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G Böck

Publications and source records attributed to G Böck.

At least 73 records · Page 4Linked to original sources
PubMed1992

Modulation of tumour marker CA-125 expression in cultured ovarian carcinoma cells.

The aim of this study was to elucidate whether proliferation of ovarian carcinoma cells may affect the biosynthesis and release of CA-125. In a cell culture model the tumour marker CA-125 expression in cytosol, surface membrane, and release into culture medium was studied in six human carcinoma cell lines. Cell cycle analysis of propidium iodide stained nuclei was performed using a fluorescent activated cell sorter. The turnover of CA-125 is very rapid, within 24 h the equivalent amount found in each cell was also released in the supernatant culture medium. A good relation between cytosolic, membrane, and released CA-125 was observed. CA-125 expression was associated predominantly with the G0/G1 phase of the cell cycle and was dependent on cell density. The results presented here demonstrate that factors associated with tumour cell proliferation could influence the CA-125 serum level in ovarian cancer patients.

Antigens, Surface↗
PubMed1992

HIV-1 gp41 contains two sites for interaction with several proteins on the helper T-lymphoid cell line, H9.

OBJECTIVE: To characterize putative binding sites for HIV-1 gp41 to H9 cells. DESIGN: Based on accumulating evidence in the literature that HIV-1 can bind to cell surfaces independent of CD4, we attempted to clarify whether gp41, the transmembrane HIV-1 protein, contributes to CD4-independent binding. We therefore tested binding of gp41 to cells. METHODS: Using fluorescence-activated cell-sorter analysis, we examined the binding of recombinant gp160 (gp160) and soluble gp41 (sgp41; Env amino acids 539-684) to H9 cells, and located the putative binding site(s) of gp41 by inhibition using 16 HIV-1 Env peptides. Putative HIV-1 receptor proteins in H9 cell lysates were Western blotted and precipitated using sgp41. RESULTS: sgp41 bound to the CD4+ H9 cells and rgp160 bound to H9 cells independent of gp120-binding sites on CD4 molecules. Two gp41 peptides (Env amino acids 591-605 and 651-665) inhibited the binding of sgp41 to H9 cells. Four bands, of 37, 40, 55 and 97 kD, were blotted in H9 cell lysates, and three bands, 40, 97 and 108 kD, were observed in the precipitation analysis using lysates of 125I-surface-labelled H9 cells and sgp41 attached to sepharose beads. CONCLUSIONS: HIV-1 gp41 contains two putative binding sites to H9 cells. These sites may be located within Env amino acids 591-605 (ERYLKDQQLLGIWGC) and 651-665 (TLLEESQNQQEKNEQ). Using two different techniques, five proteins (37, 40, 55, 97 and 108 kD) were identified in H9 lysates as possible candidates for gp41 binding.

Amino Acid Sequence↗
PubMed1992

Frequency of chicken CD4+ and CD8+ cells. Genetic control and effect of Rous sarcoma virus infection.

In chickens from congenic inbred lines CB and CC that differ only in the major histocompatibility complex (MHC), we observed significantly different percentages of CD4+ and CD8+ cells in peripheral blood lymphocytes (PBL) and spleen. Positive cells were detected by indirect immunofluorescence test as analysed by flow cytometry. In both PBL and spleen cell suspensions, the number of CD4+ cells was significantly higher in CB than in CC chickens, whereas in CC birds there was a higher percentage of CD8+ cells than in CB. These statistically significant differences were under the MHC control. We found no statistically significant influence of regressions or progression of Rous sarcoma virus-induced tumours on the percentage of peripheral T cells and on the interleukin-2 production in vitro.

Animals↗
PubMed1992

High amount of epsilon-(gamma-glutamyl)lysine cross-links in Mallory bodies.

