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Biomedical subjects

C Galanos

Publications and source records attributed to C Galanos.

At least 37 records · Page 2Linked to original sources
PubMed1997

Dietary testosterone suppresses protective responsiveness to Plasmodium chabaudi malaria.

This study investigates the effect of orally administered testosterone on serum testosterone levels and immune responses including outcome of Plasmodium chabaudi malaria. Female C57BL/10 mice were fed on a diet impregnated with 17 alpha-methyl-testosterone for 3 weeks. This raised the circulating testosterone levels from 0.28 ng/ml to 2.69 ng/ml on the average. In these mice, blood-stage infections of P. chabaudi resulted in a lethal outcome, whereas protective immunity developed in about 80% of mice fed on control diet without testosterone. Dietary 17 alpha-methyl-testosterone reduced the capacity of peritoneal cells to generate reactive oxygen intermediates after stimulation with C3b-coated zymosan and phorbol-myristate-acetate. Also, mice fed on dietary 17 alpha-methyl-testosterone responded to heat-killed Salmonella typhimurium with a higher increase in serum TNF, whereas the induced increase in the production of IL-10 by spleen cells was largely suppressed and no effect was found with respect to the production of IFN-gamma and IL-4. Our data indicate that the method of oral administration of 17 alpha-methyl-testosterone raises circulating testosterone to levels that impair protective immune responses to P. chabaudi malaria.

Animals↗
PubMed1996

Effects of granulocyte colony-stimulating factor on plasma cytokine and cytokine receptor levels and on the in vivo host response to endotoxin in healthy men.

We investigated the effects of granulocyte colony-stimulating factor (G-CSF) on cytokine and cytokine receptor plasma levels and on the in vivo host response to Salmonella abortus equi endotoxin in healthy males. Twenty volunteers received 0.8 ng/kg endotoxin and saline intravenously 1 week apart in randomized order. Twelve hours before both experiments, 10 of these subjects were pretreated with 300 micrograms G-CSF subcutaneously. G-CSF itself increased granulocyte and monocyte counts and the plasma levels of tumor necrosis factor-alpha (TNF-alpha), soluble TNF receptors (sTNF-R) p55, and p75 and interleukin-1 receptor antagonist (IL-1ra). G-CSF did not influence plasma IL-1 beta and IL-6 levels. In the G-CSF-pretreated subjects endotoxin-induced surges in rectal temperature and in the plasma levels of TNF-alpha plasma levels were about 50% increased, and surges in the plasma levels of both sTNF-Rs and IL-1ra were about twice as high as in the control group. Endotoxin-induced increases in IL-6, cortisol, and heart rate were not modified by G-CSF pretreatment. Endotoxin administration induced a transient 50% reduction in leukocyte counts in the G-CSF-pretreated subjects that was not seen in the control group. We conclude that a single stand dose of G-CSF increases the plasma levels of cytokines and cytokine receptors and considerably modifies the host response of healthy humans to a low dose of endotoxin.

Bacterial Toxins↗
PubMed1996

Induction of CD14 expression in Lpsn, Lpsd and tumor necrosis factor receptor-deficient mice.

The involvement of CD14 in lipopolysaccharide (LPS) recognition and signaling has been demonstrated in several studies. For this reason, we investigated whether the resistance of Lpsd mice to LPS might be related to an impaired CD14 expression. We compared the in vivo and in vitro expression of CD14 in Lpsn (LPS sensitive) and Lpsd mice, and its modulation by LPS, killed gram-negative and gram-positive bacteria and double-stranded (ds)RNA. Untreated Lpsn and Lpsd cultured macrophages (M phi), expressed similar amounts of CD14 mRNA and membrane-bound (m)CD14. LPS enhanced CD14 expression only in Lpsn M phi, while all bacteria, or dsRNA, enhanced CD14 in Lpsn and Lpsd M phi. Similarly, in vivo administration of LPS induced or enhanced CD14 mRNA in different organs of Lpsn mice only, while bacteria or dsRNA in both types of mouse. Furthermore, exogenous recombinant tumor necrosis factor (TNF) induced in vivo and in vitro enhanced CD14 expression in Lpsn, Lpsd and also in TNF receptor 2-deficient (TNFR2-/-) mice, but failed to do so in TNFR1-/- mice, showing that TNFR1 mediates the effect of TNF on CD14. However, LPS, bacteria and dsRNA induced CD14 in both TNFR2-/- and TNFR1-/- mice to a similar extent, revealing that this induction does not require TNF signaling.

