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K N Traill

Publications and source records attributed to K N Traill.

18 recordsLinked to original sources
PubMed1991

Correlation of lymphocyte lipid composition membrane microviscosity and mitogen response in the aged.

Healthy aged and young blood donors were investigated for the role of membrane lipid composition in the age-related increase in membrane microviscosity and decline of mitogen responsiveness. Membrane microviscosity was shown to correlate positively with membrane cholesterol/phospholipid molar ratios, which were significantly elevated in the elderly. A positive correlation also was confirmed between lymphocyte membrane microviscosity, which was measured using the probe 1,6-diphenyl 1,3,5-hexatriene, and phytohemagglutinin responsiveness of cells from the same donor. Using stepwise regression statistical analysis, the variables age, cholesterol, cholesterol/total phospholipid and phosphatidyl ethanolamine/phosphatidyl choline molar ratios were all shown to have a significant positive influence on membrane microviscosity, whereas total phospholipids had a negative effect. No statistically significant difference was seen in content of any single saturated or unsaturated fatty acid between young and old donors. After pooling, however, the proportion of all unsaturated fatty acids was significantly higher in cells from the elderly as a consequence of an increase of n-6 and n-3 polyunsaturated fatty acids. Changes in lipid composition and physical properties of lymphocyte plasma membranes may, therefore, be responsible (at least partially) for the diminution of immune reactivity in old age.

Adult↗
PubMed1990

Lipoprotein interactions with T cells: an update.

The role of plasma lipoproteins in atherogenesis is well recognized but the physiological relevance of their immunoregulatory properties is still questioned. Here Karine Traill and colleagues outline the recent advances that have been made towards unravelling the mechanisms of immunoregulation by lipoproteins in vitro and consider whether any of these mechanisms are operative in vivo. In particular they address the possible detrimental effects of high serum lipoprotein levels on immune function and the question of whether hyperlipidemia (or hypercholesterolemia) should be considered a risk factor for diminished immunity, for example in old age.

Aging↗
PubMed1990

Increased expression of high-affinity low-density lipoprotein receptors on human T-blasts.

Like all cells, lymphocytes need cholesterol for proper function, a requirement met by a finely tuned homeostasis between intracellular synthesis and uptake from the environment via low-density lipoproteins (LDL). We used flow cytometry to analyze the receptor activity of resting cells and T blasts incubated/activated in serum-free culture medium, or in medium supplemented with 25-5,000 micrograms/ml LDL. Dioctadecyl-indocarbocyanine has proved to be a useful fluorescent probe for investigating the LDL receptor activity of lymphocytes. The results show the receptor activity of day-3 resting T cells to be reduced more than 50% by 50 microgram LDL/ml, whereas 100-fold higher concentrations are necessary to achieve the same level of reduction in day-3 PHA blasts. The LDL receptor activities of individual blood donors' resting T cells, in vitro cholesterol-deprived resting T cells, and activated T blasts, were compared using two analytical techniques: spectrofluorometric analysis of detergent-solubilized cell suspensions and flow cytometric analysis of single living cells. Receptor affinity was determined by Scatchard analysis of spectrofluorometric binding curves, and by Line-weaver-Burke plots of flow cytometric data. Both methods yielded essentially identical dissociation constants (Kd) for cholesterol-deprived resting T cells and mitogen-activated T blasts, which fell in the expected range for the high-affinity LDL receptor (4.1-8.9 nM). In addition, spectrofluorometric analysis, but not flow cytometry, permitted quantification of LDL uptake.(ABSTRACT TRUNCATED AT 250 WORDS)

Adult↗
PubMed1989

Promotion of lymphocyte growth by high density lipoproteins (HDL). Physiological significance of the HDL binding site.

