Biomedical subjects

C C Curtain

Publications and source records attributed to C C Curtain.

At least 37 records · Page 2Linked to original sources

PubMed1987

Estimation of spin probe clustering in biological membranes.

An iterative spectral subtraction technique has been developed which accurately estimates the proportion of 'dilute' and 'clustered' I(12, 3) (i.e., 5-nitroxide stearate) in human erythrocyte ghosts at 37 degrees C, even if subtractant spectra free from probe-probe interactions cannot be measured due to technical limitations. Gordon et al. ((1985) J. Membrane Biol. 84, 81-95) earlier showed that I(12, 3) occupies a class of high-affinity sites in ghosts at probe/total lipid ratios (P/L) less than 1/2250. Saturation occurs with increasing probe concentration, and, at higher loading, the probe inserts itself at initially dilute sites to form membrane-bound clusters of variable size. Although this model allows determination of the dilute/clustered probe ratio, it requires subtraction of experimental spectra with a 'magnetically dilute' spectrum obtained using P/L less than 1/4600. The new methodology accurately profiles the % probe clustering in human erythrocyte ghosts over the entire P/L range, even if the lowest P/L for the subtractant spectrum contains substantial probe-probe interactions (i.e., P/L of 1/604 or 1/303). Application of either the subtraction technique in Gordon et al. (1985) or the iterative subtraction protocol described here should allow determination of probe clustering in a wide range of I(12, 3)-labeled biological membranes.

Cyclic N-Oxides↗

PubMed1987

Electron spin resonance spectroscopy in the study of lymphoid cell receptors.

ESR spectroscopy is a sensitive method which can be used to monitor a wide range of changes in and near the plasma membranes of lymphoid cells during such events as ligand-induced receptor patching and capping, activation and endocytosis and phagocytosis. The advantages of the technique are rapidity, sensitivity, small sample size (10(6)-10(7) cells), and nondestructive nature. Spin labels that are attached to a range of intrinsic and extrinsic molecules give information about the fluidity and polarity of the environment in which they are located. Because of the occurrence of interaction when probes are sequestered in restricted regions of the membrane, ESR spectroscopy is also a valuable technique for measuring the formation of domains in the cell membrane.

Cell Membrane↗

PubMed1987

ESR spectroscopy in the study of antigen processing--uptake of spin-labelled antigens by macrophages.

Macrophages were briefly pulsed with a spin-labelled synthetic polypeptide, poly(L-tyrosine:L-glutamic acid) poly DL-alanine:poly L-lysine (n-TGAL) in the presence and absence of anti-TGAL-antibody, and the electron spin resonance (ESR) spectra of the cell suspension compared with the spectrum of free n-TGAL in solution. Spectral analysis indicated two cell-associated n-TGAL pools, one composed of freely rotating label held in an aqueous environment, susceptible to protease digestion and ascorbate reduction, and a second highly concentrated pool, sequestered intracellularly, and held within a highly ordered, polar microenvironment. The ESR analyses were completed within minutes of antigen pulsing, employed very small numbers of live cells, and did not damage the cells being tested. The utility of the technique in screening fatty acid-antigen conjugates for macrophage uptake was demonstrated.

Animals↗

PubMed1985

Spin probe clustering in human erythrocyte ghosts.

A model has been developed for 5-nitroxide stearate, I(12,3), distribution in human erythrocyte ghosts which accurately predicts ESR spectral alterations observed with increased probe/total lipid (P/L) at 37 degrees C. This spin probe occupies a class of high-affinity, noninteracting sites at low loading. Saturation occurs with increasing probe concentration, and, at higher loading, the probe inserts itself at initially dilute sites to form membrane-bound clusters of variable size. No 'low' probe remains at high P/L where all I(12,3) clusters in a 'concentrated' phase. This model allows determination of the dilute/clustered probe ratio, and shows that I(12,3) segregates in erythrocytes at what might otherwise be considered low P/L (e.g., 1/359). These findings validate the earlier use of empirical parameters to estimate probe sequestration in biological membranes.

Cyclic N-Oxides↗

PubMed1985

A spin label study of the ionic strength dependent conformational change in the human Ia molecule.