Animals↗
PubMed1996

Biochemical characterization of lipopolysaccharides extracted from a hydrophobic strain of Pasteurella multocida.

Lipopolysaccharides were extracted from freeze-dried cells of Pasteurella multocida strain P-1581 (serotype 8) by the phenol-chloroform-petroleum ether method and biochemically analysed using standard procedures. The primary neutral sugars were glucose, galactose and heptose. No deoxy sugars were detected. Amino sugars included galactosamine, glucosamine and glucosamine-6-phosphate. 3-Deoxy-D-manno-2-octulosonic acid was present at a relatively low concentration. The analyses included identification and quantification of phosphate and alanine. The primary fatty acids and their approximate relative ratios were 3-hydroxytetradecanoate and tetradecanoate 2:1. Tetradecanoic acid was bound almost exclusively by ester linkages. 3-Hydroxytetradecanoic acid was bound primarily by amide linkages, although significant numbers of ester-bound residues were noted. Sodium dodecyl sulphate-polyacrylamide gel electrophoresis analyses indicated that the lipopolysaccharides were of low molecular weight.

Amino Sugars↗
PubMed1996

Phase II trial of intravenous endotoxin in patients with colorectal and non-small cell lung cancer.

We report the immunological and clinical results of a phase II trial with intravenously administered highly purified endotoxin (Salmonella abortus equi) in patients with advanced cancer. 15 patients with non-small cell lung cancer and 27 with colorectal cancer were entered into the study. 37 evaluable patients received at least four injections of endotoxin (4 ng/kg body weight) and 1600 mg ibuprofen orally in 2-week intervals. Transient renal (WHO grade 0-1) and hepatic (WHO grade 0-4) toxicities occurred in several patients. Constitutional side-effects such as fever, chills and hypotension could not be prevented completely by pretreatment with ibuprofen. 3 patients in the colorectal cancer group demonstrated objective responses (1 complete remission (CR), 2 partial remission (PR)). The complete remission has been maintained for more than 3 years, while the partial remissions were stable for 7 and 8 months, respectively. Only marginal antitumour effects were seen in the lung cancer group. Tolerance of the macrophage system to the stimulatory effect of endotoxin, as measured by human necrosis factor alpha (TNF-alpha) release into serum, built up after the first administration and remained at a steady-state level after each subsequent injection. In constrast, rising CD4:CD8 ratio and release of tumour necrosis factor beta (TNF-beta) indicated the continuing activation of the lymphocyte system by repetitive injections of endotoxin.

Adult↗
PubMed1996

Diurnal variations in the human host response to endotoxin.

To investigate diurnal variations in the host response to endotoxin, Salmonella abortus equi endotoxin (0.8 ng/kg) was given intravenously to healthy men in a placebo-controlled design at 0900 or 1900 h. The time course of rectal temperature and the plasma levels of tumor necrosis factor- alpha (TNF-alpha), interleukin-6 (IL-6), adrenocorticotropic hormone (ACTH), and cortisol were monitored for 11 h following the injections. The time of day did not affect the endotoxin-induced increase in plasma TNF-alpha or IL-6. However, subjects who received endotoxin in the evening, when endogenous glucocorticoid levels were low, showed about twice the increases in rectal temperature and plasma ACTH and cortisol levels as those who received endotoxin in the morning, when endogenous glucocorticoid levels were high. These results demonstrate diurnal variations in the human susceptibility to endotoxin that may be due to a suppression of the biologic effects of TNF-alpha and IL-6 by endogenous glucocorticoids.

