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Biomedical subjects

J H Phillips

Publications and source records attributed to J H Phillips.

At least 199 records · Page 11Linked to original sources
PubMed1983

Cytoskeletal proteins in cultured secretory cells from rat pituitary.

Cultured cells from female rat pituitaries were examined by immunofluorescence microscopy for the presence of pituitary hormones and for cytoskeletal proteins. The cells attach well to glass coverslips and, when well-dispersed initially, develop networks of branching processes. Their cytoplasm appears granular and shows granular fluorescence when the cells are permeabilized and treated with antisera to pituitary hormones. The cultures contain fibroblasts which are morphologically distinct and much larger than the secretory cells. Consequently, they provide an internal control for cytoskeletal proteins in nonsecretory cells. Fibroblasts have relatively prominent stress fibres. By contrast, the secretory cells have a diffuse cytoplasmic distribution of all cytoskeletal proteins investigated. One cell type, the gonadotrope, which secretes luteinizing hormone and follicle-stimulating hormone, was examined in particular detail.

Actinin↗
PubMed1983

NKP-15: a monoclonal antibody reactive against purified human natural killer cells and granulocytes.

A monoclonal antibody of the isotype IgG1,k, NKP-15, was produced by immunizing Balb/c mice with highly purified suspensions of human natural killer (NK) cells. By indirect immunofluorescence, NKP-15 reacted with 15 +/- 5.0% of the peripheral blood lymphocytes, 90 +/- 3.0% of the granulocytes and was non-reactive with T-cells, B-cells, monocytes, platelets and erythrocytes. NKP-15 reacted with 90 +/- 6.0% of purified large granular lymphocytes (LGL), which displayed highly enriched NK cell activity. NKP-15, however, showed no reactivity with a panel of human tumor cell lines. Analysis of NK cell-K562 tumor cell conjugates and depletion by panning of NKP-15+ cells indicated that all functionally tumorlytic NK cells expressed the NKP-15 antigen. It was also determined by competitive blocking experiments that NKP-15 recognizes a new NK cell surface antigen not delineated by the NK-specific monoclonal antibody, Leu-7, and that the epitope for NKP-15 appears to be associated with the Fc-receptor of NK cells and granulocytes.

Animals↗
PubMed1983

Adenine nucleotide and phosphoenolpyruvate transport by bovine chromaffin granule "ghosts".

Purified bovine adrenal chromaffin granule "ghosts" (resealed membranes) accumulate nucleotides and phosphoenolpyruvate in a temperature-dependent process. Equilibration takes several hours, the compounds being accumulated to not more than twice their concentration in the medium. This uptake occurs primarily by a non-saturable diffusion process, and rates and extents show little dependence on membrane energization. Uptake rates are inhibited less than 50% by compounds that discharge the "ghost" membrane potential, or by atractylate or low concentrations of Cibacron Blue. The data gave little support to the theory that chromaffin granules have a membrane potential-dependent adenine nucleotide transport process and that nucleotides are in equilibrium with the positive-inside potential across the granule membrane.

Adenine Nucleotides↗
PubMed1983

Visualization of the exocytosis/endocytosis secretory cycle in cultured adrenal chromaffin cells.

Cultured bovine adrenal medullary chromaffin cells were stimulated to secrete catecholamines by addition of veratridine or nicotine. The formation of an exocytotic pit exposes a major secretory granule membrane antigen, the enzyme dopamine beta-hydroxylase, to the external medium. By including antiserum to this enzyme in the medium, we were able to visualize sites of exocytosis by decoration of bound antibody using a fluorescent second antibody. Internalization of this antibody-antigen complex was then followed in chase experiments: approximately half the surface complex was internalized in 15-30 min. In other experiments, secretion was triggered in the absence of antiserum, and surface enzyme was revealed by binding antibodies at various times after secretion had been halted by an antagonist. Surface patches of antigen remained discrete from the bulk of the plasma membrane for at least 30 min, although a substantial proportion of the antigen was internalized within this time. Cell surface concanavalin A receptors were internalized at a roughly similar rate, suggesting that mechanisms may be similar. After internalization, chromaffin granule membranes fused to larger structures, possibly lysosomes, and were transported over a few hours to the perinuclear region of the cell.

Adrenal Medulla↗
PubMed1983

Subpopulations of human natural killer cells defined by expression of the Leu-7 (HNK-1) and Leu-11 (NK-15) antigens.

