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Biomedical subjects

L M Neckers

Publications and source records attributed to L M Neckers.

At least 91 records · Page 5Linked to original sources
PubMed1985

Phorbol ester enhances deoxynucleoside incorporation while inhibiting proliferation of K-562 cells.

Recently a new technique has been developed to study bromodeoxyuridine incorporation into DNA as a function of the cell cycle phase distribution. We have made the observation that treatment of the hematopoietic cell line K-562 results in a complete and immediate cessation of cell growth, although tritiated thymidine incorporation is markedly elevated for as long as 5 days. We have made use of that newly described technique to study this phenomenon.

Bromodeoxyuridine↗
PubMed1985

Transferrin receptor induction is required for human B-lymphocyte activation but not for immunoglobulin secretion.

Transferrin receptors are expressed on proliferating cells and are required for their growth. Transferrin receptors can be detected after, but not before, mitogenic stimulation of normal peripheral blood T and B cells. In the experiments reported here we have examined the regulation of transferrin receptor expression on activated human B cells and whether or not these receptors are necessary for activation to occur. Activation was assessed by studying both proliferation and immunoglobulin secretion. We have determined that transferrin receptor expression on B cells is regulated by a factor contained in supernatants of mitogen-stimulated T cells (probably B-cell growth factor). This expression is required for proliferation to occur, since antibody to transferrin receptor (42/6) blocks B-cell proliferation. Induction of immunoglobulin secretion, however, although dependent on PHA-treated T-cell supernatant, is not dependent on transferrin receptor expression and can occur in mitogen-stimulated cells whose proliferation has been blocked by antitransferrin receptor antibody. In addition, we have demonstrated that IgM messenger RNA induction following mitogen stimulation is unaffected by antitransferrin receptor antibody. These findings support a model for B-cell activation in which mitogen (or antigen) delivers two concurrent but distinct signals to B cells: one, dependent on B-cell growth factor and transferrin receptor expression, for proliferation, and a second, dependent on T cell-derived factors and not requiring transferrin receptors, which leads to immunoglobulin secretion.

Antibodies, Monoclonal↗
PubMed1985

Significant non-S-phase DNA synthesis visualized by flow cytometry in activated and in malignant human lymphoid cells.

The development of a monoclonal antibody to the deoxynucleoside bromodeoxyuridine (BrdU), combined with two parameter flow cytometry, has allowed us to examine large numbers of cells for non-S-phase DNA synthesis. Three human lymphoid cell populations were studied to determine the level of deoxynucleoside (dN) incorporation as a function of DNA content. In each population, non-S-phase DNA synthesis was observed. In a rapidly growing human T-lymphoblastoid cell line (CCRF-CEM), 53% of dN incorporation occurred in G0/G1 plus G2 + M. In chronic lymphocytic leukemia (CLL) cells stimulated with tetradecanoylphorbol acetate (TPA), 45% of the observed burst in thymidine incorporation was found to be localized to G0/G1 cells. Non-S-phase incorporation was not, however, limited to neoplastic cells. Normal human peripheral blood B cells treated with the Cowan strain of Staphylococcus aureus (CSA) undergo a transient burst in thymidine incorporation, but do not go on to divide in the absence of other stimuli. Flow-cytometric analysis showed that 80% of this CSA-stimulated dN incorporation was into G0/G1 cells. These data are consistent with a more dynamic state of DNA synthesis than usually envisioned. Furthermore, the data show that although thymidine incorporation levels are related to incorporation of dN into DNA, they can be unrelated to cell proliferation.

Antibodies, Monoclonal↗
PubMed1985

Induction of immunoglobulin secretion in follicular non-Hodgkin's lymphomas: role of immunoregulatory T cells.

B cell neoplasms are clonal expansions of B lymphocytes thought to be frozen at various points along the normal B cell differentiation pathway. We studied cell suspensions from lymph nodes involved by follicular (nodular) non-Hodgkin's lymphoma to determine the capacity of the malignant B cells to secrete immunoglobulin (Ig). Neoplastic B cells from all 14 follicular lymphomas secreted monoclonal immunoglobulin in culture when appropriate signals were provided. In most cases, maximal Ig secretion occurred when autologous T cells were removed by E rosette depletion, replaced with allogeneic normal T cells, and the cultures were exposed to 12-O-tetradecanoylphorbol-13-acetate. Autologous T cells exerted a suppressor effect on Ig secretion in 8/8 cases studied, diminishing the response of the malignant B cells to allogeneic T cells. This suppressor effect did not correlate with the percentage of cells staining with anti-Leu-2a or with "helper-suppressor" (Leu-3a-Leu-2a) ratios of the lymph node T cells. Our findings demonstrate that the arrested differentiation of most follicular lymphomas is reversible and implicate a T cell-mediated host immunoregulatory mechanism affecting Ig secretion in vivo. An additional contribution of our results is the demonstration of a cell culture system for synthesis of sufficient monoclonal Ig for use as an immunogen in production of anti-idiotype antibodies.

