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Biomedical subjects

D Berkovic

Publications and source records attributed to D Berkovic.

25 records · Page 2Linked to original sources
PubMed1991

Tumor necrosis factor induces rapid production of 1'2'diacylglycerol by a phosphatidylcholine-specific phospholipase C.

Tumor necrosis factor (TNF) is a proinflammatory polypeptide that is able to induce a great diversity of cellular responses via modulating the expression of a number of different genes. One major pathway by which TNF receptors communicate signals from the membrane to the cell nucleus involves protein kinase C (PKC). In the present study, we have addressed the molecular mechanism of TNF-induced PKC activation. To this, membrane lipids of the human histiocytic cell line U937 were labeled by incubation with various radioactive precursors, and TNF-induced changes in phospholipid, neutral lipid, and water-soluble metabolites were analyzed by thin layer chromatography. TNF treatment of U937 cells resulted in a rapid and transient increase of 1'2'diacylglycerol (DAG), a well-known activator of PKC. The increase in DAG was detectable as early as 15 s after TNF treatment and peaked at 60 s. DAG increments were most pronounced (approximately 360% of basal levels) when cells were preincubated with [14C]lysophosphatidylcholine, which was predominantly incorporated into the phosphatidylcholine (PC) pool of the plasma-membranes. Further extensive examination of changes in metabolically labeled phospholipids indicated that TNF-stimulated hydrolysis of PC is accompanied by the generation of phosphorylcholine and DAG. These results suggest the operation of a PC-specific phospholipase C. Since no changes in phosphatidic acid (PA) and choline were observed and the production of DAG by TNF could not be blocked by either propranolol or ethanol, a combined activation of phospholipase D and PA-phosphohydrolase in DAG production appears unlikely. TNF-stimulated DAG production as well as PKC activation could be blocked by the phospholipase inhibitor p-bromophenacylbromide (BPB). Since BPB did not inactivate PKC directly, these findings underscore that TNF activates PKC via formation of DAG. TNF stimulation of DAG production could be inhibited by preincubation of cells with a monoclonal anti-TNF receptor (p55-60) antibody, indicating that activation of a PC-specific phospholipase C is a TNF receptor-mediated event.

Calcium↗
PubMed1990

Effects of antineoplastic phospholipids on parameters of cell differentiation in U937 cells.

The proliferation of the human promonocytic leukemia cell line U937 is inhibited by several ether lipids, ether lipid analogues and by phorbol esters. An early effect of this retardation of cell growth is the induction of a basic chromosomal protein, histone H1(0). Northern blot analysis of H1(0) mRNA levels reveals an increase of the mRNA concentration within a few hours after addition of hexadecylphosphocholine and 1-O-octadecyl-2-O-methyl-rac-glycero-3-phosphocholine. This early effect on the synthesis of a subtype of H1 proteins precedes the expression of several parameters of the monocytic differentiation of U937 cells.

Antigens, Surface↗
PubMed1987

Specific membrane receptors for human interferon-gamma (IFN-gamma).

IFN-gamma is a T-cell derived lymphokine which possesses antitumoral activity for a variety of malignant cells by virtue of its direct effect on cell growth and by its immunomodulatory activity. All IFN-gamma actions are initiated by binding to high affinity cell surface receptors, which are constitutively expressed in virtually all cell lines from various tissues. Although the detailed structure of the IFN-gamma receptor is still elusive, the available data suggest that the high affinity IFN-gamma binding site is a heterodimeric molecule of 128 kDa comprised of two subunits of 53 and 75 kDa, which is invariantly expressed in distinct tumor cells, differing in their response to IFN-gamma. Thus, the capability and type of cellular response to IFN-gamma appears to be largely determined at a post-receptor level. Nevertheless, in sensitive cell lines, the magnitude of response is proportional to the quantity of receptor ligand interactions. This could be important for the definition of effective doses in clinical applications of IFN-gamma, as distinct tumor cells are heterogeneous with respect to quantity of IFN-gamma receptors, with greater 20-fold differences of the number of receptors per cell.

Affinity Labels↗
PubMed1986

IFN-gamma receptors on human tumor cells: relationship between receptor ligand interactions and induction of IFN-gamma response.

Scatchard analysis of 125I-IFN-gamma binding on fresh lymphoid and myeloid tumor cells derived from 34 leukemia patients and on normal cells obtained from 14 healthy individuals revealed similar high affinity binding with a mean Kd of around 2 X 10(-11) M in 14/14 normal and 30/34 malignant cells, but large quantitative differences in the receptor number of malignant cells with a range of 300 to 12,000 receptors/cell. In contrast, normal lymphoid and myeloid cells expressed consistantly low numbers of receptors with a mean of 300 and 1,000 receptors/cell, respectively. Kinetic studies of IFN-gamma binding in relation to induction of HLA-DR antigens in established cell lines revealed the existence of close correlations between the quantity of receptor ligand interaction and induction of IFN-gamma response, indicating that at limiting IFN-gamma concentrations the height of response is controlled by the number of expressed membrane receptors. In the light of the observed quantitative differences in IFN-gamma receptors on various tumors, this finding may have implications for the definition of therapeutically effective IFN-gamma doses.

Bone Marrow↗
PubMed1986

Quantitation and characterization of gamma-interferon receptors on human tumor cells.

The vast majority (71 of 77) of human tumor cells derived from various tissue origins were found to express specific membrane receptors for gamma-interferon (IFN-gamma). Six receptor-negative tumors were found among leukemic cells of lymphoid origin. Scatchard analysis with 125I-labeled human recombinant IFN-gamma revealed a similar binding affinity with a mean dissociation constant (Kd) of around 2 X 10(-11) M not only for various established cell lines, but also for leukemic and carcinoma cells derived from biopsy material. In contrast to similar KdS, large differences in the number of expressed IFN-gamma membrane receptors were found on distinct tumor cells of the same cell type ranging from a few hundred up to 2 X 10(4) for both carcinoma cells and leukemic cells. For comparison, the IFN-gamma receptor number on normal lymphocytes (mean, approximately 300/cell) and normal bone marrow cells (mean, approximately 1000/cell) was consistently found to be low. Cross-linking of membrane-bound 125I-IFN-gamma with disuccinimidyl suberate and subsequent sodium dodecyl sulfate-polyacrylamide gel electrophoresis analysis revealed, in both leukemia and carcinoma cells, three distinct complexes with molecular weights of approximately 70,000, 92,000, and 160,000, suggesting the existence of IFN-gamma receptor subunits. A dimeric structure of the functional IFN-gamma receptor with an estimated molecular weight of about 128,000 +/- 10,000 is proposed. Together with the Scatchard analysis, these data suggest the existence of a single class of high affinity IFN-gamma receptors in tumor cells of distinct tissue origin.

Cell Line↗