The functional and phenotypic characteristics of cells in human peripheral blood that mediate "natural killer" (NK) cytolysis have been examined with the use of multiparameter flow cytometry analysis and cell sorting. Essentially, all lymphocytes expressing NK and ADCC activity reacted with the anti-Leu-11a monoclonal antibody. The Leu-11a antigen was expressed on cytotoxic large granular lymphocytes (LGL), neutrophils, and basophils, but was not present on B cells, mitogen-activated T lymphoblasts, or Leu-1+ and Leu 4+ resting T cells. Anti-Leu-11a antibody selectively inhibited the binding of FITC heat-aggregated IgG complexes to granulocytes and LGL, and it may recognize a type of Fc receptor on these cells. Two-color FACS cell sorting indicated the existence of four lymphocyte subsets defined by the expression of Leu-11a and Leu-7 antigens. The Leu-11a+, -7- cells were highly active in 4-hr NK assays with the use of 51Cr-labeled K562 as the target. In contrast, the Leu-11a-, -7+ cells demonstrated weak activity and the Leu-11a-, -7- cells demonstrated no activity. The function of the Leu 11a+, -7+ cells varied considerably among several individuals examined. Multiparameter analysis with the use of two-color flow cytometry was used to determine the relationship between the expression of these NK-associated antigens and T and B cell-associated markers. These data indicate that considerable heterogeneity exists within human peripheral lymphocytes with regard to cell phenotype and function, but that several defined cellular subsets can be clearly revealed by using multiparameter FACS analysis and sorting.

Antibodies, Monoclonal↗
PubMed1982

Reduction of membrane-bound dopamine beta-hydroxylase from the cytoplasmic surface of the chromaffin-granule membrane.

Resealed bovine chromaffin-granule 'ghosts' were used for assaying the membrane-bound form of dopamine beta-hydroxylase. Hydroxylation of the substrate tyramine is dependent on its accumulation within the 'ghosts', where the active site of the enzyme is located. Free tyramine in the medium is at a low concentration, low ionic strength and a relatively high pH (7.0), so that even in the presence of a reducing agent (co-reductant) the unaccumulated amine is hydroxylated at a negligible rate. 'Ghosts' contain an endogenous co-reductant, which is shown to be catecholamine remaining in the membrane itself after granule lysis. Catecholamine that is free in solution in the medium or in the interior of the 'ghosts' is not effective as co-reductant, nor is ascorbate, in contrast with the situation with soluble dopamine beta-hydroxylase. Ferrocyanide is an active co-reductant, however, giving a hydroxylation rate approximately equal to the tyramine accumulation rate: it does not enter the 'ghosts', nor does the enzyme appear to utilize ferrocyanide sealed inside the 'ghosts'. A mechanism must therefore exist for transferring electrons across the membrane from the cytoplasmic surface to the matrix surface. NADH is not an electron donor for the enzyme, nor is cytochrome b-561 involved.

Animals↗
PubMed1981

Antitumor effect of tuftsin.

Tuftsin, a physiological tetrapeptide derived from the Fc region of leukophilic IgG possesses a variety of immunopotentiating properties including the ability to act as an immunotherapeutic agent against the experimental tumors, L1210 leukemia and Cloudman S-91 melanoma. Although the mechanism of action of tuftsin in vivo is not known, several types of leukocytes have been shown to become cytotoxic effector cells following activation with tuftsin. These cells presently include macrophages, natural killer cells, and granulocytes. The possibility that tuftsin can also activate other types of effector cells have not been ruled out. We feel this small peptide has a high potential (largely unrecognized) as an antitumor immunopotentiating agent. It is naturally occurring in man and appears to be relatively non-toxic. Its exact sequence (Thr-lys-Pro-Arg) is known and it can be chemically synthesized. Methods are also available to monitor the levels of tuftsin in body fluids. These properties along with its ability to control infectious disease make this agent one of the more promising immunopotentiators.

Animals↗
PubMed1981

Transport of Ca2+ and Na+ across the chromaffin-granule membrane.

Bovine chromaffin-granule ghosts accumulate 45Ca2+ in a temperature- and osmotic-shock-sensitive process; the uptake is saturable, with Km 38 microM and Vmax. 28 nmol/min per mg at 37 degrees C. Entry occurs by exchange with Ca2+ bound to the inner surface of the membrane. It is inhibited non-competitively by Na+, La3+ and Ruthenium Red (Ki 10.7 mM, 7 microM and 2 microM respectively), and competitively by Mg2+ (ki 0.9 mM). Uptake was not stimulated by ATP. Na+ induces Ca2+ efflux; Ca2+ can re-enter the ghosts by a process of Ca2+/Na+ exchange. La3+ inhibits Ca2+ efflux during Ca2+-exchange, and Ca2+ efflux induced by Na+, suggesting that Ca2+ uptake and efflux, and Ca2+/Na+ exchange, are catalysed by the same protein. Na+ enters ghosts during CA2+ efflux, but the kinetics of its entry are not exactly similar to the kinetics of Ca2+ efflux. Initially 1-2 Na+ enter per Ca2+ lost, but at equilibrium 3-4 Na+ have replaced each Ca2+. There is no evidence that either Ca2+ uptake or efflux by Ca2+/Na+ exchange is electrogenic, suggesting that the stoichiometry of exchange is Ca2+/2Na+. This exchange reaction may have a role in depleting cytoplasmic Ca2+ after depolarization-induced Ca2+ entry through the adrenal medulla plasma membrane; there is some evidence that there may be an additional entry mechanism for Na+ across the granule membrane.

Animals↗