Alcoholic hepatitis, the most severe form of alcoholic liver disease, is associated with inflammation, liver cell necrosis, and the appearance of Mallory bodies (MBs) in hepatocytes. Identical MBs can be experimentally induced in mouse livers by chronic griseofulvin or 3,5-diethoxycarbonyl-1,4-dihydrocollidine treatment. MBs are filamentous cytoplasmic inclusions containing insoluble high molecular weight protein material. Covalent polymerization of intracellular proteins may occur through formation of epsilon-(gamma-glutamyl)lysine cross-links catalyzed by Ca(2+)-dependent transglutaminases. Therefore, isolated experimentally-induced MBs were analyzed for the presence of epsilon-(gamma-glutamyl)lysine bonds. Highly purified MBs contained 19.7 nmole (griseofulvin-induced) and 14.4 nmoles (3,5-diethoxycarbonyl-1,4-dihydrocollidine induced) of isodipeptide linkage, respectively, per mg of protein. These results suggest that transglutaminase-induced cross-linking of proteins plays a major role in MB formation.

Animals↗
PubMed1991

Correlation of lymphocyte lipid composition membrane microviscosity and mitogen response in the aged.

Healthy aged and young blood donors were investigated for the role of membrane lipid composition in the age-related increase in membrane microviscosity and decline of mitogen responsiveness. Membrane microviscosity was shown to correlate positively with membrane cholesterol/phospholipid molar ratios, which were significantly elevated in the elderly. A positive correlation also was confirmed between lymphocyte membrane microviscosity, which was measured using the probe 1,6-diphenyl 1,3,5-hexatriene, and phytohemagglutinin responsiveness of cells from the same donor. Using stepwise regression statistical analysis, the variables age, cholesterol, cholesterol/total phospholipid and phosphatidyl ethanolamine/phosphatidyl choline molar ratios were all shown to have a significant positive influence on membrane microviscosity, whereas total phospholipids had a negative effect. No statistically significant difference was seen in content of any single saturated or unsaturated fatty acid between young and old donors. After pooling, however, the proportion of all unsaturated fatty acids was significantly higher in cells from the elderly as a consequence of an increase of n-6 and n-3 polyunsaturated fatty acids. Changes in lipid composition and physical properties of lymphocyte plasma membranes may, therefore, be responsible (at least partially) for the diminution of immune reactivity in old age.

Adult↗
PubMed1991

Chicken major histocompatibility complex congenic lines differ in the percentages of lymphocytes bearing CD4 and CD8 antigens.

In the experiments to be described two congenic inbred lines CB and CC and two recombinant lines CB.R1 and CC.R1 were used. All four lines differ only in regard to the major histocompatibility complex (MHC). To determine the percentage distributions of the two cell subsets in peripheral blood lymphocytes (PBL) in these lines, monoclonal antibodies to these two antigens were used. By FACScan there were more CD4+PBL in CB and CB.R1 lines (share B-F/B-L region, controlling class I/class II antigens with line CB) than CC and CC.R1, while the reverse was true with CD8+ subsets. There were more CD8+ PBL in the CC and CC.R1 lines and less in CB and CB.R1 lines. The ratio of CD4+ to CD8+ in CB chickens was 3.4 +/- 0.2 and in CC chickens 1.6 +/- 0.1.

Animals↗
PubMed1991

High molecular weight components are main constituents of Mallory bodies isolated with a fluorescence activated cell sorter.

Mallory bodies (MBs) are cytoplasmic filamentous aggregates containing cytokeratin (CK) material. They occur in hepatocytes of patients with alcoholic liver disease (i.e., alcoholic hepatitis) and can also be induced experimentally in mice by chronic griseofulvin intoxication. To further investigate components and mechanisms involved in MB formation, a new method for MB purification was established. MBs present in a liver homogenate of griseofulvin-fed mice were labeled with a murine monoclonal antibody specific for MBs and a second fluorescein isothiocyanate-conjugated (anti-mouse IgG and IgM) antibody and subsequently isolated by two sequential sorting procedures using a fluorescence activated cell sorter (FACS). Purity of MB isolates was over 90% as revealed by computer analysis of sorting signals and fluorescence and electron microscopy. Electrophoretic separation on sodium dodecyl sulfate-polyacrylamide gels revealed three MB-related polypeptides with apparent molecular masses of 48, 55, and 65 kilodaltons but most of the highly purified MB material did not enter the gel or remained at the interphase between stacking and resolving gels. Western blotting with CK-specific antibodies showed the presence of CK epitopes in the high molecular weight MB material, which has a similar amino acid composition as normal liver CKs. These results establish that very high molecular weight material is the main constituent of MBs and suggest that a post-translational modification of CKs by covalent crosslinks is a principal mechanism of MB pathogenesis.

