Search PubMed⌕ Search

Biomedical subjects

D B Wilson

Publications and source records attributed to D B Wilson.

At least 253 records · Page 14Linked to original sources
PubMed1984

Hydrolysis of polyphosphoinositides by purified sheep seminal vesicle phospholipase C enzymes.

Sheep seminal vesicles contain two immunologically distinct phospholipase C (PLC) enzymes that can hydrolyze phosphatidylinositol (PI) (Hofmann, S.L., and Majerus, P.W. (1982) J. Biol. Chem. 257, 6461-6469). One of these enzymes (PLC-I) has been purified to homogeneity; the second (PLC-II) has been purified 2600-fold from a crude extract of seminal vesicles. In the present study we have compared the ability of these purified enzymes to hydrolyze PI, phosphatidylinositol 4-phosphate (PI-4-P), and phosphatidylinositol 4,5-diphosphate (PI-4,5-P2). Using radiolabeled substrates in small unilamellar phospholipid vesicles of defined composition, the two enzymes were found to hydrolyze all three of the phosphoinositides. Hydrolysis of all three phosphoinositides by both enzymes was stimulated by Ca2+; however, in the presence of EGTA only the polyphosphoinositides were hydrolyzed. The two enzymes displayed substrate affinities in the order PI greater than PI-4-P greater than PI-4,5-P2, and maximum hydrolysis rates in the order PI-4,5-P2 greater than PI-4-P greater than PI. When present in the same vesicles, PI and the polyphosphoinositides competed for a limiting amount of either enzyme. Inclusion of phosphatidylcholine into vesicles containing the phosphoinositides resulted in greater inhibition of PI hydrolysis than polyphosphoinositide hydrolysis. When all three phosphoinositides were present in vesicles mimicking the cytoplasmic leaflet of cell membranes, there was preferential hydrolysis of the polyphosphoinositides over PI. We conclude that a single phospholipase C can account for the hydrolysis of all three phosphoinositides seen during agonist-induced stimulation of secretory cells. The cytoplasmic Ca2+ concentration and phospholipid composition of the membrane, however, may influence the relative rate of hydrolysis of the three phosphoinositides.

Animals↗
PubMed1984

Inhibition of cell-mediated lympholysis by cloned and uncloned lines of natural killer cells and cytotoxic T lymphocytes with sugars and lectins.

This study was designed to further understand the nature of the interaction between natural killer (NK) cells and their susceptible targets. To do this, a panel of sugars and two lectins was tested for the ability to inhibit the lysis of NK-sensitive targets by cloned and uncloned lines of human NK cells. Six of these sugars (beta-gentiobiose, sucrose, alpha-lactose, beta-lactose, N-acetylglucosamine, and N-acetylgalactosamine) and one lectin, wheat germ agglutinin (WGA), proved to be potent inhibitors of the lytic activity of NK cells as well as of cytotoxic T lymphocytes activated in mixed lymphocyte cultures. Both beta-gentiobiose and WGA were shown to inhibit lysis at the level of the killer cell. Finally, the inhibitory effect of WGA could be reversed by addition of its sugar ligand, N-acetylglucosamine, which is itself an inhibitor of lytic function. From these findings it is concluded that these inhibitors probably do not act at the recognition stage of lysis since all of the NK and CTL lines tested, regardless of specificity, were inhibited by the same panel of sugars and lectins. Instead, it appears more likely that these inhibitors block some postrecognition stage of the lytic mechanism. The common inhibition profile by these sugars on NK and CTL activity further suggests that these two cell types may share, at least partially, a common lytic mechanism.

Acetylglucosamine↗
PubMed1984

Analysis of T cell populations that induce and mediate specific resistance to graft-versus-host disease in rats.

F1 hybrid (A X B) rats immunized with parental strain (A) T cells, or which recover from graft-versus-host (GVH) reactions caused by parental T cells, develop strong T cell-mediated immune responses against anti-major histocompatibility complex (MHC) receptor structures on donor T cells specific for host (MHCb) alloantigens. This immune response provides the basis for a profound and specific resistance to the subsequent induction in these animals of local or systemic GVH disease. Using subset-specific monoclonal antibodies and negative selection rosetting procedures, we attempted to determine which donor T cell subset possesses the immunogenic idiotypic markers, and which host T cell subpopulation mediates GVH resistance. We show here that the immunizing donor cell population belongs to the W3/25+ helper T cell (Th) subset, the same one that causes local GVH reactions, and that both the W3/25+ Th and the OX8+ killer/suppressor (Tk/s) subsets of host T cells are able to transfer GVH resistance to secondary F1 recipients.