Antibody Formation↗
PubMed1984

In vitro enhancement of immunoglobulin gene expression in chronic lymphocytic leukemia.

B cell chronic lymphocytic leukemia (CLL) cells appear to be arrested in their differentiation so that little immunoglobulin is secreted in most cases. To determine their capacity for further differentiation we stimulated cells from a series of 10 cases of CLL with a phorbol ester and assayed for production of immunoglobulin protein, accumulation of immunoglobulin mRNA, and alterations in cell surface markers. We found that cells from all cases were induced to secret monoclonal immunoglobulin of the same heavy and light chain type as the surface membrane immunoglobulin type. Immunoglobulin secretion was preceded by a rapid increase in the levels of mRNA coding for IgM, predominantly the secretory form, mu s-mRNA, rather than the membrane form, mu m-mRNA. A similar selection of mu s- over mu m-mRNA is known to occur in plasma cells by a mechanism of differential processing of mRNA from a single mu-chain gene. Except for a decline in the expression of surface IgD, cell surface determinants remained unaffected both in terms of the percentage of positive cells and the relative number of sites per cell. In contrast to previous studies, these results indicate that CLL cells consistently retain the capacity to further differentiate toward plasma cells and secrete immunoglobulin. The immunoglobulin secretion is mediated, at least in part, by a developmentally regulated increment in mu s-mRNA.

B-Lymphocytes↗
PubMed1984

The role of the transferrin receptor in human B lymphocyte activation.

Transferrin receptors are expressed on proliferating cells and are required for their growth. Transferrin receptors can be detected after, but not before, mitogenic stimulation of normal peripheral blood T and B cells. T cells demonstrate a functional requirement for transferrin receptors in the activation process. These receptors, in turn, are induced to appear by T cell growth factor (interleukin 2). In the experiments reported here, we examined the regulation of transferrin receptor expression on activated human B cells and whether these receptors are necessary for activation to occur. Activation was assessed by studying both proliferation and immunoglobulin secretion. We determined that transferrin receptor expression on B cells is regulated by a factor contained in supernatants of mitogen-stimulated T cells (probably B cell growth factor). This expression is required for proliferation to occur, because antibody to transferrin receptor (42/6) blocks B cell proliferation. Induction of immunoglobulin secretion, however, although dependent on phytohemagglutinin-treated T cell supernatant, is not dependent on transferrin receptor expression and can occur in mitogen-stimulated cells whose proliferation has been blocked by anti-transferrin receptor antibody. These findings support a model for B cell activation in which mitogen (or antigen) delivers two concurrent but distinct signals to B cells: one, dependent on B cell growth factor and transferrin receptor expression, for proliferation; and a second, dependent on T cell-derived factors and not requiring transferrin receptors, which leads to immunoglobulin secretion.

Antibodies, Monoclonal↗
PubMed1984

Low-grade lymphomas. Expression of developmentally regulated B-cell antigens.

A series of low-grade B-cell lymphomas was analyzed for a battery of immunologic determinants by flow cytometry and immunohistochemistry. Histologically distinctive subclasses of these lymphomas, well-differentiated lymphocytic (WDL), intermediately differentiated lymphocytic (IDL), and follicular center cell (FCC) lymphoma, were found to be readily distinguishable by their expression of immunologic determinants that are known to be developmentally regulated in normal B cells. Although all cases expressed monoclonal surface immunoglobulin (sIg), HLA-DR, and the surface membrane proteins recognized by antibodies B1 (p32) and BA1, staining with other monoclonal antibodies revealed unique immunologic phenotypes for each subclass: WDL p65 (Leu 1)+, p24 (BA2)-; IDL p65+, p24+; FCC p65-, p24-. Additionally, the fluorescence intensities (number of determinants per cell) obtained for sIg, BA-1, and B1, but not HLA-DR, were significantly different among the three lymphoma subclasses. The relative fluorescence intensities of each of these three markers followed the same pattern: FCC greater than IDL greater than WDL. Taken together, these distinguishing features suggest that low-grade B-cell lymphomas represent arrested, and possibly sequential, stages of B-cell differentiation.

