A study on the interactions of surfactin with phospholipid vesicles.
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Surfactin, an acidic lipopeptide produced by various strains of Bacillus subtilis, behaves as a very powerful biosurfactant and posses several other interesting biological activities. By means of differential scanning calorimetry and X-ray diffraction the effect of surfactin on the phase transition properties of bilayers composed of different phospholipids, including lipids forming hexagonal-HII phases, has been studied. The interactions of surfactin with phosphatidylcholine and phosphatidylglycerol seem to be optimal in the case of myristoyl acyl chains, which have a similar length to the surfactin hydrocarbon tail. Data are shown that support formation of complexes of surfactin with phospholipids. The ionized form of surfactin seems to be more deeply inserted into negatively charged bilayers when Ca2+ is present, also supporting the formation of surfactin-Ca2+ complexes. In mixtures with dielaidoylphosphatidylethanolamine, a hexagonal-HII phase forming lipid, surfactin displays a bilayer stabilizing effect. Our results are compatible with the marked amphiphilic nature of surfactin and may contribute to explain some of its interesting biological actions; for instance the formation of ion-conducting pores in membranes. (+info)
Mycoplasmal lipopeptide MALP-2 induces the chemoattractant proteins macrophage inflammatory protein 1alpha (MIP-1alpha), monocyte chemoattractant protein 1, and MIP-2 and promotes leukocyte infiltration in mice.
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Natural as well as experimental infections with pathogenic mycoplasmas lead to cellular responses characterized by early polymorphonuclear leukocyte influx, which in turn is followed by infiltration of macrophages. Since some of the most potent leukocyte chemoattractants are macrophage products, we investigated whether the 2-kDa macrophage-activating lipopeptide (MALP-2) from Mycoplasma fermentans was capable of inducing chemoattractant chemokines and initiating an in vivo inflammatory effect. MALP-2 was a potent in vitro inducer of the chemokines macrophage inflammatory protein 1alpha (MIP-1alpha), monocyte chemoattractant protein 1 (MCP-1), and MIP-2, yielding a maximal response at 0.1 ng/ml (5 x 10(-11) M). Leukocyte infiltration was determined after intraperitoneal injection of MALP-2, liposome-encapsulated MALP-2, and heat-killed mycoplasmas. There was a steady increase in the number of peritoneal cells over 72 h in response to these agents. Polymorph counts were maximal by 24 to 48 h, decreasing thereafter. Monocytes/macrophages had significantly increased after 3 days. MIP-1alpha, MCP-1, and MIP-2 levels in serum or peritoneal lavage fluid were determined. MIP-1alpha and MCP-1 levels were elevated by 2 to 6 h after injection and were still above control values after 24 h. In contrast, MIP-2 levels reached their maximum at 2 h, dropping to control values after 24 h. We conclude that macrophage-stimulating mycoplasmal lipoproteins, exemplified by MALP-2, play an important role in the late phase of phagocyte recruitment at sites of infection and that this is affected by leukoattractive chemokines. (+info)
Lichenysins G, a novel family of lipopeptide biosurfactants from Bacillus licheniformis IM 1307: production, isolation and structural evaluation by NMR and mass spectrometry.
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A series of 9 lactonic lipopeptide biosurfactants was isolated from Bacillus licheniformis IM 1307 as representatives of the lichenysin group and we propose to name them lichenysins G. They were recovered from the culture medium as complex mixtures of molecules having different peptide sequences and different structures of beta-hydroxy fatty acids. Their separation was achieved by a reversed-phase HPLC method leading to eight well-separated compounds. The complete structure of individual isoforms was proposed following the results of amino acid and fatty acid analysis, LSI-MS and 2D NMR spectroscopies. Compared to surfactin, lichenysins G are at least 10 fold more efficient biosurfactants. (+info)
Oral administration of the immunomodulator JBT-3002 induces endogenous interleukin 15 in intestinal macrophages for protection against irinotecan-mediated destruction of intestinal epithelium.
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We recently reported that p.o. administration of the new synthetic bacterial lipopeptide JBT-3002 can protect mice from irinotecan (CPT-11)-induced intestinal injury, but the mechanism was not known. Because interleukin-15 (IL-15) is associated with maintenance of intestinal epithelial cell integrity, we examined whether p.o. administration of JBT-3002 elevates expression of this monocyte-derived cytokine. Four daily i.p. injections of 100 mg/kg CPT-11 were effective against liver metastases produced by CT-26 murine colon cancer cells, but severe damage to the intestinal epithelium and early death of the mice also resulted. Three consecutive daily p.o. doses of JBT-3002 prior to i.p. injection of irinotecan prevented the undesirable side effects of irinotecan without reducing its ability to eradicate liver metastases. Immunohistochemical analyses of the intestines of mice treated with JBT-3002 and CPT-11 demonstrated an increase in the number of dividing cells in the crypts and enhanced expression of IL-15 in lamina propria cells; the increase correlated with increased expression of the IL-15 gene as determined by semiquantitative reverse transcriptase-PCR. In vitro studies demonstrated that JBT-3002 induced expression of IL-15 in peritoneal macrophages but not in normal intestinal epithelial cells (IEC-6). Moreover, the presence of IL-15 decreased irinotecan-mediated cytotoxicity of IEC-6 epithelial cells. These data show that the p.o. administration of JBT-3002 induces expression of IL-15 by macrophages in the lamina propria, which can prevent irinotecan-induced injury to the intestinal mucosa. (+info)
Cytosolic phospholipase A2 activation is essential for beta 1 and beta 2 integrin-dependent adhesion of human eosinophils.