Animals↗
PubMed1984

Negative selection in vivo reveals expression of strong Mls determinants in mice with X-linked immunodeficiency.

Evidence is presented that mice with X-linked immunodeficiency (xid) express strong Mlsa,d determinants, a putative marker of the mature subset of B cells. Although young (3-5 wk) (CBA/N X DBA/2)F1 male (xid+) mice stimulated only very weak mixed lymphocyte reactions (MLR) to Mlsa,d determinants, older mice (greater than 7 wk) regularly elicited conspicuous responses, despite being totally unresponsive to TNP-Ficoll. Expression of Mlsa,d determinants by xid+ mice was also detected by the procedure of negative selection in vivo. Thus BALB/c T cells were totally depleted of Mlsa,d reactivity after blood to lymph recirculation through 10-wk old irradiated xid+ (CBA/N X DBA/2)1 male mice. Significantly, a marked (90%) reduction in the anti-Mlsa,d response also occurred with T cell filtration through 3-wk xid+ mice, i.e., mice that elicit only minimal primary MLR; filtration through 3-wk xid- normal female mice led to near-complete (99%) negative selection. Collectively these data indicate either, (a) that xid+ mice contain appreciable numbers of cells with at least some of the properties of mature B cells, or (b) that the expression of Mlsa,d determinants is not restricted to mature B cells. In either case, B cells from xid mice cannot be viewed as a simple model for immature normal B cells.

Aging↗
PubMed1984

Monoclonal antibodies directed against HLA molecules affect the lytic and proliferative behavior of a cloned line of human natural killer cells.

Monoclonal antibodies with specificity for HLA class I and class II antigens were generated against a cloned line of human natural killer cells (NK 3.3). These antibodies were selected for their ability to either inhibit or enhance the lytic activity of NK 3.3. Antibody pretreatment and binding studies have established that the antibodies act at the level of the killer cell; we show in this paper that the nature of the target cell also plays a role in whether the antibody causes enhancement or inhibition of lysis. We provide evidence that these antibodies act at an early stage of lysis prior to the Ca++ dependent programming for lysis step, and describe the effects of these antibodies on the proliferation of cloned and uncloned lines of natural killer cells.

Antibodies, Monoclonal↗
PubMed1984

Isolation and characterization of the Salmonella typhimurium LT2 xylose regulon.

Salmonella DNA was partially digested with EcoRI, and the digest was fractionated to obtain fragments larger than 8 kilobases (kb). These were ligated into EcoRI-cut pBR322, and the mixture was used to transform Salmonella Xyl- cells selecting for ampR xyl+ transformants. A 21- and a 27-kb plasmid were isolated, both of which contained the entire xylose regulon. The xylose regulon was localized to a 6.3-kb segment of a 13.5-kb EcoRI fragment. Subclones were constructed which contained either the genes for D-xylose isomerase and D-xylulokinase or the genes for the D-xylose transport and the D-xylose regulatory factors. The gene order determined by the subcloning experiments is consistent with that determined by genetic mapping. The spots corresponding to D-xylose isomerase and D-xylulokinase subunits were identified in two-dimensional gels of several xylose-induced strains. Each of them had a molecular weight of 45,000 and an isoelectric point of 6.2 +/- 0.1.

Aldose-Ketose Isomerases↗
PubMed1984

Biosynthesis of coagulation Factor V by a human hepatocellular carcinoma cell line.

A human hepatocellular carcinoma line, HepG2, was found to secrete coagulation Factor V. Factor V activity in HepG2 culture fluid increased nearly linearly during a 20-h time course (5 ng Factor Va/h per 10(6) cells). Thrombin treatment increased Factor V activity in HepG2 culture medium six- to ninefold, indicating that the medium accumulates a mixture of Factors V and Va. To demonstrate de novo synthesis of Factor V, HepG2 cells were incubated in culture medium containing [35S]methionine. Labeled Factor V was immunoprecipitated from the medium and was shown to co-migrate with purified plasma Factor V upon sodium dodecyl sulfate polyacrylamide gel electrophoresis and fluorography. When medium was treated with thrombin before immunoprecipitation and fluorography, the 330,000-Mr [35S]methionine-labeled Factor V was converted to Factor Va. Factor Va coagulant activities from HepG2 cells and human plasma were inhibited in parallel by anti-Factor V antibody, indicating that HepG2 and plasma Factor Va have the same intrinsic activity. If normal hepatocytes synthesize Factor V at the same rate as HepG2 cells, then hepatocyte secretion can account for the total Factor V present in plasma. The production of Factor V by cultured human umbilical vein endothelial cells was also examined. Spent culture medium from endothelial cells contained only Factor Va and the amount was less than 1% of the activity found in medium from HepG2 cells under comparable conditions. The amount of Factor V activity in endothelial cell culture fluid did not change with time in culture.

