Activation of murine macrophages by lipoprotein and lipooligosaccharide of Treponema denticola.
(9/20782)
We have recently demonstrated that the periodontopathogenic oral spirochete Treponema denticola possesses membrane-associated lipoproteins in addition to lipooligosaccharide (LOS). The aim of the present study was to test the potential of these oral spirochetal components to induce the production of inflammatory mediators by human macrophages, which in turn may stimulate tissue breakdown as observed in periodontal diseases. An enriched lipoprotein fraction (dLPP) from T. denticola ATCC 35404 obtained upon extraction of the treponemes with Triton X-114 was found to stimulate the production of nitric oxide (NO), tumor necrosis factor alpha (TNF-alpha), and interleukin-1 (IL-1) by mouse macrophages in a dose-dependent manner. Induction of NO by dLPP was at 25% of the levels obtained by Salmonella typhosa lipopolysaccharide (LPS) at similar concentrations, while IL-1 was produced at similar levels by both inducers. dLPP-mediated macrophage activation was unaffected by amounts of polymyxin B that neutralized the induction produced by S. typhosa LPS. dLPP also induced NO and TNF-alpha secretion from macrophages isolated from endotoxin-unresponsive C3H/HeJ mice to an extent similar to the stimulation produced in endotoxin-responsive mice. Purified T. denticola LOS also produced a concentration-dependent activation of NO and TNF-alpha in LPS-responsive and -nonresponsive mouse macrophages. However, macrophage activation by LOS was inhibited by polymyxin B. These results suggest that T. denticola lipoproteins and LOS may play a role in the inflammatory processes that characterize periodontal diseases. (+info)
Potent immunoregulatory effects of Salmonella typhi flagella on antigenic stimulation of human peripheral blood mononuclear cells.
(10/20782)
A key function of monocytes/macrophages (Mphi) is to present antigens to T cells. However, upon interaction with bacteria, Mphi lose their ability to effectively present soluble antigens. This functional loss was associated with alterations in the expression of adhesion molecules and CD14 and a reduction in the uptake of soluble antigen. Recently, we have demonstrated that Salmonella typhi flagella (STF) markedly decrease CD14 expression and are potent inducers of proinflammatory cytokine production by human peripheral blood mononuclear cells (hPBMC). In order to determine whether S. typhi and soluble STF also alter the ability of Mphi to activate T cells to proliferate to antigens and mitogens, hPBMC were cultured in the presence of tetanus toxoid (TT) or phytohemagglutinin (PHA) and either killed whole-cell S. typhi or purified STF protein. Both whole-cell S. typhi and STF suppressed proliferation to PHA and TT. This decreased proliferation was not a result of increased Mphi production of nitric oxide, prostaglandin E2, or oxygen radicals or the release of interleukin-1beta, tumor necrosis factor alpha, interleukin-6, or interleukin-10 following exposure to STF. However, the ability to take up soluble antigen, as determined by fluorescein isothiocyanate-labeled dextran uptake, was reduced in cells cultured with STF. Moreover, there was a dramatic reduction in the expression of CD54 on Mphi after exposure to STF. These results indicate that whole-cell S. typhi and STF have the ability to alter in vitro proliferation to soluble antigens and mitogens by affecting Mphi function. (+info)
Gamma interferon stimulates rat alveolar macrophages to kill Pneumocystis carinii by L-arginine- and tumor necrosis factor-dependent mechanisms.
(11/20782)
Pneumocystis carinii pneumonia remains a serious complication for immunocompromised patients. In the present study, P. carinii organisms interacted with gamma interferon (IFN-gamma)-stimulated alveolar macrophages (AMs) to activate the L-arginine-dependent cytocidal pathway involving reactive nitrogen intermediates (RNI) that were assayed as nitrite (NO2-). Unstimulated cultures of AMs produced negligible quantities of RNI. Addition of P. carinii organisms to IFN-gamma-primed AMs resulted in greatly enhanced production of RNI. NO2- levels increased from 0.8 +/- 0.4 to 11.1 +/- 3.8 microM as early as 6 h after P. carinii organisms were incubated with IFN-gamma-stimulated AMs and to 35.1 +/- 8.9 microM after a 24-h incubation, a near-maximum level. High levels of NO2- were produced by AMs primed with as little as 10 U of IFN-gamma per ml in the presence of P. carinii, and a 20-fold increase in IFN-gamma concentration resulted in only a further 65% increase in NO2- production. RNI-dependent killing of P. carinii was demonstrated by both a 51Cr release assay and a [35S]methionine pulse immunoprecipitation assay. Addition of either monoclonal tumor necrosis factor alpha (TNF-alpha) neutralizing antibody or 200 microM NG-monomethyl-L-arginine (L-NGMMA), a competitive inhibitor of the L-arginine-dependent pathway, significantly decreased NO2- production and reduced P. carinii killing. TNF-alpha alone had no effect on P. carinii viability. These results suggest that (i) the specific interaction of P. carinii organisms with IFN-gamma-primed AMs triggers the production of RNI, (ii) RNI are toxic to P. carinii, and (iii) TNF-alpha likely plays a central role in mediating P. carinii killing by IFN-gamma-stimulated AMs. (+info)
Effect of transforming growth factor beta on experimental Salmonella typhimurium infection in mice.
