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Biomedical subjects

V Lehmann

Publications and source records attributed to V Lehmann.

At least 55 records · Page 3Linked to original sources
PubMed1987

Lethal toxicity of lipopolysaccharide and tumor necrosis factor in normal and D-galactosamine-treated mice.

The toxic properties of human recombinant tumor necrosis factor (TNF) were investigated in mice made hypersensitive to endotoxin by treatment with D-galactosamine. C3H/TifF mice treated with D-galactosamine were rendered sensitive to the lethal effects of submicrogram amounts of TNF. In the absence of D-galactosamine, TNF caused approximately 80% lethality with 500 micrograms. The duration of sensitization to TNF lasted up to 8 h after D-galactosamine administration, that towards LPS, up to 4 h. As with LPS, with TNF sensitization could be inhibited by uridine administered up to 2 h after D-galactosamine/TNF, showing that the early biochemical alterations in the liver known to be necessary for sensitization to LPS are also necessary for sensitization to TNF. In contrast to LPS, the toxicity of TNF was expressed also in D-galactosamine-treated endotoxin-resistant C3H/HeJ mice. The susceptibility of these mice to TNF was identical to that of endotoxin sensitive mice. In the absence of D-galactosamine the toxicity of TNF in C3H/HeJ mice was comparable to that obtained in C3H/TifF mice, being lethal with amounts of the order of 500 micrograms. The present results support the hypothesis that TNF is a mediator of lethal toxicity of endotoxin.

Animals↗
PubMed1987

Tumor necrosis factor augments the immunogenicity and the production of L-ornithine by peritoneal macrophages.

The release of ornithine by macrophages and its correlation with their immunogenicity after treatment with recombinant human tumor necrosis factor-alpha (TNF) were analyzed. TNF was found to augment the capacity of peritoneal macrophages to convert 14C-labeled arginine into L-ornithine and to release L-ornithine into the culture medium. In agreement with the previous observation that L-ornithine potentiates immune responses in vivo, it was also found that treatment of peritoneal exudate cells with TNF augments the capacity of these cells to immunize mice in vivo for a subsequent secondary in vitro cytotoxic response against non-major-histocompatibility antigens.

Animals↗
PubMed1986

Demonstration of membrane receptors for human natural and recombinant 125I-labeled tumor necrosis factor on HeLa cell clones and their role in tumor cell sensitivity.

Samples of natural and recombinant tumor necrosis factor (TNF) were labelled with 125I by use of Iodobeads and were used to study TNF receptor expression on different HeLa clones. Both labeled preparations retained cytotoxic activity after 125I-labelling and were indistinguishable from their unlabeled counterparts on sodium dodecylsulfate/polyacrylamide gel electrophoresis. They bound specifically to TNF-sensitive cells but not to HeLa clones that were resistant to TNF. The binding of 125I-TNF was competed up to 90% by the addition of non-radiolabeled TNF. The binding reached a half-maximal level at 10 pM and saturation at 100 pM. These concentrations approximated those required for cell destruction. Scatchard analysis of the binding data yielded linear plots suggesting that TNF binds to homogeneous TNF receptor sites. The number of receptor sites ranged between 770 and 2200 sites on TNF-sensitive cells. The surface expression of these TNF receptors appeared to be necessary but not sufficient for the cells to become sensitive against the cytotoxic action of TNF. One of the HeLa clones was found to express the same number of TNF receptors as the most sensitive clone, but was nevertheless resistant to the toxic effect of TNF.

Clone Cells↗
PubMed1986

Analysis of protective and cytotoxic immune responses in vivo against metabolically inactivated and untreated cells of a mutagenized tumor line (requirements for tumor immunogenicity).

The immunogenicity of a mutagenized subline (ESb-D) of the weakly immunogenic T-cell lymphoma L 5178 Y ESb has been characterized. The injection of 10(6) ESb-D cells ip did not establish lethal tumors in untreated DBA/2 mice but established tumors in sublethally irradiated mice. Injection of ESb-D cells into otherwise untreated DBA/2 mice established also a state of protective immunity against the subsequent injection of otherwise lethal doses of ESb tumor cells. Protection was only obtained after injection of intact but not UV-irradiated or mitomycin-C-treated ESb-D cells. A direct T-cell-mediated cytotoxic activity was also demonstrable in the spleen cells of DBA/2 mice after injection of ESb-D cells but not ESb cells. The cytotoxic activity was variant specific for ESb-D target cells, and it was induced only with intact but not UV-irradiated or mitomycin C-treated ESb-D cells. This suggested that the induction of protective and cytotoxic immunity may require the persistence of the antigen or unusually high antigen doses. The in vivo priming for a secondary in vitro cytotoxic response, in contrast, was achieved with intact and also with mitomycin C-treated ESb-D cells but again not with UV-irradiated ESb-D cells. This indicated that the metabolic activity was a minimal requirement for the in vivo immunogenicity of the ESb-D tumor line. The secondary cytotoxic activity was demonstrable on ESb-D and ESb target cells and could be restimulated in vitro about equally well with ESb-D and ESb cells. But the in vivo priming was again only obtained with ESb-D cells and not with ESb cells. These experiments thus demonstrated that the requirements for immunogenicity are more stringent in vivo than in vitro, and more stringent for the induction of direct cytotoxic and protective immunity in vivo than for the in vivo priming for secondary in vitro responses.