Carcinoma, Hepatocellular↗
PubMed1984

Selected minerals in foods surveys, 1974 to 1981/82.

The levels of 11 essential minerals in food commodity groups and total diets of adult males, infants, and toddlers were summarized from the Selected Minerals in Foods Survey, Fiscal Years (FY) 1974 through mid-1982, of the Food and Drug Administration (FDA) to evaluate the sources and adequacy of those minerals in the diets. The three diets were adequate in calcium, phosphorus, magnesium, potassium, manganese, and selenium; low in copper; and elevated in sodium and iodine. Iron was adequate in adult male but not in infant or toddler diets, and zinc was adequate in infant but somewhat low in adult male and toddler diets. Dairy products; meat, fish, and poultry; grain and cereal products; and fruits and vegetables were the major sources of the 11 minerals. Some statistically significant differences in mineral levels were found among years for total diets and food commodity groups. Regression analysis indicated several linear trends in the mineral content of food commodity groups but no linear trends in the mineral levels of total diets. FDA continues to monitor the levels of minerals in food and diets through the revised Total Diet Study program, which began in mid-FY82.

Adult↗
PubMed1984

Fine structural aspects of the cranial neuroepithelium in early embryos of the rhesus monkey.

Temporal changes in the ultrastructure of cranial neuroepithelial cells and their developmental interactions with adjacent chordamesodermal cells were analyzed during early neurogenesis in rhesus monkey embryos at stages 8-12 (19-28 days of gestation). In the neural plate and early neural fold stages, microtubules and apical microfilaments are not as prominent in the neuroepithelial cells as they are after the neural tube closes, whereas gap junctional vesicles are more evident prior to closure. Initially, the association between neuroepithelial cells and the presumptive notochordal cells is relatively closer than that between neuroepithelial cells and mesenchymal cells. After neural tube closure, the notochord becomes more distant. Cytoplasmic processes from neuroepithelial cells, notochordal cells, and mesenchymal cells interact via the neural basal lamina. In the rhesus monkey embryo, the ultrastructural features of neuroepithelial cells and their interactions with adjacent chordamesodermal cells are similar to those which have been observed in the mouse at comparable stages of gestation.

Animals↗
PubMed1983

Epitopes on H-2Dd somatic cell mutants recognized by cytotoxic T cells.

We have generated several cell lines that express an altered H-2Dd molecule. These cell lines, which were selected for by the failure to express the serological specificity reacting with the monoclonal antibody 34-2-12, have also undergone alterations in epitopes recognized by CTL. One of the mutants, 2.12(-4) was not killed by an allogeneic anti-Dd CTL line, CTLL-A2, even though this line was cytotoxic for the parental cell line and two other 34-2-12- mutant lines. Two of the 34-2-12- mutant lines had an identical serological profile using other monoclonal Dd antibodies, however these two mutants differed markedly in their susceptibility to cytotoxicity by CTLL-A2. In addition to the determinants recognized by allogeneic CTL we also examined the effect of the mutation on the determinants involved in restricting the anti-FITC modified-self-cytotoxic response. An anti-FITC-Dd CTL line did not react with two of the mutants and reacted only weakly with the other mutant, demonstrating not only that the Dd epitopes recognized by this cell line and the allogeneic CTL were different, but also that it is possible for a H-2 class I molecule to express epitopes recognized by allogeneic CTL but not epitopes that function as restricting elements to certain antigens. The observation that both T cell- and B cell-defined determinants were altered in these mutant cell lines is in contrast to the findings, with the mutant mouse strains which were selected for by changes in T cell-defined determinants, which show few, if any, alterations to serological specificities. Characterization of T cell-recognized epitopes expressed on serologically selected somatic cell variants may therefore prove to be most useful for the study of structure-function relationships of H-2 class I molecules.

Animals↗
PubMed1983

Fine structure of pituitary blood vessels in embryos of the dwarf (dw) mutant mouse.