(12/20782)
We have investigated the effect of the in vivo administration of recombinant transforming growth factor beta (rTGF-beta) on the pathogenic mechanisms involved in Salmonella typhimurium experimental infection in mice. The protective response elicited by macrophages was induced by rTGF-beta1 by 2 days after experimental infection, as demonstrated by an increased NO production, while the humoral protective effect began with cytokine mRNA expression 2 days after the challenge and continued after 5 days with cytokine release and lymphocyte activation. We demonstrated that all mice who received rTGF-beta1 survived 7 days after infection. The number of bacteria recovered in the spleens and in the livers of rTGF-beta1-treated mice 2 and 5 days after infection was significantly smaller than that found in the same organs after phosphate-buffered saline (PBS) inoculation. Furthermore, 2 and 5 days after infection, splenic macrophages from rTGF-beta1-treated mice showed a greater NO production than did those from PBS-treated mice. The effect of rTGF-beta1 on S. typhimurium infection in mice was correlated with the expression of cell costimulatory CD28 molecules. Five days after S. typhimurium infection, the percentage of CD28(+)-expressing T cells in splenic lymphocytes from rTGF-beta1-treated mice increased with respect to that from control mice. Gamma interferon (IFN-gamma) mRNA was present in a greater amount in spleen cells from rTGF-beta1-treated mice after 2 days, although the intensity of the band decreased 5 days after the challenge. A similar pattern was obtained with the mRNAs for interleukin-1alpha (IL-1alpha), IL-6, TGF-beta, and inducible nitric oxide synthase, which showed greater expression in cells obtained from rTGF-beta1-treated and S. typhimurium-infected mice 2 days after challenge. The treatment with rTGF-beta1 induced an increase in IL-1alpha and IFN-gamma release in the supernatant of splenocyte cultures 5 days after the experimental infection with S. typhimurium. Moreover, we demonstrated that 5 days after infection, the IFN-gamma titer was significantly greater in the sera of rTGF-beta-treated mice than in those of PBS-treated mice. Also, hsp60 showed greater expression 2 days after the challenge in splenocytes from rTGF-beta1-treated mice. The role played by proinflammatory and immunoregulatory cytokines and by CD28 is discussed. (+info)
Inhibition of transforming growth factor beta production by nitric oxide-treated chondrocytes: implications for matrix synthesis.
(13/20782)
OBJECTIVE: Nitric oxide (NO) is generated copiously by articular chondrocytes activated by interleukin-1beta (IL-1beta). If NO production is blocked, much of the IL-1beta inhibition of proteoglycan synthesis is prevented. We tested the hypothesis that this inhibitory effect of NO on proteoglycan synthesis is secondary to changes in chondrocyte transforming growth factor beta (TGFbeta). METHODS: Monolayer, primary cultures of lapine articular chondrocytes and cartilage slices were studied. NO production was determined as nitrite accumulation in the medium. TGFbeta bioactivity in chondrocyte- and cartilage-conditioned medium (CM) was measured with the mink lung epithelial cell bioassay. Proteoglycan synthesis was measured as the incorporation of 35S-sodium sulfate into macromolecules separated from unincorporated label by gel filtration on PD-10 columns. RESULTS: IL-1beta increased active TGFbeta in chondrocyte CM by 12 hours; by 24 hours, significant increases in both active and latent TGFbeta were detectable. NG-monomethyl-L-arginine (L-NMA) potentiated the increase in total TGFbeta without affecting the early TGFbeta activation. IL-1beta stimulated a NO-independent, transient increase in TGFbeta3 at 24 hours; however, TGFbeta1 was not changed. When NO synthesis was inhibited with L-NMA, IL-1beta increased CM concentrations of TGFbeta1 from 24-72 hours of culture. L-arginine (10 mM) reversed the inhibitory effect of L-NMA on NO production and blocked the increases in TGFbeta1. Anti-TGFbeta1 antibody prevented the restoration of proteoglycan synthesis by chondrocytes exposed to IL-1beta + L-NMA, confirming that NO inhibition of TGFbeta1 in IL-1beta-treated chondrocytes effected, in part, the decreased proteoglycan synthesis. Furthermore, the increase in TGFbeta and proteoglycan synthesis seen with L-NMA was reversed by the NO donor S-nitroso-N-acetylpenicillamide. Similar results were seen with cartilage slices in organ culture. The autocrine increase in CM TGFbeta1 levels following prior exposure to TGFbeta1 was also blocked by NO. CONCLUSION: NO can modulate proteoglycan synthesis indirectly by decreasing the production of TGFbeta1 by chondrocytes exposed to IL-1beta. It prevents autocrine-stimulated increases in TGFbeta1, thus potentially diminishing the anabolic effects of this cytokine in chondrocytes. (+info)
Effect of tumor necrosis factor alpha on vascular resistance, nitric oxide production, and glucose and oxygen consumption in perfused tissue-isolated human melanoma xenografts.
