Eda T. Bloom

Impaired natural killer cell activity during bereavement

Natural killer (NK) cell activity, a component of the immune surveillance system, was compared in women whose husbands had recently died with that found in age-matched women who had not experienced recent adverse life events. Bereaved women had significantly lower NK activity than women whose husbands were healthy. In a second study, depressive symptoms and NK activity were measured longitudinally in women before and after the death of their husbands. Our results suggest that depressive symptoms, not merely the death of the spouse, are related to a reduction in NK activity during bereavement.

Plasma cortisol and natural killer cell activity during bereavement

Natural killer cell (NK) activity, which is important in the defense against tumors and viral infections, is reduced in women undergoing conjugal bereavement. The relationship between NK activity and plasma cortisol was investigated in three groups of subjects: women who were anticipating the death of their husbands, women whose husbands had recently died, and controls. Bereaved women showed reduced NK activity and increased plasma cortisol levels as compared to controls. Anticipatory bereaved women also showed significant reductions in NK activity, but had levels of plasma cortisol comparable to those of controls. The reduction of NK activity during anticipatory and actual bereavement cannot be explained solely on the basis of increased cortisol secretion.

Quantification of natural cytotoxicity by human lymphocyte subpopulations isolated by density: Heterogeneity of the effector cells☆

Natural cell mediated cytotoxicity has been expressed as percent cytotoxicity, as the slope of the titration curve obtained by testing different effector: target cell ratios, and as lytic units. Objections can be raised to each method as used. The present report involves the study of cytotoxicity by subpopulations of lymphocytes obtained by Percoll density gradient centrifugation. The subpopulations vary greatly in cytotoxic activity, making accurate comparisons by traditional means difficult. A method was therefore developed for making objective comparisons between activities of subpopulations of lymphocytes. Cytotoxicity titration curves, which were sigmoidal on linear-linear plots, were found to best fit the sigmoidal curves described by the Von Krogh equation. A best fitting scale family of curves having the identical maximum cytotoxicity and shape parameter was fitted simultaneously to cytotoxicity measurements obtained by titrating all subpopulations of effector cells obtained from each patient. Values expressing relative cytotoxicities were obtained from the ratios of the scale parameters of the different curves or by obtaining lytic units from the fitted curves. In addition to the enriched natural cytotoxic activity found among the lighter cells obtained by Percoll density gradient centrifugation, activity was also increased among cells sedimenting at the very bottom of the gradient. This was found in subpopulations of cells obtained from 16/21 gradients in tests against K562 and in subpopulations obtained from both of 2 gradients in tests against Daudi cells. These findings are consistent with the existence of at least 2 subpopulations of lymphocytes which can mediate natural cytotoxicity, and which are separable by density.

Monocyte-mediated augmentation of human natural cell-mediated cytotoxicity☆

Normal human monocytes can significantly and rapidly augment natural cell-mediated cytotoxicity (NCMC) against K562 target cells. Approximately 50% augmentation was observed after direct mixture of monocytes with autologous null cells in the 4-hr chromium-release assay. This effect was dependent on the number of monocytes, and B cells and granulocytes were not effective. Coculture of null cells with monocytes and subsequent recovery of null cells for use as effector cells also produced significantly elevated cytolytic activity. This effect was dependent upon the number of monocytes, the length of time of coculture, and the cell donor. Augmentation of NK activity was rapid and observed after 0.5-12 hr of coculture, but suppression was observed after 36 hr; augmentation was observed with high monocyte:null cell (1:1, 1:2) ratios, and no effect was generally observed with lower ratios (1:8). At the single-cell level, the augmentation was associated with an increase in the proportion of target-binding cells which were lytically active. The augmentation of NK activity by monocytes required close cellular proximity, was mediated by a factor which was active or induced only in close proximity of the effector and producer cells, and/or was mediated by a soluble factor with a molecular weight greater than 50,000. This new demonstration that monocytes can augment as well as suppress NCMC may represent another avenue by which NK cell activity may be modulated in vivo.

Delayed effects of low-dose radiation on cellular immunity in atomic bomb survivors residing in the United States.