Adrenocorticotropic Hormone↗
PubMed1995

Interferon beta, a cofactor in the interferon gamma production induced by gram-negative bacteria in mice.

The interferon (IFN) gamma production of splenocytes from closely related C57BL/10ScSn (Sn) and C57BL/10ScCr (Cr) mice was compared. Concanavalin A and CD3 monoclonal antibodies induced high levels of IFN-gamma in both Sn and Cr splenocytes. By contrast, treatment with gram-negative bacteria induced IFN-gamma only in Sn splenocytes; in Cr splenocytes, the IFN-gamma response was heavily impaired. The IFN-gamma induction by bacteria requires the cooperation of IFN-gamma-producing cells with macrophages. Depletion of macrophages from Sn splenocytes resulted in the loss of ability to produce IFN-gamma after bacterial stimulation. Reconstitution with new Sn macrophages restored the IFN-gamma responsiveness, whereas reconstitution with Cr macrophages failed to do so. Normal function of IFN-gamma-producing cells and a defective function of macrophages of Cr mice was demonstrated by evidence showing that whole or macrophage-depleted Cr splenocytes, when supplemented with Sn macrophages, acquire the ability to produce IFN-gamma in response to bacteria. A similar effect was achieved by supplementing Cr splenocytes with supernatants of bacteria-stimulated Sn macrophages or with recombinant murine IFN-beta or IFN-alpha. Preincubation of active macrophage supernatants with antibodies to IFN-beta suppressed the helper activity for Cr splenocytes. Moreover, the bacteria-induced production of IFN-gamma by Sn splenocytes could be inhibited by antibodies to murine IFN-beta. The results provide evidence that IFN-beta is an important cofactor of IFN-gamma induction, which is not induced in Cr mice by gram-negative bacteria.

Animals↗
PubMed1995

Lipid A binding to neonatal and adult red blood cells.

Lipid A is responsible for the endotoxic activities of gram-negative bacteria. Binding of lipid A (50 micrograms/ml) to RBC was studied using a passive hemolysis test. RBC from adults, cord and venous RBC from full-term infants and RBC from preterm infants were studied. Lipid A sensitized RBC were hemolysed with anti-lipid A and guinea pig complement. Hemolysis was expressed as hemoglobin concentration (absorbance at 546 nm) in the supernatant after centrifugation. 50 micrograms/ml lipid A did not increase spontaneous hemolysis (< 3%) after 60 min of incubation in any of the four groups. The passive hemolysis test did not result in additional hemolysis (5%) of umbilical cord RBC. RBC of preterm infants showed a significant increase in hemolysis (24%) after 60 min of incubation. In RBC of full-term neonates, increased hemolysis (14 and 46%) was detected after 30 and 60 min of incubation. Adult RBC hemolysed stronger (26 and 64%) after 30 and 60 min than neonatal RBC. We conclude that lipid A binds less to neonatal RBC compared to adults.

Adult↗
PubMed1995

Endotoxin (Salmonella abortus equi) in cancer patients. Clinical and immunological findings.

1. LPS phase I trial revealed MTD I of 1.0 ng/kg body weight and MTD II of 5.0 ng/kg body weight, the latter given together with ibuprofen (1,600 mg). 2. LPS phase II trial, using 4,0 ng/kg body weight plus ibuprofen in a biweekly schedule didn't show any response in patients with non-small cell lung cancer but 1 CR and 2 PR (13% response rate) in colorectal cancer patients. 3. The LPS tolerance is specific for each cytokine and mediator in regard to the kinetic and degree of its development. INF-gamma prevents the tolerance development to several cytokines with the exception of IL-8. 4. No tolerance was found in cell adhesion, phospholipase A2, and soluble TNF receptor I and II. Priming of ex vivo cytokine production of cytokines was found in mononuclear cells. 5. Synthetic LPS partial structure SDZ-MRL 953 (I) induces a cytokine pattern, which is profoundedly different from that of LPS, (II) with an inverse TNF to G-CSF relation, (III) is (without any ibuprofen treatment) remarkably low toxic, (IV) and downregulates the TNF-response to LPS markedly.