(14/20782)
The effect of tumor necrosis factor alpha (TNF-alpha) on vascular resistance, nitric oxide production, and consumption of oxygen and glucose was examined in a perfused tissue-isolated tumor model in nude mice. One experimental group was perfused with heparinized Krebs-Henseleit buffer, a second one was perfused with TNF-alpha (500 microgram/kg) 5 h before perfusion. The vascular resistance increased significantly 5 h after TNF-alpha injection. The increase in vascular resistance did not seem to be mediated by a decrease in tumor nitric oxide production, as determined by perfusate nitrate/nitrite concentrations, but may be due to aggregation of leukocytes, platelets, and erythrocytes and/or endothelial consumption among the three experimental groups. The oxygen consumption was linearly dependent on the amount of available oxygen in the perfusate, whereas the glucose consumption was constant and independent of the glucose delivery rate. The present experiments provide new insights into physiological and metabolic mechanisms of action of TNF- alpha for optimization of future treatment schedules involving TNF-alpha. (+info)
Nitric oxide in the endometrium.
(15/20782)
Nitric oxide (NO) is an important mediator of paracrine interactions, especially within the vascular system. It is a powerful inhibitor of platelet aggregation and a potent vasodilator. NO is also a neurotransmitter and it plays a role in cell-mediated cytotoxicity. NO-generating enzymes (nitric oxide synthases, NOS) have been described in the endometrium of a number of species, suggesting that NO might be involved in endometrial function. In human endometrium, endothelial NOS and inducible NOS have been localized to glandular epithelium in the non-pregnant uterus. Weak inducible NOS immunoreactivity has been observed in decidualized stromal cells. NO might participate in the initiation and control of menstrual bleeding. Furthermore, it may play a part in the inhibition of platelet aggregation within the endometrium, where menstrual haemostasis is thought to occur primarily by vasoconstriction rather than clot organization. Endometrially derived NO could also suppress myometrial contractility. Recent attention has focused on the part that NO might play in maintaining myometrial quiescence during pregnancy. NO also appears to relax the non-pregnant myometrium, an action which could be exploited for the medical treatment of primary dysmenorrhoea. (+info)
Control and assessment of the uterus and cervix during pregnancy and labour.
(16/20782)
Preterm labour and resultant preterm birth are the most important problems in perinatology. Countless efforts have failed to establish a single effective treatment of preterm labour, partly because the mechanisms regulating the uterus and cervix during pregnancy are not well understood. New knowledge is needed to inhibit early progression of labour (uterine contractility and cervical ripening), and adequate quantitative tools to evaluate the uterus and cervix during pregnancy are lacking. In this review, we outline studies showing that the uterus (myometrium) and cervix pass through a conditioning step in preparation for labour. This step is not easily identifiable with present methods to assess the uterus or cervix. In the uterus, this seemingly irreversible step consists of changes in the electrical properties to make muscle more excitable and responsive to produce forceful contractions. In the cervix, the step consists of softening of the connective tissue components. Progesterone appears to have a dominant role in controlling both the uterus and cervix, as antiprogestins induce early, preterm conditioning leading to preterm labour. Apparently, nitric oxide (NO) also controls conditioning of the uterus and cervix. In the uterus, NO, in concert with progesterone, inhibits uterine contractility. At term, NO production by the uterus and placenta are decreased and allow labour to progress. In contrast, NO in the cervix increases at the end of pregnancy and it may be the final pathway for stimulating cervical ripening by activation of metalloenzymes. The progress of labour can be assessed non-invasively using electromyographic (EMG) signals from the uterus (the driving force for contractility) recorded from the abdominal surface. Uterine EMG bursts detected in this manner characterize uterine contractile events during human and animal pregnancy. A low uterine EMG activity, measured transabdominally throughout most of pregnancy, rises dramatically during labour. EMG activity also increases substantially during preterm labour in humans and rats. This method may be used one day to predict impending preterm labour and identify control steps and treatments. A quantitative method also assesses the cervix, using an optical device which measures collagen fluorescence in the cervix. The collascope estimates cervical collagen content from a fluorescent signal generated when collagen cross-links are illuminated with excitation light of about 340 nm. The system has proved useful in rats and humans at various stages of pregnancy, and indicates that cervical softening occurs progressively in the last one-third of pregnancy. In rats, collascope readings correlate with resistance measurements made in the isolated cervix, which may help to assess cervical function during pregnancy, and indicate control and treatments. (+info)