Search PubMed⌕ Search

Biomedical subjects

G Bruchelt

Publications and source records attributed to G Bruchelt.

At least 73 records · Page 4Linked to original sources
PubMed1987

Methods for the determination of the interferon-induced enzyme 2'-5' oligoadenylate synthetase in mononuclear blood cells.

The determination of 2'-5'-oligoadenylate synthetase in peripheral blood mononuclear cells is used as a biological response parameter during therapy with interferon and in the diagnosis of diseases related to the interferon system. In this communication, some general aspects concerning the preparation of 2'-5'-oligoadenylate synthetase from peripheral blood mononuclear cells and the incubation conditions of the 2'-5'-oligoadenylate synthetase reaction are reported. Four analytical procedures for the determination of the products formed during the 2'-5'-oligoadenylate synthetase reaction were comparatively investigated and the advantages and limitations of the assays are discussed. As an example of possible clinical application, the levels of 2'-5'-oligoadenylate synthetase were determined in the mononuclear cell fraction of peripheral blood from healthy persons as well as from children with chronic myelogenous leukaemia.

2',5'-Oligoadenylate Synthetase↗
PubMed1987

Kinetics of restoration of interferon production after bone marrow transplantation in man.

Peripheral blood mononuclear cells (PBMC) from 21 patients after bone marrow transplantation (BMT) were studied for their capacity to produce interferon (IFN) in vitro. The basal and IFN-stimulated 2-5 A synthetase activity was also investigated as a marker of the cells' ability to respond to exogenous IFN. All but one patients received cyclosporin A as a prophylaxis against graft-versus-host disease (GVHD). GVHD was diagnosed in three patients. IFN production in response to stimulation with phytohemagglutinin or poly I:C was not detectable in most patients without GVHD until 7 months after grafting. However, in a proportion of recipients without GVHD, studied early after BMT, transient normal IFN production was observed. In contrast to patients without GVHD, PBMC from patients with GVHD produced stable high levels of IFN when stimulated in vitro. The impairment of IFN production did not correlate with conditioning regimens, infection, plasma cyclosporin levels or the lymphocytes' blastogenic response to the mitogens. Addition of interleukin-2 (IL-2) to culture medium of fresh unresponsive PBMC restored only partially the defective IFN production. Similarly, T-cell lines propagated in IL-2 conditioned medium, from unresponsive PBMC, produced low levels of IFN gamma when stimulated with PHA. The basal activity of 2-5 A synthetase in PBMC from patients without GVHD could not be stimulated, during the first 3 months after BMT, by the cultivation of cells with IFN alpha.(ABSTRACT TRUNCATED AT 250 WORDS)

2',5'-Oligoadenylate Synthetase↗
PubMed1986

Clinical experiences in the treatment of neuroblastoma with 131I-metaiodobenzylguanidine.

Treatment of neuroblastoma is an unsolved problem of pediatric oncology. In spite of highly intensified chemotherapy, the long-term survival rate of children with a metastatic neuroblastoma is below 10%. We therefore used 131I-metaiodobenzylguanidine (MIBG) for the first time to treat children with a neuroblastoma in relapse or primary unresponsiveness to chemotherapy. We had previously demonstrated that MIBG is useful for the scintigraphic imaging of neuroblastoma lesions and had investigated the cytotoxicity and uptake of MIBG in various neuroblastoma cell lines. We treated 6 children with neuroblastoma in a total of 19 courses. Three of the children suffered from a relapse of neuroblastoma; 3 had never gained a remission. Four of the 6 children lost their bone pain and fever during the first 3 days. In 5 of the 6 children the solid tumor as well as the bone marrow infiltration responded to MIBG treatment, with responses ranging from transitory decrease of the tumor mass to complete disappearance of abdominal tumors. We also witnessed a stabilization of osteolytic lesions, a decrease in elevated serum catecholamines, and a decrease in bone marrow infiltration. Five of the 6 children died of tumor progression 55-249 days after the first MIBG treatment.

3-Iodobenzylguanidine↗
PubMed1985

The role of reactive oxygen compounds derived from 6-hydroxydopamine for bone marrow purging from neuroblastoma cells.