Animals↗
PubMed1985

Free flow electrophoresis as a tool for enrichment of mutants with temperature-dependent lethal mutations in lipid A synthesis.

Free flow electrophoresis was shown to be a useful tool to enrich for mutants conditionally defective in lipid A synthesis. The method was based on the observation that electrophoretic mobility of bacterial cells is dependent on the structure of lipopolysaccharides and is influenced by lesions in the synthesis of the O-specific chains as well as by lesion in the synthesis of the complete 3-deoxy-D-manno-octulosonic acid (dOclA) lipid A region. Using this procedure a new mutant conditionally defective in dOclA-8-P synthesis was isolated (mutant Ts5). Following a shift to nonpermissive conditions it accumulates a mixture of at least two equally represented lipid A precursor structures. One is made up of glucosamine, phosphate and 3-hydroxymyristic acid in a molar ratio 1.0:1.0:2.0 and lacks dOclA and the nonhydroxylated fatty acids lauric, myristic and palmitic acid. The precursor preparation derived from mutant Ts5 thus differs from previously described lipid A intermediates by the relatively high substitution by palmitic acid. The implications of the above findings to the biosynthesis of lipid A are discussed.

Aldehyde-Lyases↗
PubMed1985

Isolation and structural analysis of two lipid A precursors from a KDO deficient mutant of Salmonella typhimurium differing in their hexadecanoic acid content.

The extraction, purification and structural characterization of two lipid A precursors (Ia and Ib) differing only in one hexadecanoic acid are described. Both precursors were synthesized at elevated temperatures by a new mutant of Salmonella typhimurium (mutant Ts5) which is conditionally defective in synthesis of the 3-deoxy-D-manno-octulosonic acid region of lipopolysaccharides. Both precursors were purified by repeated phenol/chloroform/petroleum ether (PCP) extractions followed by thin layer chromatography. The precursor preparation was free of lipopolysaccharides and phospholipids and contained less than 0.1% protein. Structural analysis which included chemical degradation procedures as well as positive ion laser desorption (LDMS) mass spectroscopy of dephosphorylated lipid A precursors showed together that precursor Ia represents a diphosphorylated glucosamine disaccharide containing two ester, two amide-linked residues of 3-hydroxytetradecanoic acid and lacks the ester-linked dodecanoic, tetradecanoic and hexadecanoic acid as well as 3-deoxy-D-manno-octulosonic acid. Precursor Ib has the same basic structure as precursor Ia, but contains in addition one mol of hexadecanoic acid per mol disaccharide which is linked to the 3-hydroxy group of the amide-bound 3-hydroxy-tetradecanoic acid of the reducing, terminal glucosamine residue. The structure of precursor Ib supports the conclusion that hexadecanoic acid incorporation occurs at an early stage in lipid A biosynthesis prior to the attachment of 3-deoxy-D-manno-octulosonic acid and/or other polar substituents.

Chromatography, DEAE-Cellulose↗
PubMed1985

Comparison of the capacity of two lipid A precursor molecules to express the local Shwartzman phenomenon.

It has been shown recently that a Salmonella lipid A precursor molecule (Ia) and its synthetic counterpart are inactive in expressing the local Shwartzman reaction in both homologous and heterologous systems in combination with lipid A. Precursor Ia contains a bisphosphoryl-beta-1,6-glucosamine disaccharide substituted by 4 mol of (D)-3-hydroxytetradecanoyl residues. Escherichia coli lipid A, on the other hand, which contains two additional non-hydroxylated acyl residues in the form of two 3-acyloxyacyl units, is highly active. We have recently isolated a lipid A precursor molecule (Ib) with the same basic structure as precursor Ia, which contains, however, one additional non-hydroxylated (hexadecanoic) fatty acid forming one 3-acyloxyacyl residue. A comparison of precursor Ia and Ib in homologous and cross-reacting Shwartzman systems confirmed that precursor Ia completely lacked the capacity to prepare the skin for, or to elicit, the Shwartzman reaction. In contrast, precursor Ib was strongly active in inducing the local Shwartzman reaction both when administered intradermally as a preparatory agent and when administered intravenously as a provocatory agent. The results indicate that the additional presence of at least one fatty acid either as such or as an acyloxyacyl residue (as in precursor Ib) is a prerequisite for the ability of the molecule to induce the local Shwartzman phenomenon.