Blood vessels in the pars distalis of the embryonic pituitary gland of the dwarf (dw) mutant mouse were analyzed by means of electron microscopy. Dwarf (dw/dw) embryos ranging in age from 15 through 18 days of gestation were obtained from matings of dwarf homozygotes in which reproductivity was induced by means of hormone supplementation and grafts of normal pituitary to the kidney capsule. Endothelial cells in vessels of the dwarf pituitary were retarded in development as indicated by a retention of macrovesicles and marginal flaps beyond embryonic stages when they normally subside in normal embryos, as well as a lag in the development of endothelial cell attenuations. This retardation may be causally related to the pituitary pathology involving faulty cytodifferentiation of somatotrophs and mammotrophs.

Animals↗
PubMed1983

Early development of the otocyst in an exencephalic mutant of the mouse.

Development of the otic anlage was studied by means of scanning electron microscopy in exencephalic loop-tail (Lp/Lp) embryos and in their normal (+/+; Lp/+) littermates at 8.5-9.5 days of gestation. In the abnormal embryos, the cells of the otic pit lack the rounded prominent surfaces and dense arrangement of microvilli which characterize the normal otic pit, and the overall configuration of the otic anlage is also distorted. The relatively early manifestation of abnormalities in the otic anlage of this mutant suggests that the defective neural tube may exert an influence as early as the otic placode stage, or that a common factor may simultaneously affect both otic and neural tube development in this mutant.

Animals↗
PubMed1983

High affinity esterification of eicosanoid precursor fatty acids by platelets.

We have examined the relative rates of uptake of several fatty acids into washed, human platelets by measuring incorporation into cellular phospholipids. In the presence of 15 microM fatty acid-free albumin and with radioactive fatty acid concentrations of 5-500 nM, esterification into phospholipid was linear with time and platelet concentration and saturable with respect to fatty acid concentration. Two distinct classes of uptake rate were observed. Arachidonate and 5,8,11,14,17-eicosapentaenoate exhibited high affinity, relatively rapid incorporation into platelet phospholipids at pH 6.5: apparent Michaelis constant (Km) = 30 nM, apparent maximum velocity (Vmax) = 28 pmol/min per 10(9) platelets. Two other eicosanoid precursors, 5,8,11-eicosatrienoate and 8,11,14-eicosatrienoate, exhibited the same Vmax, but Km of 85 and 60 nM, respectively. Under the same conditions, stearate, oleate, and linoleate were incorporated into phospholipids much less efficiently (Vmax approximately 8 pmol/10(9) cells per min, apparent Km greater than or equal to 170 nM). Qualitatively similar results were found at pH 7.4. Uptake of radiolabeled, rapid-uptake fatty acids was not diminished by the presence of excess, unlabeled, slow-uptake fatty acids. Thus, the specificity of this esterification system resembles that of the arachidonate-specific, long-chain acyl-CoA synthetase present in platelets. It may represent the expression in vivo of the synthetase, although the apparent affinity of the synthetase for fatty acid is much less. This esterification system probably represents the physiologic mechanism for platelet arachidonate uptake, whereby arachidonate is collected from plasma, despite the fact that its concentration is considerably lower than that of other plasma fatty acids.

8,11,14-Eicosatrienoic Acid↗
PubMed1983

Age-related alterations in thyroid hormone binding properties of porcine skeletal muscle cytosol.

Thyroid hormone binding properties were evaluated in cytosol prepared from semitendinosus muscle of 1-d, 3-wk and 5-mo-old Yorkshire-Duroc crossbred pigs. Optimum thyroxine (T4) binding was obtained with 35 ug cytosolic protein in a solution containing 50 mM Tris-HCl buffer, pH 7.4, and 25 mM KCl and 10 mM MgCl2. The half-time of maximum T4 binding was approximately 10 min. Protease digestion of cytosol eliminated specific T4 binding. Muscle cytosol preferentially bound T4 over triiodothyronine (T3). At equimolar concentrations the amount of T3 binding relative to T4 was .1, 50 and 10% at 1 d, 3 wk and 5 mo, respectively. Scatchard analysis of T4 binding revealed a similar high-affinity, low-capacity component in all age groups. However, at 3 wk of age the concentration of the low-affinity, high-capacity binding component for T4 was 12-fold greater than at either 1 d or 5 mo of age. The affinity of this high-capacity binding component did not change with age. The function of thyroid hormones in regulating skeletal muscle growth and metabolism have not been elucidated. These data demonstrate that thyroid hormone binding properties of skeletal muscle do change with age and further research is needed to investigate the possible relationship of these findings to growth regulatory mechanisms.

Age Factors↗