Antibodies, Monoclonal↗
PubMed1984

Suppression of immunoglobulin synthesis by lymphocyte subpopulations in patients with Crohn's disease.

In previous studies, patients with mild or inactive Crohn's disease were found to have increased suppressor T-cell activity. To further characterize suppressor T cells in Crohn's disease, studies were carried out with the use of monoclonal antibodies. Excessive suppressor activity was eliminated by removal of OKT 8+ lymphocytes by complement-mediated lysis. However, the percentage of OKT 8+ (or Leu 2a+) cells and the ratio of OKT 4+ to OKT 8+ (or Leu 3a+ to Leu 2a+) cells were not significantly different from normal. Although the subgroup of patients with increased suppression of immunoglobulin synthesis had a significantly lower mean Leu 3a to Leu 2a ratio than that of normal subjects, in the whole group of Crohn's patients studied, neither the percentage of Leu 2a+ cells nor the ratio of Leu 3a+ to Leu 2a+ cells correlated with excessive suppression of immunoglobulin synthesis. A subpopulation of Leu 2a+ lymphocytes reactive with the monoclonal antibody HNK-1 (Leu 2a+ HNK-1+) was increased in patients with Crohn's disease. Furthermore, elimination of HNK-1-reactive lymphocytes by complement-mediated lysis diminished the excessive suppressor cell function in patients with Crohn's disease. The percentage of Leu 2a+ HNK-1+ lymphocytes correlated significantly with the suppression of pokeweed mitogen-stimulated immunoglobulin synthesis in vitro. Thus, patients with mild Crohn's disease have an increased suppressor cell activity in vitro which correlates with the presence of a subset of lymphocytes that have an HNK-1+ Leu-2a+ phenotype.

Adolescent↗
PubMed1984

Protein synthesis but not DNA synthesis is required for tumor cell invasion in vitro.

The effects of protein and DNA synthesis inhibitors on in vitro tumor cell invasion was studied in a quantitative invasion system using human amnion. Highly invasive M5076 tumor cells were incubated on the basement membrane (BM) of the amnion in the presence of cycloheximide, aphidicolin or sodium butyrate. Tumor cell penetration through the entire thickness of the amnion was measured after 24 h. Protein synthesis in the M5076 cells was inhibited 95% within 1 h by 10(-6)M cycloheximide while DNA synthesis was unaffected. Treatment with cycloheximide reduced the number of spontaneously invading cells by 82% and by 66% in the presence of the chemoattractant, formylmethyl leucine-phenylalanine (FMLP), known to stimulate invasiveness. DNA synthesis in the M5076 tumor cells was selectively inhibited with aphidicolin (10 micrograms/ml). Although the DNA synthesis was greatly reduced, no significant effect on invasiveness with or without FMLP was observed. A less specific inhibitor of DNA synthesis, sodium butyrate (1 mM), arrested cell proliferation in the G1 phase of the cell cycle. When the butyrate-treated M5076 cells were tested in the invasion assay, a 50% increase in the number of invasive cells was observed. All three inhibitors were used in concentrations that did not affect cell viability or cell attachment to the amnion. These data indicate that protein synthesis but not DNA synthesis and not cell proliferation are required for tumor cells to invade native connective tissue barriers in vitro.

Amnion↗
PubMed1983

Lymphokine-activated killer cell phenomenon. III. Evidence that IL-2 is sufficient for direct activation of peripheral blood lymphocytes into lymphokine-activated killer cells.

Purified interleukin 2 (IL-2) was found to be sufficient for direct activation of peripheral blood lymphocytes into lymphokine-activated killer (LAK) cells. The LAK activation factor was directly and consistently associated with IL-2 activity using classic protein purification techniques, adsorption to IL-2-dependent cell lines, and inhibition with anti-Tac antibody. As yet, no other cytokines have been found that perform the same role.

Antibodies, Monoclonal↗
PubMed1983

Polymorphonuclear neutrophils express the common acute lymphoblastic leukemia antigen.

Monoclonal antibodies J5, VIL-A1, and BA-3, known to react with the common acute lymphoblastic leukemia antigen (CALLA) were found to specifically stain normal human polymorphonuclear neutrophils (PMN). The antigen detected on PMN had a molecular weight (95,000-110,000 mol wt) close to that of CALLA (95,000-100,000 mol wt) and thus these surface membrane antigens are likely related, if not identical. The fluorescent staining intensity of PMN is comparable to that of CALLA-positive leukemic cells and the presence of PMN in patient samples could potentially produce false-positive results in diagnosis.