Amino Acids↗
PubMed1990

Glucocorticoid-regulated gene expression in the immune system. Analysis of glucocorticoid-regulated transcripts from the mouse macrophage-like cell line P388D1.

To further elucidate the molecular mechanisms underlying glucocorticoid-mediated immune suppression, we have exploited cDNA cloning and subtractive screening methods to identify glucocorticoid-regulated transcripts in the mouse macrophage-like cell line, P388D1. Two of the three isolated glucocorticoid-regulated mRNA species corresponded to genes potentially important to immunoregulation: one glucocorticoid-suppressed mRNA species probably encoded the previously uncloned 3-hydroxy-3-methylglutaryl coenzyme A reductase, an enzyme that appears important for in vitro immune responses. The other mRNA species showed glucocorticoid-increased mRNA steady-state levels and was transcribed from an endogenous ecotropic type C retroviral locus. This transcript gives rise to a protein (transmembrane retroviral protein, formerly p15E), which, along with its feline and human homologs, has been implicated in immunosuppression caused by mouse, cat, and human retroviruses. Our results raise the possibility that the immunosuppressive activity of glucocorticoids might be mediated, in part, by regulating the expression of the above immunoregulatory proteins.

Animals↗
PubMed1990

Tumor necrosis factor alpha maintains the viability of murine epidermal Langerhans cells in culture, but in contrast to granulocyte/macrophage colony-stimulating factor, without inducing their functional maturation.

Freshly isolated murine epidermal Langerhans cells (LC) are weak stimulators of resting T cells but increase their stimulatory capacity 10-30-fold upon 2-3 d of culture together with other epidermal cells. This maturation of LC is mediated by two keratinocyte products. Granulocyte/macrophage colony-stimulating factor (GM-CSF) maintains viability and increases function. IL-1 alone does not keep LC alive, but when combined with GM-CSF further enhances their stimulatory activity. We have now searched for a cytokine that would keep LC in a viable, but functionally immature state. When LC (enriched to greater than 75%) were cultured in the presence of GM-CSF (2 ng/ml) or murine (TNF-alpha) (plateau effect at 62 U/ml), the recovery of viable LC after 72 h was identical. The LC cultured in murine TNF-alpha, however, were 10-30 times less active in stimulating resting T cells. A series of experiments demonstrated that this phenomenon was not due to the induction of insufficient amounts of GM-CSF, the induction of a suppressor factor, or a toxic effect of TNF-alpha. Interestingly, the observed TNF-alpha activity exhibited a species preference, as human TNF-alpha was not active at comparable doses. We have observed an unexpected effect of TNF-alpha on LC in vitro. Though we found that freshly prepared epidermal cells express TNF-alpha mRNA, further studies are needed to establish whether TNF-alpha plays a role in vivo by keeping resident LC in a viable, but functionally immature state.

Animals↗
PubMed1990

Increased expression of high-affinity low-density lipoprotein receptors on human T-blasts.

Like all cells, lymphocytes need cholesterol for proper function, a requirement met by a finely tuned homeostasis between intracellular synthesis and uptake from the environment via low-density lipoproteins (LDL). We used flow cytometry to analyze the receptor activity of resting cells and T blasts incubated/activated in serum-free culture medium, or in medium supplemented with 25-5,000 micrograms/ml LDL. Dioctadecyl-indocarbocyanine has proved to be a useful fluorescent probe for investigating the LDL receptor activity of lymphocytes. The results show the receptor activity of day-3 resting T cells to be reduced more than 50% by 50 microgram LDL/ml, whereas 100-fold higher concentrations are necessary to achieve the same level of reduction in day-3 PHA blasts. The LDL receptor activities of individual blood donors' resting T cells, in vitro cholesterol-deprived resting T cells, and activated T blasts, were compared using two analytical techniques: spectrofluorometric analysis of detergent-solubilized cell suspensions and flow cytometric analysis of single living cells. Receptor affinity was determined by Scatchard analysis of spectrofluorometric binding curves, and by Line-weaver-Burke plots of flow cytometric data. Both methods yielded essentially identical dissociation constants (Kd) for cholesterol-deprived resting T cells and mitogen-activated T blasts, which fell in the expected range for the high-affinity LDL receptor (4.1-8.9 nM). In addition, spectrofluorometric analysis, but not flow cytometry, permitted quantification of LDL uptake.(ABSTRACT TRUNCATED AT 250 WORDS)