Purified human Ia molecules were labelled with maleimide or isothiocyanate spin labels or by reacting "TEMPAMINE" spin label with the neuraminic acid of their carbohydrate residues. It was found that increasing the ionic strength from 0.05 to 0.75 markedly increased the dipolar interaction between the maleimide-attached labels, but no effect was found of ionic strength or motion on dipolar interaction with the other two labels. The effect of increasing ionic strength could be blocked by the prior addition of Ia-specific antibody, but could not be reversed by the addition of antibody after ionic strength was increased. These findings complement an earlier finding that increasing ionic strength over the range 0.05-0.75 has an inhibitory effect on the combination of Ia with its antibody. Because the maleimide spin labels attach predominantly to SH groups it is suggested that increasing ionic strength causes conformational changes in the immunoglobulin loop region which alter the accessibility of the Ia antibody-binding site.

Antibody Specificity↗

PubMed1983

Changes in the ordering of lipids in the membrane of Dunaliella in response to osmotic-pressure changes. An e.s.r. study.

Changes in the ordering and motion of lipids in response to changes in the external solute concentration have been studied by using the 5-nitroxide stearate (5NS) and 16-nitroxide stearate (16NS) spin probes in the plasma membrane of the halotolerant unicellular alga Dunaliella salina. Increases in ordering of the 5NS probe and decreases in motion of the 16NS probe were observed in cells equilibrated over 18 h at increasing NaCl concentrations. These changes probably resulted from the influence of the high NaCl concentration on the charged phospholipid head groups of the membrane. A short-term (less than 100 min) decrease in the order parameter, S, of the 5NS probe was observed for cells swollen by exposure to a sudden decrease of NaCl concentration from 5.0 to 2.5 M. After 100 min the value of S for 5NS was close to the value obtained in cells that had been equilibrated in 2.5 M-NaCl for 18 h. Since the cells had regained their original size and shape by 100 min it was assumed that the short-term decrease in S was associated with the swelling. A similar result was obtained when the cells were suddenly changed from 3.0 M- to 1.5 M-sorbitol. Conversely, an increase in S was observed for cells shrunk when the external solute concentration was doubled from 1.5 M- to 3.0 M-NaCl. As the cells regained their original size and shape the value of S decreased to the value observed in cells that had been equilibrated in 3.0 M-NaCl for 18 h. It is suggested that the changes in S are related to the movement of lipid into or out of a reservoir of membrane material as the membrane shrinks or expands. This movement of lipid maintains the tension of the membrane below the value at which it is disrupted. Such changes in lipid ordering could provide a mechanism whereby information about external osmotic-pressure changes is transmitted across the cell wall.

Ascorbic Acid↗

PubMed1980

Lipid domain formation and ligand-induced lymphocyte membrane changes.

Spectral parameters of spin-labelled phosphatidylcholine, ceramide and cerebroside in the plasma membranes of human blood lymphocytes were measured before and after treatment with various ligands, which included concanavalin-A and phytohemagglutinin. It was found that ligand treatment led to a significant decrease in order of the hydrocarbon chains of the phospholipids. This was accompanied by a clustering of the labelled spingolipids, as estimated by spin-spin interaction, and an increase in the order of their hydrocarbon chains. In the untreated cells the cerebroside fatty acid chain was more ordered than that of the phosphatidylcholine. It was considered that the decrease in phospholipid order was brought about by the sequestration of the more rigid sphingolipids into the patches and caps formed by receptor-ligand complexes. The significance of these changes in lipid distribution and ordering is discussed in relation to the activation of membrane enzyme systems by mitogenic ligands.

Cell Membrane↗

PubMed1979

Lymphocyte surface modulation and glycosphingolipids.

Using the fluorescent antibody method, identical localization of ligand and glycosphingolipids was found in human peripheral blood B lymphocytes patched and capped with polyvalent anti-immunoglobulin. cAMP also showed the same pattern of localization. It is suggested that glycosphingolipids are hydrogen bonded to receptor-bearing membrane macromolecules and are borne with them into the ligand-induced patches and caps. Here the asymmetric carbon chains of the glycosphingolipids modify the local lipid environment, leading to activation of the adenyl cyclase in the membrane which produces the cAMP.

