular antigen was, consequently, markedly reduced.  相似文献   

14.
Summary Reticulum cell sarcoma (RCS) cells of SJL/J (H-2s) mice have been shown to express antigens that are cross-reactive with allogeneic cells of the H-2d and H-2b haplotypes by cell-mediated cytotoxicity, antibody-mediated cytotoxicity, immunofluorescence, and quantitative absorption assays. These alien antigens have been detected on both spontaneous and in vivo- and in vitro-passaged RCS cells to varying degrees.The in vitro cell lines were able to stimulate a syngeneic cytotoxic T cell response detected in a 4-h 51Cr release assay. The cytotoxic cells reacted with in vitro RCS tumor targets but not with in vivo or spontaneous RCS tumors. Furthermore, the cytotoxic cells lysed H-2d and to a lesser extent H-2b target cells, but not H-2k, H-2p, or H-2r cells. The cross-reactivity was also observed with SJL/J anti-BALB/c cytotoxic cells, which can lyse in vitro RCS targets effectively. The in vivo tumors were not stimulatory in cytotoxic responses and did not serve as targets.H-2d specificities were also detected in cultured RCS tumor cells by cytotoxic antibody. Both allogeneic SJL/J anti-BALB/c, C57B1/6 anti-BALB/c sera reacted with RCS tumor cells and not normal SJL/J cells. Furthermore, monospecific Dd sera were also cytotoxic against RCS lines. The cytotoxic activity could be absorbed by BALB/c cells and RCS cells but not with normal SJL/J cells. The H-2d specificities were also detected on the in vivo lines by indirect immunofluorescence. The majority (60%) of spontaneously arising tumors expressed either H-2d or H-2b allospecificities in the immunofluorescence assays. Although these antigens may not be inappropriate for the SJL/J strain, their differential expression on tumor cells may be significant in the etiology of the tumor.  相似文献   

15.
《Life sciences》1995,57(26):PL401-PL406
Pre-incubation of J774 murine macrophages with 5,6-dihydroxyindole-2-carboxylic acid (DHICA), a diffusible intermediate in the biosynthesis of eumelanins, leads to a marked increase in the levels of nitric oxide (NO) produced by lipopolysaccharide (LPS)-induced NO-synthase (iNOS). The effect varies with DHICA concentratior being maximum at a concentration of 1 × 10−6M, and is suppressed by the NOS inhibitor NG-monomethyl-L-arginine (L-NMMA). No stimulation is observed when macrophages are exposed to DHICA after activation with LPS, indicating that the indole does not affect the catalytic activity of iNOS. These results point to a hitherto unrecognized role of DHICA as a chemical messenger mediating interaction between active melanocytes and macrophages in epidermal inflammatory and immune responses.  相似文献   

16.
Several tumor-derived murine macrophage cell lines were evaluated as cloned prototypes of tissue macrophages for their ability to metabolize arachidonic acid. Unexpectedly, two cell lines, J774A.1 and WR19M.1, rapidly converted exogenous 14C-arachidonic acid (AA) to a single major prostaglandin metabolite. The compound, PGD2, was positively identified by TLC, HPLC, and GC-MS. The enzymatic formation of the PGD2 was shown by inhibition of its formation by indomethacin and reduced formation of 14C-PGD2 and 14C-PGD2 in boiled cells. When J774A.1 cells were prelabeled with 3H-AA, cultured for 24 hours, and stimulated with lipopolysaccharide (LPS), PGD2 was again the predominant product. No other tumor derived cell lines, including several other murine macrophage lines, produced significant amounts of PGD2. Elicited and activated murine peritoneal macrophages produced only small amounts of PGD2, but resident peritoneal macrophages produced modest amounts of PGD2. Exaggerated formation of PGD2 by J774A.1 and WR19M.1 cells may be a consequence of neoplastic transformation or the clonal expansion of a minor subpopulation of normal tissue macrophages.  相似文献   