The characteristics and physiological relevance of the high density lipoprotein (HDL) binding site on unstimulated and mitogen activated human peripheral blood lymphocytes have been investigated. At 37 degrees C, specific binding/uptake of fluorescent (dioctadecylin-docarbocyanine, DiI) HDL was observed by cells from healthy donors as well as by those from low density lipoprotein receptor-defective patients; mitogen activated T-blasts exhibited a markedly elevated DiI-HDL uptake compared to resting T-cells. Binding was saturable at 37 degrees C and of high affinity, with a Kd of 5 x 10(-8) M. It was blocked by anti-apoAI polyclonal antibodies (F(ab)2 fraction), but not by anti-apolipoprotein (apo)E, anti-apoAII, or anti-apoB, and was inhibited competitively by HDL apoproteins and an apoAI-protein A fusion protein. T-cell associated DiI-HDL was increased by trypsin treatment (of the cells) and decreased by activation in the presence of HDL or low density lipoprotein. Comparison of the concentration dependencies of growth promotion and specific cell association of HDL indicated that two mechanisms of lipid exchange may be in operation: one a binding-dependent mechanism of cholesterol exchange, with maximal effect in the HDL concentration range (20-200 micrograms/ml) in which specific binding increases rapidly, and the other a binding-independent exchange of lipids effective at concentrations in which specific binding is saturated (300-5000 micrograms/ml).

Adult↗
PubMed1989

Use of a FACS III for fluorescence depolarization with DPH.

We have previously demonstrated age-related differences in human lymphocyte membrane fluidity, by use of steady-state polarization measurements on bulk cell suspensions with the fluorescence probe DPH. However, for exact analysis of the possible functional importance of these changes, single-cell measurements were deemed of interest. We have now used an analog division device to measure fluorescence depolarization "p" of DPH in real time with a FACS III flow cytometer. The measurements are reliable, as we have been able to confirm the differences in DPH "p" between monocytes and lymphocytes previously shown in bulk suspension and to demonstrate the expected differences in fluidity of lipid-modulated cells. We also found significant differences in DPH "p" between lymphocytes of young and elderly blood donors. Lymphocyte subsets did not differ in polarization values but did differ in fluorescence intensity with Th less than Ts less than B = NK cells.

Adult↗
PubMed1989

Immunodeficiency in old age.

Aging is a multi-facetted process, but the deterioration of the immune function seems to play a central role in this context. Paradoxically, immune reactivity against exogenous antigens declines during aging while autoimmune reactivity increases. One of the aims of our investigations on the function of the senescent immune system was to define immune parameters of 'normal' aging, i.e. those not dependent on underlying diseases. Specifically, our interest was focused on the possible role of an altered lipid metabolism of cells of the immune system during aging. The known decrease of plasma membrane fluidity of lymphoid cells and monocytes in higher age may be one of the factors responsible for the nonoptimal functioning of the immune system. This property, in turn, seems to be based on an altered lipid metabolism. Specifically, we have evidence that the finely tuned balance between the transport of cholesterol to and from lymphoid cells via the environment and the intracellular cholesterol biosynthesis seems to be disturbed with increasing age. This conclusion is drawn from experiments where low-density lipoprotein (LDL) and high-density lipoprotein (HDL) receptor activity is assessed using fluorescently labeled lipoproteins in fluorescence-activated cell sorter (FACS) analyses. LDL receptor uptake is unexpectedly increased in the elderly, but LDL receptor regulation, and serum LDL composition itself seem to be normal. Preliminary data point out the possibility that the efflux of cholesterol via HDL may be insufficient. Attempts to modulate plasma membrane fluidity by means of the phospholipid mixture 'active lipid 721' (AL 721) showed that this drug, in contrast to literature reports, is not a 'membrane fluidizer' but rather exerts this effect as a nutrient for lymphocytes and monocytes.

Aging↗
PubMed1988

Approach to define "normal aging" in man. Immune function, serum lipids, lipoproteins and neopterin levels.