Adult↗
PubMed1989

Modulation of ovarian carcinoma tumor marker CA-125 by gamma-interferon.

Interferons are known to modulate several cellular functions by the induction of different proteins. In our study a gamma-interferon-mediated presentation of one of the most important ovarian tumor markers (CA-125) on the cell surface was demonstrated using two ovarian carcinoma cell lines in vitro (HTB-77 and SKOV-3). This induction was found to be dependent on an intact protein biosynthesis. The gamma-interferon effect reached a maximum on the third day of treatment. Under such conditions the CA-125 concentration was increased intracellularly, on the cell surface, and in the supernatant culture medium. The surface antigen was shed rapidly with a half-life of 1 day. The addition of dexamethasone to gamma-interferon treated HTB-77 cells improved CA-125 expression synergistically. HLA-DR and CA-125 expression was found to be regulated by interferons in different ways. The demonstration of CA-125 expression provides an important insight into the potential regulatory mechanism governing this tumor marker. Since interferons are naturally occurring substances as well as therapeutically administered agents, it seems necessary to pay attention to possible tumor marker modulation.

Antigens, Tumor-Associated, Carbohydrate↗
PubMed1989

Promotion of lymphocyte growth by high density lipoproteins (HDL). Physiological significance of the HDL binding site.

The characteristics and physiological relevance of the high density lipoprotein (HDL) binding site on unstimulated and mitogen activated human peripheral blood lymphocytes have been investigated. At 37 degrees C, specific binding/uptake of fluorescent (dioctadecylin-docarbocyanine, DiI) HDL was observed by cells from healthy donors as well as by those from low density lipoprotein receptor-defective patients; mitogen activated T-blasts exhibited a markedly elevated DiI-HDL uptake compared to resting T-cells. Binding was saturable at 37 degrees C and of high affinity, with a Kd of 5 x 10(-8) M. It was blocked by anti-apoAI polyclonal antibodies (F(ab)2 fraction), but not by anti-apolipoprotein (apo)E, anti-apoAII, or anti-apoB, and was inhibited competitively by HDL apoproteins and an apoAI-protein A fusion protein. T-cell associated DiI-HDL was increased by trypsin treatment (of the cells) and decreased by activation in the presence of HDL or low density lipoprotein. Comparison of the concentration dependencies of growth promotion and specific cell association of HDL indicated that two mechanisms of lipid exchange may be in operation: one a binding-dependent mechanism of cholesterol exchange, with maximal effect in the HDL concentration range (20-200 micrograms/ml) in which specific binding increases rapidly, and the other a binding-independent exchange of lipids effective at concentrations in which specific binding is saturated (300-5000 micrograms/ml).

Adult↗
PubMed1989

Expression of 7F7-antigen, a human adhesion molecule identical to intercellular adhesion molecule-1 (ICAM-1) in human carcinomas and their stromal fibroblasts.