17.
Net flux of cholesterol represents the difference between efflux and influx and can result in net cell-cholesterol accumulation, net cell-cholesterol depletion, or no change in cellular cholesterol content. We measured radiolabeled cell-cholesterol efflux and cell-cholesterol mass using cholesterol-normal and -enriched J774 and elicited mouse peritoneal macrophage cells. Net cell-cholesterol effluxes were observed when cholesterol-enriched J774 cells were incubated with 3.5% apolipoprotein (apo) B depleted human serum, HDL3, and apo A-I. Net cell-cholesterol influxes were observed when cholesterol-normal J774 cells were incubated with the same acceptors except apo A-I. When incubated with 2.5% individual sera, cholesterol mass efflux in free cholesterol (FC)-enriched J774 cells correlated with the HDL-cholesterol (HDL-C) concentrations (r2 = 0.4; P=0.003), whereas cholesterol mass influx in cholesterol-normal J774 cells correlated with the LDL cholesterol (LDL-C) concentrations (r2 = 0.6; P<0.0001) of the individual sera. A positive correlation was observed between measurements of [3H]cholesterol efflux and reductions in cholesterol mass (r2 = 0.4; P=0.001) in FC-enriched J774 cells. In conclusion, isotopic efflux measurements from cholesterol-normal or cholesterol-enriched cells provide an accurate measurement of relative ability of an acceptor to remove labeled cholesterol under a specific set of experimental conditions, i.e., efflux potential. Moreover, isotopic efflux measurements can reflect changes in cellular cholesterol mass if the donor cells are enriched with cholesterol.  相似文献   

18.
Alloantibodies specific for non-H-2 histocompatibility antigens of the mouse have been produced. Immunization (BALB/cJ×DBA/2J)F1 anti-B10.D2/n was conducted, followed by hemagglutination, immunofluorescence, and mixed hemabsorption tests on absorbed and unabsorbed sera. The results indicate that antibodies specific for H-3a and H-8a antigens are present. In addition, H-8a antigenic determinants were detected on erythrocyte membrane surfaces, as well as on cells of other body tissues.  相似文献   

19.
The plant lectins, concanavalin A (conA), wheat germ agglutinin (WGA), and phytohemagglutinin (PHA) stimulate steroidogenesis in cultured adrenal tumor cells. ConA maximally stimulated steroidogenesis at 100 μg/ml following an approximate 4 h lag phase. ConA stimulation was completely inhibited by α-methyl-d-mannopyranoside and the WGA effect was prevented by N-acetyl-d-glucosamine. It was also found that conA alone did not cause a measurable increase in either intra- or extracellular cyclic adenosine 3′5′-monophosphate (cAMP) production. In addition, conA when added simultaneously with adrenocorticotropin (ACTH) doubled the intra- and extracellular cAMP production over controls treated with ACTH alone. This enhancement effect was dose dependent. When Y-1 cells were preincubated with conA and then treated with either ACTH or cholera enterotoxin (CT) there was a dose- and time-dependent inhibition of induced cAMP production. In the case of CT, the inhibitory effect occurred even with simultaneous addition of conA and CT. This effect was reversed by addition of both α-methyl-d-mannopyranoside and washing with Eagle's minimal essential medium (MEM) 1 h after CT had bound to its receptor. This reversal was not apparent for the inhibitory effect of conA on ACTH-induced cAMP production which occurred after 2 h of preincubation with conA. These results demonstrate that conA, as well as the other plant lectins, interact with specific membrane receptors to reversibly stimulate steroid production as well as enhancing or inhibiting ligand-induced cAMP production in cultured adrenal tumor cells.  相似文献   

20.
Peritoneal cells from highly susceptible BALB/c mice were infected with Leishmania major and cultured for various times in vitro. The culture supernatants contained significant levels of IL-1 which were consistently higher than those in the cell cultures stimulated with an optimal concentration of LPS. This finding extends to a macrophage cell line, P388D1, and peritoneal exudate cells stimulated with starch in vivo. However, the level of IL-1 produced was significantly reduced when the cells were preincubated with a lymphokine preparation (supernatant of Con A-stimulated rat spleen cells). The level of IL-1 produced seems to be directly correlated with the degree of parasitization of the macrophages. A similar and dose-dependent reduction in IL-1 production by infected macrophages could also be obtained when the cells were preincubated with IFN-gamma. This finding is in direct contrast to that of visceral leishmaniasis in which peritoneal macrophages from BALB/c mice infected with Leishmania donovani not only fail to produce IL-1 but also lose the capacity to produce IL-1. This apparent discrepancy is discussed in terms of a possible difference in the induction of cell-mediated immunity between the two leishmanial diseases.  相似文献   

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