In 53 healthy women with mean age 25.1 years (age range 21-34 years) and in 51 healthy women with mean age 82.1 years (age range 75-91 years), a panel of immunological and biochemical tests was performed. These tests, comprising lymphocyte mitogen responsiveness, phenotyping of lymphocytes, uptake of low density lipoprotein by T cells, serum levels of neopterin, lipids and lipoproteins, as well as routine blood chemistry, were investigated for a possible effect of age and of the classification according to the SENIEUR Protocol of admission criteria by the European Economic Community's Concerted Action Program on Aging (EURAGE). A highly significant effect of age on serum levels of neopterin, lipids and lipoproteins was found. No clear effects, however, of SENIEUR status on these variables was detected. As expected, age had a significant impact on mitogen responsiveness of T cells. Proportional numbers of helper/inducer and cytotoxic/suppressor T cells (as well as antigen density on these cells) were not influenced by age. SENIEUR classification did not affect these immunologic variables. Thus, most of the tested variables that are not included in the SENIEUR admission criteria appear to present information not yet covered by the SENIEUR variables. Various ways for a possible revision or extension of the SENIEUR Protocol are discussed.

Adult↗
PubMed1988

Lipid requirements of human T lymphocytes stimulated with mitogen in serum-free medium. Membrane "fluidity" changes are an artefact of lipid (AL721) uptake by monocytes.

A correlation has been sought between the effects of lipids on membrane fluidity and mitogen responsiveness of human peripheral blood lymphocytes (PBL). Cholesterol and the reputedly potent membrane fluidizing agent AL721 (1) were used for these studies. However, the large AL721 induced increase in membrane "fluidity", assessed by steady state polarization of the probe 1,6-diphenyl-1,3,5-hexatriene (DPH), was found to be an artefact arising from lipid uptake by monocytes. Mitogen responses were enhanced by AL721 but unaffected by cholesterol. It is concluded that AL721 does not exert its effect through enhanced triggering of cells by altered membrane fluidity but rather that lymphocytes require an exogenous source of phospholipids/triglycerides for optimal growth in vitro, although they can synthesize sufficient cholesterol to meet their own needs.

Adult↗
PubMed1987

Analysis of fluorescent low density lipoprotein uptake by lymphocytes. Paradoxical increase in the elderly.

Uptake of dioctadecylindocarbocyanine (DiI)-labelled low density lipoproteins (LDL) by peripheral blood lymphocytes (PBL) from young healthy donors has been characterized by flow cytometric analysis. The receptor positive cells were primarily (greater than 70%) T cells. Saturation and competition studies were performed with freshly isolated PBL as well as after a 2-3-day incubation in cholesterol-free medium. In both cases uptake was specific for LDL and not high density lipoprotein. It was also abrogated by chemical modification of apo B, and was not shown by PBL from a patient with familial hypercholesterolemia. DiI-LDL-uptake by cells from elderly donors was compared with that of PBL from young donors. There was a clear increase in uptake by freshly isolated PBL from aged donors which was shown not to stem from underlying "sickness". In contrast, uptake by pre-incubated cells was very variable, in terms of percentage receptor-positive cells and the level of uptake by those cells. However, LDL rescued mevinolin-suppressed mitogen responses from both old and young donors indicating that there is no impairment of uptake or degradation of LDL by PBL from the elderly.

Adult↗
PubMed1987

High density lipoprotein uptake by freshly isolated human peripheral blood T lymphocytes.

A high density lipoprotein (HDL) receptor/binding site has been identified on peripheral blood lymphocytes, and some of its properties were compared with those of HDL receptors on other cell types. Binding studies were performed using fluorescent (dioctadecylindocarbocyanine)-labelled HDL (DiI-HDL) and analyzed by fluorescence microscopy and quantitative flow cytometry. Uptake of low levels of DiI-HDL during a 2 h incubation at 37 degrees C was a property of all lymphocytes, i.e. not of one particular subset only. Visual inspection of these cells in the fluorescence microscope revealed both membrane and cytoplasmic fluorescence, indicating that DiI-HDL become internalized during the 2 h incubation; internalization appeared to be a receptor-mediated process. In competitive binding studies, apo E-free HDL competed effectively for DiI-HDL binding, whereas LDL competed very weakly. Two features of DiI-HDL uptake are demonstrated which are unique to lymphocytes: 1) it was enhanced 3-6-fold by inclusion of EDTA in the incubation medium or by incubating in Ca2+/Mg2+ free medium, and 2) it was saturable at 37 degrees C.