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We examined the role of cytosolic phospholipase A2 (cPLA2) during human eosinophil adherence to ICAM-1- or VCAM-1-coated plates. IL-5-stimulated eosinophils adhered to ICAM-1 through the beta 2 integrin CD11b/CD18, while nonstimulated eosinophils did not. By contrast, nonstimulated eosinophils adhered to VCAM-1 through the beta 1-integrin VLA-4/CD29. Both IL-5-induced adhesion to ICAM-1 and spontaneous adhesion to VCAM-1 corresponded temporally to cPLA2 phosphorylation, which accompanied enhanced catalytic activity of cPLA2. The structurally unrelated cPLA2 inhibitors, arachidonyl trifluoromethylketone and surfactin, significantly inhibited eosinophil adhesion to ICAM-1 and VCAM-1 in a concentration-dependent manner. Inhibition of secretory PLA2, 5-lipoxygenase, or cyclooxygenase did not affect eosinophil adhesion. Addition of arachidonic acid to eosinophils after cPLA2 inhibition with arachidonyl trifluoromethylketone or surfactin did not reverse the blockade of adhesion to ICAM-1 or VCAM-1. However, CV-6209, a receptor-specific antagonist of platelet-activating factor, inhibited all integrin-mediated adhesion. The activated conformation of CD11b as identified by the mAb, CBRM1/5, as well as quantitative surface CD11b expression were up-regulated after IL-5 stimulation. However, cPLA2 inhibition neither prevented CBRM1/5 expression nor blocked surface Mac-1 up-regulation caused by IL-5. Our data suggest that cPLA2 activation and its catalytic product platelet-activating factor play an essential role in regulating beta 1 and beta 2 integrin-dependent adhesion of eosinophils. This blockade occurs even in the presence of up-regulated eosinophil surface integrin. (+info)
Nanometer scale organization of mixed surfactin/phosphatidylcholine monolayers.
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Mixed monolayers of the surface-active lipopeptide surfactin-C(15) and of dipalmitoyl phosphatidylcholine (DPPC) were deposited on mica and their nanometer scale organization was investigated using atomic force microscopy (AFM) and x-ray photoelectron spectroscopy (XPS). AFM topographic images revealed phase separation for mixed monolayers prepared at 0.1, 0.25, and 0.5 surfactin molar ratios. This was in agreement with the monolayer properties at the air-water interface indicating a tendency of the two compounds to form bidimensional domains in the mixed systems. The step height measured between the surfactin and the DPPC domains was 1.2 +/- 0.1 nm, pointing to a difference in molecular orientation: while DPPC had a vertical orientation, the large peptide ring of surfactin was lying on the mica surface. The N/C atom concentration ratios obtained by XPS for pure monolayers were compatible with two distinct geometric models: a random layer for surfactin and for DPPC, a layer of vertically-oriented molecules in which the polar headgroups are in contact with mica. XPS data for mixed systems were accounted for by a combination of the two pure monolayers, considering respective surface coverages that were in excellent agreement with those measured by AFM. These results illustrate the complementarity of AFM and XPS to directly probe the molecular organization of multicomponent monolayers. (+info)
Antiviral and hemolytic activities of surfactin isoforms and their methyl ester derivatives.
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Inactivation of enveloped viruses (VSV, SFV, and SHV-1) by surfactin lipopeptides was dependent on the hydrophobicity, i.e. the number of carbon atoms of the fatty acid, and on the charge of the peptide moiety as well as on the virus species. Surfactins with fatty acid chains of 13 carbon atoms showed very low antiviral activity in comparison to C14 and C15 isoforms. C15 surfactin monomethyl ester also inactivated SFV which was resistant to the mixture of surfactin isoforms as produced by Bacillus subtilis. In contrast, the dimethyl ester showed no virus-inactivation capacity. Disintegration of viral structures as determined by electron microscopy after inactivation of VSV and SFV was comparable to the titer reduction. The effect of the surfactin isoforms and methyl esters on erythrocyte hemolysis correlated with the virus-inactivation capacity. Surfactins with a fatty acid chain moiety of 15 carbon atoms and one negative charge showed the highest antiviral activity. (+info)
Effect of MALP-2, a lipopeptide from Mycoplasma fermentans, on bone resorption in vitro.
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Mycoplasmas may be associated with rheumatoid arthritis in various animal hosts. In humans, mycoplasma arthritis has been recorded in association with hypogammaglobulinemia. Mycoplasma fermentans is one mycoplasma species considered to be involved in causing arthritis. To clarify which mycoplasmal compounds contribute to the inflammatory, bone-destructive processes in arthritis, we used a well-defined lipopeptide, 2-kDa macrophage-activating lipopeptide (MALP-2) from M. fermentans, as an example of a class of macrophage-activating compounds ubiquitous in mycoplasmas, to study its effects on bone resorption. MALP-2 stimulated osteoclast-mediated bone resorption in murine calvaria cultures, with a maximal effect at around 2 nM. Anti-inflammatory drugs inhibited MALP-2-mediated bone resorption by about 30%. This finding suggests that MALP-2 stimulates bone resorption partially by stimulating the formation of prostaglandins. Since interleukin-6 (IL-6) stimulates bone resorption, we investigated IL-6 production in cultured calvaria. MALP-2 stimulated the liberation of IL-6, while no tumor necrosis factor was detectable. Additionally, MALP-2 stimulated low levels of NO in calvaria cultures, an effect which was strongly increased in the presence of gamma interferon, causing an inhibition of bone resorption. MALP-2 stimulated the bone-resorbing activity of osteoclasts isolated from long bones of newborn rats and cultured on dentine slices without affecting their number. In bone marrow cultures, MALP-2 inhibited the formation of osteoclasts. It appears that MALP-2 has two opposing effects: it increases the bone resorption in bone tissue by stimulation of mature osteoclasts but inhibits the formation of new ones. (+info)