Animals↗
PubMed1985

Suppression of cytotoxic T lymphocyte activation by L-ornithine.

The effect of L-ornithine on several types of immune reactions was analyzed. L-ornithine was found to suppress the activation of cytotoxic T lymphocytes (CTL) in vivo and in vitro. This suppressive effect was not observed with the structural analogues D-ornithine, L-lysine, or putrescine or with the amino acids L-histidine or L-alanine. The concentration of 9 X 10(-3) M L-ornithine was found to mediate a practically complete suppression of the cytotoxic response in vitro if applied on day 0 or day 1 of the culture, but a comparably weak suppression if applied on day 3. The same concentration of L-ornithine had no effect on the production of the lymphokines interleukin 2 (IL 2) and gamma-interferon (IFN-gamma). This concentration of ornithine had also no substantial effect on several types of proliferative responses, including the allogeneic mixed lymphocyte reaction, the concanavalin A-activated IL 2-dependent proliferation of thymocytes, and IL 2-dependent proliferation of the T cell clone W-2. These observations suggest that L-ornithine inhibits selectively the differentiation of CTL effector cells. By the criteria tested, the immunosuppressive effect of L-ornithine is more selective than that of cyclosporine A, which was previously found to suppress not only the activation of cytotoxic activity but also proliferative responses and the production of the lymphokines IL 2 and IFN-gamma.

Animals↗
PubMed1984

Endotoxic properties of chemically synthesized lipid A part structures. Comparison of synthetic lipid A precursor and synthetic analogues with biosynthetic lipid A precursor and free lipid A.

Synthetic lipid A part structures corresponding structurally to a biosynthetic lipid A disaccharide precursor have been analyzed for endotoxic activity in several systems in vivo and in vitro. It was found that a synthetic beta-1,6-linked D-glucosamine disaccharide, which carries four molar equivalents of (R)-3-hydroxytetradecanoyl residues in positions 2, 3, 2' and 3' and phosphoryl groups in positions 1 and 4' (preparation 406), exhibited lethal toxicity, B lymphocyte mitogenicity, the capacity to engender prostaglandin formation in macrophages and to induce endotoxic tolerance, as well as serological lipid A antigenicity. On a weight basis, preparation 406 was of comparable activity to lipid A precursor and bacterial free lipid A. Preparation 406, like lipid A precursor, lacked, however, the ability to induce the local Shwartzman phenomenon and both preparations were of moderate pyrogenicity. Two further synthetic analogues which contained only one phosphoryl group (preparation 404 at C-4', preparation 405 at C-1) showed comparable or diminished activity depending on the test system employed, except in the capacity to inactivate complement where they exhibited, in contrast to preparation 406, significant activity. The results show that the endotoxic principle of lipopolysaccharides, as postulated previously is embedded in the lipid A component. Our results also suggest initial conclusions on the structural requirements for the expression of endotoxin activities.

Animals↗
PubMed1984

Human opsonins to meningococci after vaccination.

Two groups of volunteers were immunized with either a serogroup A plus C meningococcal polysaccharide vaccine or a combined serogroup B polysaccharide-serotype 2 protein vaccine. Serum opsonin responses were measured by chemiluminescence of polymorphonuclear leukocytes exposed to opsonized live meningococci. Two of the six volunteers immunized with the A plus C vaccine had an increase in serum opsonins to group A meningococci, four responded to group C meningococci, and none to group B meningococci. Five other volunteers who were immunized with the combined group B polysaccharide-serotype 2 protein vaccine responded with an increase in serum opsonins to group B meningococci of two different protein serotypes, as well as to a group C-serotype 2 meningococcal strain. Although no booster effect was observed after a second dose of the combined vaccine, both the polysaccharide and the protein components appear to be able to stimulate an opsonin response.

Bacterial Vaccines↗
PubMed1983

Degranulation and enzyme release during phagocytosis of inert particles and of bacteria by polymorphonuclear neutrophil granulocytes.

The degranulation and release of lysosomal (myeloperoxidase, beta-glucuronidase, lysozyme) and cytoplasmic (lactate dehydrogenase-LDH) enzymes from polymorphonuclear neutrophil granulocytes (PMG) during phagocytosis of inert latex particles or bacteria were studied. Degranulation was much faster and more pronounced by phagocytosis of bacteria than of inert particles. A high frequency of lysosome-lysosome as well as lysosome-phagosome fusions suggested that granular material was transported by lysosome- lysosome- phagosome fusions. During bacterial phagocytosis there was evidence of release of granular material into cytoplasm causing enzymatic disintegration. After 60 minutes cell lysis occurred in about 5 per cent of the cells during bacterial phagocytosis. There was non-specific release of LDH during phagocytosis of inert particles, probably due to erythro-phagocytosis. After 60 minutes the release during bacterial phagocytosis amounted to 20-30 per cent of the enzyme content of the cells. A nearly equal release of lysosomal and cytoplasmic enzymes gave support for the idea that cell lysis was the main mechanism of enzyme release.