Adult↗
PubMed1986

Lymphocyte membrane lipid composition and mitogen responsiveness in chickens: role of membrane "fluidity".

After establishing optimal conditions for measuring the membrane lipid packing density ("fluidity") of chicken peripheral blood lymphocytes, the fluidity was modulated in vitro by incubation in cholesterol or phospholipid ("active lipid", AL)-enriched serum-free tissue culture medium. The effect of these lipids on mitogen responsiveness was then investigated, the aim being to determine whether the observed enhancement/suppression was membrane mediated, i.e. explainable by fluidity changes. Chicken peripheral blood lymphocytes exhibited no requirement for exogenous cholesterol; low concentrations did not affect the mitogen response while the higher concentrations, which induced a measurable decrease in membrane fluidity, were usually mildly suppressive. Pre-incubation did not increase this suppressive effect and we believe it not to be membrane mediated. AL, at low concentrations which induced no changes in membrane fluidity, prolonged the phytohemagglutinin response, enhancement being evident only after the peak; we interpret this as a nutrient effect. At the higher concentrations, which induced large increases in fluidity, a transient enhancement was followed by suppression; suppression was delayed in onset when AL was added 4 h after phytohemagglutinin stimulation. It is therefore an early event which may be mediated through changes in membrane fluidity.

Animals↗
PubMed1986

Simple method for comparing large numbers of flow cytometry histograms exemplified by analysis of the CD4 (T4) antigen and LDL receptor on human peripheral blood lymphocytes.

We have developed a simple method for comparing the relative fluorescence intensity (FI) of flow cytometry histograms. It entails assessment of the FI (equivalent to the fluorescence-activated cell sorter (FACS) channel) of the 50th or 75th percentiles of either positively stained cells or the total cell population. We illustrate the method with dilution curves of 1) monoclonal antibodies against the T4 surface antigen of human peripheral blood lymphocytes and 2) fluorescent low density lipoprotein (LDL) binding to the human peripheral blood lymphocytes LDL receptor. We demonstrate the versatility of the method by characterizing the binding properties of fluorescent LDL to their receptors. Binding was shown to be specific and of high affinity, and to reach a steady state plateau at about 2 hr; the affinity of fluorescent LDL for the receptor was found to be two to three times higher than that of the unlabeled LDL.

Antibodies, Monoclonal↗
PubMed1985

Age-related changes in lymphocyte subset proportions, surface differentiation antigen density and plasma membrane fluidity: application of the eurage senieur protocol admission criteria.

Peripheral blood lymphocytes from 260 "apparently healthy" males and females, aged 20-97, have been investigated for age-related changes in a number of immunological parameters (percent and number of lymphocytes, OKT 4+ and OKT 8+ cells; OKT 4/8 ratio; intensity of fluorescence of OKT 4 and OKT 8 stained cells; membrane fluidity). Data were then reassessed after exclusion of people who did not conform to the SENIEUR PROTOCOL admission criteria of EURAGE (European Economic Community's Concerted Action Program on Aging) in order to investigate whether the differences observed were attributable to underlying disease. A slight decrease in the number and percentage of lymphocytes, OKT 4+ and especially OKT 8+ cells was found. Intensity of fluorescence of OKT 4 and OKT 8 stained cells from the elderly was reduced and may explain the lower percentages. Membrane fluidity was decreased in old persons (over the age of 75); the free cholesterol/phospholipid molar ratio in the serum increased up to the age of 75 and then declined, and it did not correlate with decreased membrane fluidity. Exclusion of the 20-60% of the study groups who were not eligible for admission according to the SENIEUR PROTOCOL criteria did not affect these results. The feasability and applicability of the PROTOCOL is discussed.

Adult↗
PubMed1984

Ontogeny of surface markers on functionally distinct T cell subsets in the chicken.