6-Hydroxydopamine(6-OHDA), a specific neurotoxin against sympathetic nerve cells, is a drug already used for purging of bone marrow from neuroblastoma cells before autologous bone marrow transplantation. However, we could not detect significant differences in the toxicity of 6-OHDA against neuroblastoma and other tumor cells under the purging conditions clinically used. In contrast, bone marrow stem cells were much more resistant. The unspecific toxic effect of 6-OHDA is caused by H2O2 or H2O2-derived products which are generated by auto-oxidation in the incubation medium before a significant amount of 6-OHDA is taken up by the cells. Withdrawal of oxygen during the incubation period and subsequent incubation with an oxygen containing medium led to a more specific destruction of neuroblastoma cells which can take up 6-OHDA selectively.

Ascorbic Acid↗
PubMed1985

Specific uptake of m-[125I]iodobenzylguanidine in the human neuroblastoma cell line SK-N-SH.

The uptake of m-[125I]iodobenzylguanidine (mIBG), a compound structurally analogous to the antihypertensive drug guanethidine, was examined in various human cell lines. Of three neuroblastoma lines, SK-N-LO, IMR-32, and SK-N-SH, only the last showed specific uptake of the compound. In contrast, only a nonspecific uptake could be demonstrated for the other neuroblastoma lines, as well as for an osteogenic sarcoma line (SAOS-2) and a melanoma line (IgR 3). Based on analyses of uptake characteristics from Lineweaver-Burk plots it is evident that two different transport mechanisms are responsible for mIBG uptake into SK-N-SH cells: a nonspecific diffusion mechanism, and a specific, active uptake system. The latter was dramatically reduced at 4 degrees compared to 37 degrees, as well as in the presence of ouabain or the absence of oxygen. A competitive inhibition of the transport of mIBG by norepinephrine was observed. When drug-treated SK-N-SH cells were incubated in fresh medium, 20 to 30% of mIBG was still retained in the SK-N-SH cells 24 h after the end of incubation with mIBG, whereas no mIBG was detectable in SK-N-LO cells already after 1 h.

3-Iodobenzylguanidine↗
PubMed1984

Comparative studies on the oxidative processes during phagocytosis measured by luminol-dependent chemiluminescence.

Four different methods, using luminol-enhanced chemiluminescence, are described for assessment of the oxidative processes associated with the phagocytosis of opsonized zymosan by granulocytes. The techniques described in the literature using whole blood as a substrate gave unreliable results due to quench effects of contaminating red blood cells, unpredictable serum interferences, or varying ratios of granulocytes and zymosan. In order to avoid these side effects, rapid and simple methods for the isolation of the granulocytes were applied, e.g. lysis of erythrocytes by ammonium chloride, or the isolation of granulocytes on a percoll gradient. The latter procedure proved to be reliable, simple and sufficiently quick for a routine application.

Ammonium Chloride↗
PubMed1983

Phagocytic activity of granulocytes and alveolar macrophages after burn injury measured by chemiluminescence.

Despite substantial progress in handling the acute phase, about 50 per cent of all severely burned patients are still subject to lethal infections during the later stages of the burn disease. Such patients frequently succumb to infections by opportunistic bacteria and viruses of normally low virulence indicating that the antiinfectious host defences are severely compromised. The present study was conducted in order to evaluate the effects of severe thermal injury on the two major categories of phagocytic cells, the circulating phagocytes of the blood and the alveolar macrophages as one population of the fixed phagocytes of the reticulo-endothelial system. The cells were isolated from burned and unburned rats. For quantitative assessment of the phagocytic function the chemiluminescence associated with the phagocytosis of opsonized zymosan particles was measured, using luminol as a chemiluminigenic probe. It turned out that as a consequence of the thermal trauma the phagocytic activity as measured by chemiluminescence is reduced in granulocytes as well as in alveolar macrophages.

Animals↗
PubMed1983

Studies on the metabolic conversion of ascorbate.