The 7F7-antigen is a widespread 85-kDa membrane adherence molecule involved in heterotypic adhesion between PHA-blasts and fibroblasts. Immunofluorescence analysis of COS cells transfected with clone pICAM-I indicated that 7F7-antigen is identical with ICAM-I, the ligand for the contact molecule LFA-I. We have investigated the expression of ICAM-I on several human carcinomas. Tumor cells in most carcinomas, with the exception of squamous-cell carcinomas, expressed very little ICAM-I or none at all. In contrast, marked expression of this molecule was observed on fibrous tissue in the vicinity of carcinoma cells, its intensity correlating with lymphatic infiltration in these tumors. We also examined the expression of ICAM-I on carcinoma cell lines and its induction by treatment with interferon-gamma (IFN-gamma). The inducibility of ICAM-I expression on cultured fibroblasts by several lymphokines suggests that the expression of ICAM-I in the vicinity of carcinoma cells is effected by lymphokines produced by activated lymphocytes/macrophages within the tumor.

Antigens, Surface↗
PubMed1989

An orally administered bacterial immunomodulator primes rabbit neutrophils for increased oxidative burst in response to opsonized zymosan.

To assess the potential effect of an orally administered immunomodulator, consisting of a lysate of seven different bacteria, on polymorphonuclear leukocyte (PMN) function, rabbits were fed this preparation for five consecutive days via a gastric tube. On day 6, PMN were separated from peripheral blood and oxidative burst was triggered by opsonized zymosan or 12-O-tetradecanoylphorbol-13-acetate and quantitated on a single-cell basis. This study presents the extension of an existing flow cytometric method, leading to the possibility of quantitating single-cell oxidative burst triggered by particulate (instead of only soluble) stimuli. By this means, treated animals showed statistically significant increased oxidative burst reactions compared with the control group. The data provide evidence that oral application of a bacterial immunomodulator leads to a primed state in PMN for increased oxidative activity in response to a particulate stimulus. This offers the possibility that the beneficial effect of similar treatment in humans may in part be due to comparable mechanisms.

Adjuvants, Immunologic↗
PubMed1989

Use of a FACS III for fluorescence depolarization with DPH.

We have previously demonstrated age-related differences in human lymphocyte membrane fluidity, by use of steady-state polarization measurements on bulk cell suspensions with the fluorescence probe DPH. However, for exact analysis of the possible functional importance of these changes, single-cell measurements were deemed of interest. We have now used an analog division device to measure fluorescence depolarization "p" of DPH in real time with a FACS III flow cytometer. The measurements are reliable, as we have been able to confirm the differences in DPH "p" between monocytes and lymphocytes previously shown in bulk suspension and to demonstrate the expected differences in fluidity of lipid-modulated cells. We also found significant differences in DPH "p" between lymphocytes of young and elderly blood donors. Lymphocyte subsets did not differ in polarization values but did differ in fluorescence intensity with Th less than Ts less than B = NK cells.

Adult↗
PubMed1989

Immunodeficiency in old age.

Aging is a multi-facetted process, but the deterioration of the immune function seems to play a central role in this context. Paradoxically, immune reactivity against exogenous antigens declines during aging while autoimmune reactivity increases. One of the aims of our investigations on the function of the senescent immune system was to define immune parameters of 'normal' aging, i.e. those not dependent on underlying diseases. Specifically, our interest was focused on the possible role of an altered lipid metabolism of cells of the immune system during aging. The known decrease of plasma membrane fluidity of lymphoid cells and monocytes in higher age may be one of the factors responsible for the nonoptimal functioning of the immune system. This property, in turn, seems to be based on an altered lipid metabolism. Specifically, we have evidence that the finely tuned balance between the transport of cholesterol to and from lymphoid cells via the environment and the intracellular cholesterol biosynthesis seems to be disturbed with increasing age. This conclusion is drawn from experiments where low-density lipoprotein (LDL) and high-density lipoprotein (HDL) receptor activity is assessed using fluorescently labeled lipoproteins in fluorescence-activated cell sorter (FACS) analyses. LDL receptor uptake is unexpectedly increased in the elderly, but LDL receptor regulation, and serum LDL composition itself seem to be normal. Preliminary data point out the possibility that the efflux of cholesterol via HDL may be insufficient. Attempts to modulate plasma membrane fluidity by means of the phospholipid mixture 'active lipid 721' (AL 721) showed that this drug, in contrast to literature reports, is not a 'membrane fluidizer' but rather exerts this effect as a nutrient for lymphocytes and monocytes.

Aging↗