Three subsets of chicken peripheral T cells (T1, T2 and T3) have been identified in peripheral blood of adult chickens on the basis of fluorescence intensity after staining with certain xenogeneic anti-thymus cell sera (from turkeys and rabbits). They differentiate between 3-10 weeks of age in parallel with development of responsiveness to the mitogens concanavalin A (Con A), phytohemagglutinin (PHA) and pokeweed mitogen (PWM). Functional tests on the T subsets, sorted with a fluorescence-activated cell sorter, have shown that T2, 3 cells respond to Con A, PHA and PWM and are capable of eliciting a graft-vs.-host reaction (GvHR). In contrast, although T1 cells respond to Con A, they respond poorly to PHA and not at all to PWM or in GvHR. There was some indication of cooperation between T1 and T2,3 cells for the PHA response. Parallels between these chicken subsets and helper and suppressor/cytotoxic subsets in mammalian systems are discussed.

Aging↗
PubMed1984

Lack of correlation between serum cholesterol levels, lymphocyte plasma membrane fluidity and mitogen responsiveness in young and aged chickens.

Chickens were studied in an attempt to demonstrate correlations between serum lipid levels and peripheral blood lymphocyte (PBL) plasma membrane fluidity and mitogen responsiveness: (a) in the laying hen; (b) during aging; and (c) following dietary manipulation of serum cholesterol of young and aged chickens. The membrane fluidity of PBL from laying hens was significantly greater than that of immature birds. However, no direct correlation was found between serum lipid levels, nor the serum free cholesterol/phospholipid (FC/Pl) mole composition and PBL membrane fluidity in any of the age-groups tested. Likewise, no correlation was found either between serum FC/Pl mole ratio or membrane fluidity and mitogen responsiveness of PBL from birds up to 5 years of age nor was there any evidence for a decline in mitogen responsiveness up to this age. Supplementation of diets with 1% cholesterol induced hypercholesterolemia, mainly in the very low density lipoprotein (VLDL) fraction, but membrane fluidity and mitogen responsiveness remained unaffected.

Aging↗
PubMed1983

Chicken thrombocytes. Isolation, serological and functional characterisation using the fluorescence activated cell sorter.

Different procedures for enrichment of peripheral blood leukocytes (PBL) from whole chicken blood have been compared in terms of the percentage yield of lymphocytes and thrombocytes. The yield of thrombocytes was low in the buffy coat cells of heparinised blood, but high in PBL enriched from heparinised blood over a Ficoll-Paque density gradient and in PBL prepared from citrated blood by either technique. The fluorescence activated cell sorter (FACS) has been used to separate thrombocytes from lymphocytes in PBL by sorting of the negative cells after staining with specific anti-T and anti-B cell sera (or anti-immunoglobulin). The sorted cells were more than 99% viable, more than 98% thrombocytes by antigenic and morphological criteria and did not respond in vitro to T and B cell mitogens. Absorption studies on a turkey anti-bursa cell serum (ABST) which is cross-reactive for B cells and thrombocytes (8) indicated that the cross-reactivity is attributable to two (or more) antibody specificities in the serum rather than to one antibody directed at shared determinants on the two cell types.

Animals↗
PubMed1981

A simple micromethod for MLC and CML with low numbers of murine lymph node and peripheral blood lymphocytes.

A fast and simple method is described for the mixed lymphocyte culture (MLC) of low numbers of murine lymph node (LN) and peripheral blood lymphocytes (PBL). Cultures were set up with 1 X 10(4) - 4 X 10(4) responder cells in wells of inverted Terasaki plates where the cells settled at the meniscus of a hanging drop. [3H]thymidine incorporation and 51Cr-release from appropriate allogeneic target cells were determined either directly in the same hanging drop or after cell transfer and serial 2-fold dilutions in new Terasaki plates. Replicate measurements were reproducible by both methods. Responder and stimulator cell surface antigens could also be tested by addition of antisera with complement, either at the initiation or at the effector stage of culture. Since culture volumes were only 15 microliter, almost negligible quantities of antisera were required. The method was thus particularly valuable for testing with antisera of limited availability. Examples are given where inclusion of an anti-responder alloantiserum plus complement on day 0 of the MLC eliminated the day 4 cytotoxic response, and where treatment of cytotoxic effector cells with alloantiserum plus complement for 1 h prior to the 51Cr-release assay eliminated the effector cell activity.

Animals↗