In the intact animal guinea pigs metabolize (1-14C)ascorbic acid much faster to (14C)carbon dioxide (peak exhalation at 30 min) than rats (peak exhalation at 2 to 3 h) following single oral administration, but total excretion was comparable. This finding might be related to the differences in the absorption mechanism of ascorbic acid in these species. The large difference in retention capacity of ascorbic acid in the liver is suggested to be due to multiple recirculation of ascorbic acid in the guinea pig when compared to the rate. Homogenate preparations of rat stomach, small intestine or liver as well as cultured intestinal microbial flora did not cause metabolic degradation on incubation with (1-14C)ascorbic acid to (14C)carbon dioxide. It is therefore suggested that the observed excretion of (14C)carbon dioxide is due to spontaneous non-enzymatical reaction in liver and possible other tissues. Analysis of ascorbic acid metabolites formed on incubation by analytical isotachophoresis suggests that ascorbic acid is a rather stable substrate whereas dehydroascorbate and 2,3-diketogulonic acid are rapidly degraded. This allows the assumption that in vivo metabolism of ascorbic acid might not involve dehydroascorbic acid. Our data do not support the hypothesis that ascorbic acid undergoes presystemic metabolism to carbon dioxide in the intestinal wall.

2,3-Diketogulonic Acid↗
PubMed1980

Analytical isotachophoresis: a rapid and sensitive method for determination of urinary oxalate.

A variety of procedures have been described in the literature for quantitative determination of urinary oxalate concentrations. However, a rapid and sufficiently simple technique for routine determinations is lacking. This study presents an isotachophoretic technique for measurement of urinary oxalate. Complete separation of the oxalate anion from accompanying ions is achieved in a PTFE-capillary between two electrodes according to differences in the net mobility. Short analysis time, little pretreatment of the urine samples, and extremely high resolving power and accuracy are the significant advantages of this newly developed method. This new technique was shown to be of significant value in studying the role of oxalate in urinary stone disease.

Electrophoresis↗
PubMedDate not supplied

Effects of granulocytes on human neuroblastoma cells measured by chemiluminescence and chromium-51 release assay.

We investigated whether polymorphonuclear leukocytes (PMN) are able to kill human neuroblastoma cells either directly or if coated with antibody MAb 14.18 that recognizes ganglioside GD2 present on the cell surface of most neuroblastoma cells. Neuroblastoma cells could not be destroyed directly, whereas in the antibody-dependent reaction (ADCC-reaction) they were easily eliminated. In order to answer the question whether reactive oxygen intermediates are involved in this process, chemiluminescence measurements were performed. Compared to the signals that could be measured using opsonized zymosan as stimulus, only weak CL-signals could be registered during the ADCC reaction. Pretreatment of PMN with granulocyte-macrophage colony stimulating factor (GM-CSF) enhanced the CL-signals, catalase and SOD reduced it; however, cell killing was only slightly influenced in the presence of catalase and superoxide dismutase. These data suggested that reactive oxygen compounds do not play a prominent role in the killing process. Definitive evidence for this suggestion could be obtained using PMN from a patient with chronic granulomatous disease (CGD): MAb 14.18 coated neuroblastoma cells could be killed effectively, but no CL-signal could be registered, either in the ADCC-reaction or using opsonized zymosan as stimulus.

Catalase↗
PubMedDate not supplied

Measurement of endogenous and TNF alpha-mediated H2O2 production in supernatants of SK-N-SH neuroblastoma cells with an enhanced chemiluminescence assay.

A sensitive peroxidase-dependent luminol-enhanced chemiluminescence (ECL) assay for determination of hydrogen peroxide (H2O2) generation by tumour cells was established. This test system allows determination of H2O2 in concentrations as low as 25 pmol (50 nmol/L) and yields results which are comparable to those obtained using a less sensitive photometric method and a previously described scopoletin fluorescence assay. After 3 h incubation time 10(4) SK-N-SH neuroblastoma cells released 60 +/- 5 pmol H2O2 in the supernatant and this level was significantly (p < 0.025) increased by about 70% in the presence of 5 pmol (100 ng/mL) recombinant tumour necrosis factor alpha (TNF alpha). In contrast, H2O2 production was slightly reduced by TNF alpha at a very low concentration of 0.5 fmol (0.01 ng/mL).