Carcinoma, Non-Small-Cell Lung↗
PubMed1994

Discordant adaptation of human peritoneal macrophages to stimulation by lipopolysaccharide and the synthetic lipid A analogue SDZ MRL 953. Down-regulation of TNF-alpha and IL-6 is paralleled by an up-regulation of IL-1 beta and granulocyte colony-stimulating factor expression.

Human peritoneal macrophages were exposed to increasing doses of LPS or a synthetic lipid A analogue (SDZ MRL 953) and production of the cytokines IL-1 beta, IL-6, TNF-alpha, and G-CSF was assessed at the protein and mRNA level. Cells were also prestimulated with low doses of LPS and SDZ MRL 953 to study their adaptation to a secondary challenge with high doses of LPS. The ability of macrophages to produce high levels of TNF-alpha and IL-6 after stimulation with LPS could be relieved almost completely by preincubating cells with low doses of LPS. Decreases of TNF-alpha and IL-6 production resulted from inhibition of gene transcription and/or changes in mRNA stability, as transcript levels of these cytokines were down-modulated by the process of LPS adaptation. Surprisingly, however, adapted cells were able to synthesize even larger quantities of G-CSF and IL-1 beta when exposed to a secondary LPS challenge. mRNA levels of the adapted cells remained unaltered for IL-1 beta, but were slightly increased for G-CSF as assessed by Northern blot analysis. High doses of the synthetic lipid A analogue SDZ MRL 953 were also able to adapt macrophages to a secondary LPS challenge by down-regulating TNF-alpha and IL-6 production, whereas priming secretion of G-CSF and IL-1 beta as well. We describe here the discordant adaptation of human peritoneal macrophages to a secondary LPS stimulus in vitro. These findings appear to have ramifications for the in vivo endotoxin response during inflammation and also Gram-negative septicemia.

Dinoprostone↗
PubMed1994

Influence of the glucose concentration on the yield of biomass and lipopolysaccharide in Salmonella cultures.

Two Salmonella S-forms and two R-mutants were cultivated in complex medium supplemented with different amounts (0.5-3%) glucose. Cultivation was performed batchwise in a fermentor under aerobic conditions. With all strains investigated, the yield of bacterial mass increased with increasing concentration of glucose. In the case of three strains, the % LPS content of bacteria also increased, thus achieving the aim of this investigation. The synthesis of bacterial mass and LPS did not proceed in parallel and differed from strain to strain. At optimal glucose concentration, the yield of LPS could be increased up to 250%. The chemical composition of the LPS was independent of the glucose concentration. The individual strains exhibited an identical composition with regard to lipid A and polysaccharide when cultured at different glucose concentrations. The uniformity of the molecular distribution of LPS could also be confirmed by SDS-polyacrylamide gel electrophoresis. In the S-form LPSs, also the proportion of the unsubstituted R-form LPS was not affected by the glucose concentration in the culture medium. The present results demonstrate that optimisation of the cultivation conditions with respect to the glucose concentration of the medium would be of advantage especially for Salmonella strains that are cultivated frequently.

Biomass↗
PubMed1994

A 14,000 MW lipoprotein and a glycolipid-like structure of Borrelia burgdorferi induce proliferation and immunoglobulin production in mouse B cells at high frequencies.