Escherichia coli↗
PubMed1982

Factors influencing human polymorphonuclear leukocyte chemiluminescence.

Divalent cations (Mg2+, Ca2+) regenerated chemiluminescence (CL) of zymosan-stimulated human polymorphonuclear leukocytes (PMNs) abolished by 0.1 mM EGTA. Phenol red, erythrocytes and free haemoglobin reduced CL. To obtain maximal CL response, the reaction mixture should contain small amounts of protein and glucose. High protein concentrations reduced CL. The test had a pH optimum of 7.1. With respect to stability, however, the most favourable pH was 7.4. The CL response was highly temperature-dependent, and expressed as the initial rate constant of the CL time curve it increased linearly with increasing temperatures from 20 to 37 degrees C. Several factors influence the CL response by human PMNs and test procedures should be standardized.

Blood Platelets↗
PubMed1981

[Differential peridural analgesia for vaginal delivery. A randomised study of its influence on the progress and mode of delivery (author's transl)].

This randomised study of 611 deliveries deals with the side effects of a modified extradural analgesic technique. The modification is based on a flexible reduction of local anaesthetic concentration (bupivacaine) which allows a reliable analgesia without a marked loss of labour sensation and bearing down reflex. Randomisation was achieved by offering this analgesia to the parturients in an alternating 24 hours rhythm. Statistical comparisons were carried out for the following groups of deliveries: 1. for deliveries on both of the two alternating days (Group A). 2. for deliveries with and without extradural analgesia of the peridural days (Group B). - 3. for all deliveries with and without extradural analgesia (Group C). - 4. For deliveries without extradural analgesia on "peridural days" and on "alternating days" (Group D). - The main subjects of investigation were the duration of the two stages of delivery, the dynamics of both the cervical dilation and the movement of the fetal head and the frequency, modes and causes of operative and instrumental deliveries. The safety of the analgesic technique was investigated by evaluating its influence on the status of the newborn and on the maternal regulation of body temperature and circulation. The results were obtained as follows: 1. In the group of parturients who had chosen extradural analgesia several antenatal factors accumulated which evidently had a greater influence on the course of delivery than the analgesic technique itself. - 2. The low concentration of local anaesthetic adapted to the labour pain produced a sufficient analgesia in almost every case. - 3. The duration of both stages of delivery was not altered directly by this technique. An average increase of the duration of the second stage by 116 minutes was due to postponed indications of instrumental and operative deliveries by analgesia. - 4. There was no influence on the overall frequency of operative and instrumental deliveries. Extradural analgesia, however, significantly reduced the frequency of caesarean sections to the same extent as it increased instrumental deliveries from the bottom of the pelvis. - 5. The extradural analgesia had no influence on the status of the newborn. - 6. The regulatory mechanisms of circulation and body temperature were not altered by this dosage of extradural analgesia.

Adult↗
PubMed1980

Selection of bacterial mutants from Salmonella specifically recognizing determinants on the cell surface of activated T lymphocytes. A novel system to define cell surface structures.

The isolation of bacterial mutants from Salmonella is described with specific binding capacities to allogeneically or concanavalin A-activated T lymphocytes. The enrichment of these mutants was achieved by separation of T cell blasts with adhering mutants from nonresponsive lymphocytes and nonadhering bacteria through 1 x g sedimentation. Binding of the mutants was specific for T cells early after antigen or mitogen stimulation. No adherence was observed with unstimulated T or B lymphocytes and with B cell blasts. Further results suggested that the binding of Salmonella mutants was mediated by the heteropolysaccharide moiety of lipopolysaccharides with a specificity for protein receptor sites on activated T lymphocytes. Significantly, these heteropolysaccharides also inhibited the differentiation of prekiller to killer cells from allogeneic mixed lymphocyte cultures but did not depress the proliferative response or the activity of cytotoxic effector cells. Bacterial adherence, as well as polysaccharide activity in functional tests, showed strain specificity since reactivity could only be obtained with activated T cells from AKR, C57BL/6, C3H but not with BALB/c and A/J strain mice. It is discussed whether the heteropolysaccharides mimic the structure of naturally occurring molecules and thus compete for their receptor sites. Selection of bacterial mutants with adherence properties may become a general procedure for detecting cell surface molecules on lymphoid and nonlymphoid cells.

Animals↗