Cell Line↗
PubMedDate not supplied

Toxic reactions of oxidized LDL on cells of acute myeloid leukemia.

In AML patients LDL concentration of the serum is reduced due to the high LDL receptor activity of the AML cells. This phenomenon enables the use of LDL particles as vehicles for drug targeting. Toxic lipid peroxides and aldehydes were introduced into LDL particles by the simple but effective oxidation with 10 microM CuSO4. Up to 250 nmol peroxides and 6 nmol malondialdehyde were formed per mg LDL protein within 30 h of oxidation. This oxidized LDL is effectively taken up by AML cells of the FAB type M3 and M5 indicating the presence of scavenger receptors on these cells. Within 96 h 61-84% of the AML cells are killed by the oxidized LDL. Our results open a possibility to achieve specificity for targeting lipophilic antineoplastic drugs towards AML cells using oxidized LDL as vehicles. The use of oxidized LDL as drug carrier is recommended for purging of AML bone marrow because hematopoietic stem cells that don't possess scavenger receptors are protected from toxic action.

Copper Sulfate↗
PubMedDate not supplied

[The role of interferons in neuroblastoma. 1: Antiproliferative effects].

Antiproliferative effects of interferon alpha, beta and gamma were investigated on several human neuroblastoma cell lines using the soft agar colony forming assay and the MTT-test. Investigations were carried out in order to prove whether there is any relationship between antiproliferative effects, inhibition of N-myc expression and the 2-5A system. Growth of neuroblastoma cells was inhibited by all three kinds of interferons in a concentration-dependent manner, however, rather high concentrations were necessary in some cell lines. Expression of N-myc oncogen was not inhibited by interferon-beta and no relationship between antiproliferative effects and the 2-5A system was observed. A vector containing a small N-myc fragment in antisense direction was constructed and transferred into the interferon insensitive human neuroblastoma cell line LS. After transformation, LS cells became sensitive to interferon beta: Proliferation as well as N-myc expression were inhibited and these processes are most probably associated with activation of the 2-5A system.

2',5'-Oligoadenylate Synthetase↗
PubMedDate not supplied

[The role of interferons in neuroblastoma. 2: Immunomodulatory effects].

The role of Interferon-gamma (IFN-gamma) in the immunotherapy of neuroblastoma was investigated. In vitro experiments showed that IFN-gamma augments the cytotoxicity of Natural Killer (NK) cells and of interleukin 2 (IL-2)-activated NK (LAK) cells against neuroblastoma target cells. Incubation of the neuroblastoma cells with IFN-gamma resulted in an increased susceptibility of these target cells to NK and LAK cells. Additionally, the IFN-gamma-treated neuroblastoma cells showed an increased susceptibility to the antibody-dependent cellular cytotoxicity (ADCC). In patients who have been treated with continuous infusions of IL-2, IL-2-induced secretion of IFN-gamma was detected by measuring the elevation of the 2-5 A synthetase activity in peripheral mononuclear cells or the 2-5 A oligoadenylates in the serum, although IFN-gamma itself was not detectable. From these results we conclude that IFN-gamma may play an important role in the immunotherapy of neuroblastoma in combination with IL-2 and/or with monoclonal antibodies.

Antibody-Dependent Cell Cytotoxicity↗
PubMedDate not supplied

[Effect of lithium on the proliferation of fibroblasts and tumor cell lines in vitro].

The specificity of a proliferation-inducing effect of lithium was investigated. Cell lines of embryonal and adult solid tumors as well as fibroblasts were cultured in lithium-concentrations ranging from 0,5 to 5,0 mmol/l. Neuroblastoma-cell-lines SK-N-SH, SK-N-LO, IMR 32, osteosarcoma-cell-line SAOS 2, melanoma-line IgR 3 and a fibroblast-line were used in this study. Cell proliferation was measured with a 3H-TdR-incorporation-assay and a tumor-stem cell-assay, the fibroblast-proliferation was measured following growth as monolayer respectively. No stimulation of proliferation was observed. The data of this in vitro study are basic for the clinical evaluation of the benefit of lithium in attenuation of chemotherapy induced leukopenia in patients with solid tumors.

Cell Division↗