Sonicated preparations of Borrelia burgdorferi are able to stimulate unselected resting BALB/c spleen cells to proliferate and to produce immunoglobulin in vitro. FACS analysis of target cells prestained with an integrated cell-surface marker as well as cell-depletion experiments demonstrate that the majority of responding lymphocytes are B cells. Limiting dilution analyses of resting B cells revealed high frequencies of cells producing IgM (F 1/11-1/62) or IgG (F 1/5-1/163) in response to B. burgdorferi sonicate (B.b. sonicate). These numbers were similar to those obtained with lipopolysaccharide (LPS) (IgM: F 1/20-1/84; IgG: F 1/14-1/85) or a synthetic lipopeptide of Braun's Escherichia coli lipoprotein (IgM: F 1/15, 1/19; IgG: F 1/148, 1/34). The mitogenic structure(s) expressed by B. burgdorferi is distinct from LPS, as similar proliferative responses were obtained with B cells from LPS-resistant (C57BL/10ScCr and C3H/HeJ) and LPS-susceptible (C57BL/10ScSn, C3H/HeN) mice. Furthermore, B-cell mitogenic properties were also found in two distinct fractions of a phenol-chloroform-petroleum ether extract of B. burgdorferi: they consisted of a lipoprotein distinct from the outer surface proteins (Osp) A and B and glycolipid-like structures, respectively. These data suggest that spirochetes express a multitude of distinct structures with mitogenic activity for B cells including various lipoproteins as well as glycolipid(s).

Animals↗
PubMed1994

Contribution of tumor necrosis factor-alpha and glucocorticoid in hydrazine sulfate-mediated protection against endotoxin lethality.

Hydrazine sulfate pretreatment has previously been shown in our laboratory to protect normal mice against endotoxin and D-galactosamine-sensitized mice against both exogenous tumor necrosis factor (TNF) and endotoxin. An intact pituitary is required for manifestation of the protective effects. Further, we have demonstrated that hydrazine sulfate pretreatment specifically modulates the TNF response to lipopolysaccharide (LPS) in mouse macrophages in vitro. This in vivo study was performed to test whether a reduced TNF response and/or increased glucocorticoid response may contribute to hydrazine sulfate protection against LPS-induced lethality in vivo. The results presented here establish that hydrazine sulfate pretreatment selectively attenuates circulating TNF levels following LPS challenge. Moreover, adrenalectomy abrogates hydrazine sulfate protection but does not prevent hydrazine sulfate attenuation of circulating TNF levels in response to LPS. Hydrazine sulfate-mediated protection is, however, restored permissively by corticosterone. Thus, the mechanism by which hydrazine sulfate protects against LPS lethality in adrenalectomized mice includes TNF modulation in response to endotoxin, as well as a pivotal requirement for glucocorticoid.

Adrenalectomy↗
PubMed1993

Mechanisms of endotoxin shock and endotoxin hypersensitivity.

Endotoxins (lipopolysaccharide, LPS) are biologically active substances present in Gram-negative bacteria. Injection of purified LPS into experimental animals leads to the development of many biological activities that can lead to shock with lethal outcome. The biological activities of LPS are not direct effects of the LPS molecule since LPS usually expresses no direct cytotoxic activity. The toxic and other biological properties of LPS are caused indirectly through the action of endogenous mediators that are formed following interaction of LPS with cellular targets, macrophages occupying a key position in the development of endotoxin shock. The interaction of LPS with macrophages may proceed directly leading to activation of these cells, with subsequent synthesis and secretion of a number of endogenous mediators which initiate the different biological activities of LPS. Tumor necrosis factor alpha (TNF-alpha), a macrophage derived cytokine, is a primary mediator of the lethal action of endotoxin. Sensitivity to LPS is genetically determined, varying considerably among different species. The sensitivity of normal animals (mice) to endotoxin may be enhanced considerably under different experimental conditions that include treatment with live (infection) or killed Gram-negative and -positive bacteria. Sensitization to endotoxin proceeds in all LPS-responder strains investigated and in the LPS-resistant mice of the strain C3H/HeJ. It does not proceed in a second LPS-resistant strain, C57BL/10ScCr. The absence of sensitization in the latter mice was found to be due to an impaired IFN-gamma production. IFN-gamma could be identified as the mediator of endotoxin hypersensitivity induced by bacteria.

Animals↗
PubMed1993

Cross-binding activity and protective capacity of monoclonal antibodies to lipid A.