Adult↗
PubMed1983

Plasma phenylalanine, tyrosine, and tryptophan in schizophrenia.

Plasma phenylalanine, tyrosine, and tryptophan concentrations were measured in chronic schizophrenic patients, normal controls, and heterozygotes for phenylketonuria. Schizophrenic patients' plasma concentrations of these amino acids could not be distinguished from those of normal controls, either when fasting or following oral or intravenous (IV) phenylalanine challenge. No neuroleptic effect was observed. Plasma phenylalanine-tyrosine ratios following IV phenylalanine challenge could easily distinguish heterozygotes from schizophrenic and normal control subjects but could not distinguish schizophrenic subjects from normal control subjects. No overlap between heterozygotes' values and those of the schizophrenic and normal subjects was observed. These studies find no evidence of abnormal phenylalanine metabolism in schizophrenic persons. Phenylalanine challenge did not affect the abstraction or judgment capacities of the subjects.

Administration, Oral↗
PubMed1983

Transferrin receptor induction in mitogen-stimulated human T lymphocytes is required for DNA synthesis and cell division and is regulated by interleukin 2.

Transferrin is required by many cells for growth. Mitogen-induced T lymphocyte proliferation is dependent on the presence of both interleukin 2 (IL-2; T-cell growth factor) and transferrin, even though resting lymphocytes do not have receptors for either. Exposure to mitogen (phytohemagglutinin) alone is sufficient to induce transient appearance of IL-2 receptors on lymphocytes. Using monoclonal antibodies to the IL-2 receptor and to the transferrin receptor, we examined those signals required for transferrin receptor induction during T lymphocyte proliferation. Our study has revealed that (i) monocytes, or a monocyte substitute such as the phorbol ester tetradecanoylphorbol 13-acetate, are required for transferrin receptor expression after mitogen exposure; (ii) the presence of IL-2 receptors is necessary for transferrin receptor induction; (iii) antibody to the IL-2 receptor inhibits thymidine incorporation (DNA synthesis) in lymphocytes, but only if administered before transferrin receptors have appeared; and (iv) antitransferrin receptor antibody inhibits DNA synthesis but has minimal effect on IL-2 receptor expression. Thus, IL-2 receptor induction leads to transferrin receptor induction and subsequent initiation of DNA synthesis. These data indicate that IL-2 stimulates T lymphocyte proliferation, at least in part, by induction of transferrin receptors on these cells.

Antibodies, Monoclonal↗
PubMed1983

Rearrangement and expression of immunoglobulin genes and expression of Tac antigen in hairy cell leukemia.

The origin and exact stage of differentiation of the neoplastic cells that comprise hairy cell leukemia have remained uncertain. As Ig heavy and light chain genes must both undergo a DNA rearrangement during B-cell development but rarely do so within other hematopoietic lineages, we examined these genes in this leukemia. The neoplastic cells of all eight cases demonstrated rearranged heavy and light chain genes and, in two cases examined, contained the corresponding mRNA for heavy and light chain Ig. Consistent with this B-cell genotype, all cases displayed cell surface HLA-DR and B-cell-associated antigens. Unexpectedly, all cases demonstrated cell surface Tac antigen, which previously had been restricted predominantly to select T-cell malignancies and activated T cells. Prior studies suggested that the anti-Tac monoclonal antibody recognized a peptide associated with the binding of interleukin 2 (T-cell growth factor) in such T cells. Immunoprecipitation with anti-Tac and NaDodSO4/polyacrylamide gel electrophoresis revealed an antigen on leukemic hairy cells with a Mr of 53,000-57,000, identical in size to the receptor on activated T cells. This apparent biphenotypic status might reflect a transformation-associated expression of the Tac antigen in this leukemia. Alternatively, hairy cell leukemia may be a malignancy of a unique stage of normal B-cell differentiation in which the Tac antigen is expressed.

Antibodies, Monoclonal↗
PubMed1982

NAD glycohydrolase: enzyme characterization using intact mammalian erythrocytes.

1. NADase activity has been determined on intact erythrocytes of several species. 2. Although a wide range in maximum velocity exists across species, Michaelis constants observed are very similar. 3. The enzyme is found on the outer surface of the erythrocyte plasma membrane. 4. It is inhibited by substrate after an apparent permanent modification. 5. This modification may be due to self ADP-ribosylation. 6. We have also demonstrated the presence of an ADP-ribosyltransferase on the outer surface of the sheep erythrocyte membrane.

Animals↗