Antibodies, Anti-Idiotypic↗

PubMed1978

Changes in lipid ordering and state of aggregation in lymphocyte plasma membranes after exposure to mitogens.

An electron spin probe study was made of the effect of a number of mitogenic agents on the ordering and state of aggregation of the plasma membrane lipids of lymphocytes. These agents, which included phytohemagglutinin, Concanavalin A, the calcium ionophore A23187 and periodate, caused a 20% decrease in lipid ordering in the region of the bilayer probed by 5-nitroxide stearic acid. The corresponding methyl ester probe showed marked probe-probe interaction under the same conditions indicating an aggregation of lipids in the area probed by this label. Studies with mixed lipid vesicles and ganglioside-free cells indicate that these areas are rich in glycolipids capable of hydrogen bonding to the ester probe. The decrease in ordering and the increase in aggregation of the membrane lipids were correlated with the patching and capping of the ligand-receptor complexes. Furthermore, the disappearance of fluorescent ligand from the surface of treated cells corresponded with the return of the spectral parameters of the probes to control cell values. It was concluded that glycolipids might play an important role in ligand-induced cell surface changes either as bearers of receptor groups, as in the case of some gangliosides, or in association by hydrogen-bonding with receptor proteins.

Animals↗

PubMed1977

Proteins retained with hyaluronic acid during ultrafiltration of synovial fluid.

The identity of the proteins associated with hyaluronic acid after ultrafiltration of bovine synovial fluid was examined. With discontinuous filtration, alpha2 macro- and IgM globulins were retained in hyaluronic complex both from synovial fluid and from a similar mixture of hyaluronic acid and serum. With continuous filtration and stirring to prevent formation of gels, protein was retained with hyaluronic acid in similar proportions. This protein showed immunologic identity with serum albumin, but differed in amino acid composition. After comparison with results of gel filtration, it was concluded that the identity of the retained proteins can be determined by the concentration of hyaluronic acid without variation in other ionic conditions.

Albumins↗

PubMed1976

The binding of dehydroheliotridine to DNA and the effect of it and other compounds on repair synthesis in main and satellite band DNA.

This study was aimed at elucidating the mechanisms of the preferential depression of satellite DNA synthesis by dehydroheliotridine (DHH). DHH was found to induce repair synthesis to the same extent in both main and satellite band DNA in cultured sheep lymphocytes. This was also the case with acridine orange, nitrogen mustard (HN2) and ethyl methane-sulphonate (EMS). Using analytical equilibrium ultracentrifugation no difference was found between the extents of in vitro binding of DHH by main and satellite band DNA. From these results it was concluded that the depression of the synthesis of satellite DNA could not be explained by either its preferential binding of DHH or by less effective repair mechanisms. Radiolabelled DHH when added to synchronized cultures of ovine kidney cells was found to be preferentially bound to the satellite DNA (one DHH molecule to 6000 nucleotides) compared with the main band DNA (one to 10 000). When 5-bromodeoxyuridine (BUdR) was added to the cultures no DHH label was found in the heavy, semiconservatively replicated DNA band. From these findings it is suggested that attack may occur during mitosis where all of the satellite DNA may be undergoing synthesis at the same time, thus explaining the increased amount of DHH bound to the satellite.

Animals↗

PubMed1975

Decreased synthesis of hepatic satellite DNA in pyrrolizidine alkaloidosis of the sheep.

the proportion of heavy satellite in the DNA isolated from the livers of sheep suffering from experimental pyrrolizidine alkaloidosis is significantly lower (3.5%) than that found in the DNA from lovers of normal sheep (12%). Dehydroheliotridine, the major unbound, relatively stable metabolite of lasiocarpine and heliotrine, the alkaloids used in the study, was found to inhibit selectively the semiconservative replication of the satellite DNA in cultures of ovine kidney cells. It is suggested that the inhibition of the synthesis of satellite DNA may be related to an attack by the metabolite on the pericentromeric region where the majority of the satellite sequences are located.

Animals↗

PubMed1974