Six hybridoma clones (1M, 4M, 9M, 11M, 18M and 31G), secreting monoclonal antibodies (mAbs) against lipid A were obtained after fusion between cells of mouse myeloma line X63-Ag8.653 and spleen cells from BALB/c mice immunized with acid treated Salmonella minnesota bacteria coated with additional free lipid A. The specificity and cross-binding activity of the mAbs were characterized in ELISA by using synthetic lipid A analogs as well as different lipid A and lipopolysaccharides (LPS) extracted from R- and S-form bacteria. It was found that the antibodies recognize epitopes in which phosphate groups, especially those at the C4' position of the glucosamine backbone of lipid A, were present. These epitopes were accessible also for the antibodies in purified intact LPS. By using a set of core glycolipids with increasing completion of the core region of the molecule and S-LPSs it was shown that the mAbs cross-reacted with a variety of R- and S-form LPS. The binding activity decreased with increasing length of the polysaccharide chain. The mAb did not prevent ultimate lethality of mice challenged with Klebsiella pneumoniae B and Salmonella typhimurium C5. However a delay of mortality rate of mice pretreated with antibodies 18M and 31G and infected with K. pneumoniae was seen.

Animals↗
PubMed1993

Antibodies from chronically infected cystic fibrosis patients react with lipopolysaccharides extracted by new micromethods from all serotypes of Pseudomonas aeruginosa.

Micromethods were developed to extract lipopolysaccharide (LPS, endotoxin) from single bacterial colonies of the 20 recognized Pseudomonas aeruginosa type strains. The appearance of these LPSs in polyacrylamide gel electrophoresis (PAGE) and their reactivity with serum of cystic fibrosis (CF) patients chronically infected with P. aeruginosa was studied. Silver staining of LPS after PAGE showed that 13 of the P. aeruginosa LPSs had high numbers of O-repeating units arranged in 1-4 clusters of banding. Low-molecular-weight LPS fractions were more prominent in six of the serotype strains, of which O:7 and O:14 appeared semi-rough. Corresponding immunoblots using the CF sera showed LPS patterns very similar to the silver-stained appearance, indicating an immune reaction to all P. aeruginosa LPS including that from the newly discovered O:18, O:19 and O:20. This was unexpected since only a few serotype strains (mostly O:3, O:6 and O:9) had been isolated from the patients. Absorption experiments using purified and chemically defined P. aeruginosa rough LPS and smooth LPS suggested these immune reactions were due to antibodies cross-reactive to core/lipid A as well as to lower molecular weight O-polysaccharides or "A-bands". However, in some cases O:3, O:6, and O:9 LPSs were also found to contain additional distinct O-epitopes. Immune recognition of various polyagglutinable P. aeruginosa LPSs seemed also to be caused by cross-reactive antibodies. The described microextraction methods, followed by PAGE and silver staining or immunoblotting, are easy and convenient techniques with which to study antibodies against LPS epitopes and to screen for LPS phenotypic appearance using only a few bacterial colonies from larger numbers of Gram-negative bacterial strains.

Antibodies↗
PubMed1993

Endogenous and exogenous glucocorticoids have different roles in modulating endotoxin lethality in D-galactosamine-sensitized mice.

Endotoxin sensitivity and dexamethasone protection have been assessed in mice that were adrenalectomized and also treated with D-galactosamine at the time of endotoxin challenge. Our data establish that adrenalectomy did not detectably alter the magnitude of the increased sensitivity induced by D-galactosamine alone. Furthermore, protection provided by acute exogenous glucocorticoid treatment was still demonstrable in these mice and was not influenced by chronic experimentally induced glucocorticoid deficiency. Our data confirm that the adrenalectomized mouse model of endotoxin lethality is characterized by increased sensitivity to endotoxin and establish that the magnitude of this sensitizing effect is more than 100-fold. We also show for the first time that adrenalectomy causes an appreciable kinetic shift in the endotoxic crisis and that dexamethasone, given at the time of endotoxin challenge, will significantly reverse the increased sensitivity to lethality. Our results indicate that the protective effects of corticosteroids may involve important chronic as well as acute responses. In particular, we conclude that endogenous glucocorticoid need not always increase host resistance to endotoxin, nor does such a circumstance eliminate the possibility for exogenous glucocorticoid-mediated protective effects.

Adrenalectomy↗