Several parameters of cellular immune function were assessed among persons who survived the 1945 atomic bombs in Hiroshima and Nagasaki but who now reside in the United States. The subjects in this study were exposed to various low doses (T65D) of radiation at the time of the bomb. More than half received an estimated 0 Gy (S0 group). Of those exposed to more radiation (S+ group), nearly 90% received less than 0.50 Gy (50 rad). Lymphocytes were isolated from the peripheral blood of these individuals and were assessed for the following parameters of cellular immunity: mitogenic response to phytohemagglutinin, mitogenic response to allogeneic lymphocytes, natural cell-mediated cytotoxicity (NCMC), and interferon production. In every case, the response of the S+ group was greater than that of the S0 group, although only the difference for NCMC was statistically significant. Results of studies presently being performed on A-bomb survivors residing in Hiroshima do not confirm this difference. Therefore, it is difficult to say whether the increase in natural cytotoxicity observed among the American and not the Japanese A-bomb survivors exposed to very low doses of radiation is a hormetic effect which was modulated by post-radiation environmental conditions or a result of selective migration.

Age-related decline in the lethal hit but not the binding stage of cytotoxic T-cell activity in mice

The activity of allogeneic cytotoxic T lymphocytes (CTL) declines with aging, as measured at the population level in chromium release assays, and is associated with a decreased proliferative response to the inducing alloantigens. At the single cell level, no difference was noted between target binding cells obtained from alloantigen-stimulated spleen cells of young and old mice. However, a consistent, large, and significant difference was observed between CTL generated from the spleens of young and old mice in the percentage of bound effector cells with lysed targets, i.e., the percentage of killer cells. These results show age-related changes in the lethal hit stage and not the binding stage of CTL activity. This change could be used as a probe to identify the important mechanism(s) which mediates CTL activity.

Rapid kinetics of lysis in human natural cell-mediated cytotoxicity: some implications

The entire lytic process of natural cell-mediated cytotoxicity against sensitive target cells can occur rapidly, within minutes. This was demonstrated by 51chromium release and in single-cell assays. At the cellular level, most of the target cell lysis occurred within 15-30 min after binding to effector cells. The enriched natural killer cell subpopulation of lymphocytes obtained Percoll density gradient centrifugation (containing greater than 70% large granular lymphocytes (LGL)) was the most rapidly lytic population by 51chromium release. However, in the single-cell assay, the rate of lysis of bound target cells was quite similar for the LGL-enriched effector subpopulation and the higher density subpopulation of effector cells recognized previously. Both the light and dense effector cells contained similar numbers of target binding cells. Therefore, that the light subpopulation effected lysis more rapidly and to a greater extent than the dense subpopulation suggested that the low-density effector cells probably recycled more rapidly than those of higher density. This was corroborated by the finding that when conjugates were formed at 29 degrees C for the single-cell assay, a significant number of dead unconjugated targets could be observed only on the slides made with the LGL-enriched effector cells but not on those made with dense effector cell. Lysis continued to increase in the chromium-release assay probably because of recycling, recruitment, and/or heterogeneity of the effector cells, and/or because of heterogeneity or delayed death of the target cells.

Lymphokine-activated killer cells and aging in mice: significance for defining the precursor cell.

The present study was undertaken to define the cell populations which mediate lymphokine-activated killer (LAK) cell activity in mice. Because old mice exhibit markedly decreased to nondetectable natural killer (NK) cell activity, this age-associated change provided an advantageous system to examine the contribution of NK and T cells to LAK activity. Spleen cells from either young (6-9 weeks) or old (20-26 months) mice were cultured with 1000 units/ml of recombinant interleukin 2 (rIL 2) for 3-5 days. The cells were then tested in a 51 Cr-release assay for their cytotoxicity against NK-resistant fresh tumor cells (MCA-102). The LAK activity exhibited by spleen cells from old mice following 5 days of culture was equivalent to that developed by spleen cells of young mice. This result was contrary to what would be anticipated if mature NK cells comprise the primary precursors of LAK activity, and required further elucidation. The Thy-1 and asialo GM1 (ASGM1) phenotypes of LAK precursor and effector cells were therefore examined by depletion techniques using the appropriate antibodies plus complement. The results using spleen cells harvested after 5 days of culture with rIL 2 showed that LAK effector cells which developed from spleen cells of both young and old mice were predominantly Thy-1+ (85.3% young; 91.8% old) and some coexpressed ASGM1. Spleen cells were treated prior to culture to study the precursor cells. Development of LAK activity by spleen cells from both young and old mice was greatly reduced by pretreatment with anti-ASGM1 plus complement. However, since spleen cells of old mice exhibit very low mature NK activity, these data suggest that the LAK precursors, at least in old mice, may be ASGM1+ NK precursor cells rather than mature ASGM1+ NK effector cells. In addition, treatment with anti-Thy-1 plus complement inhibited generation of a significant proportion of LAK activity only in the spleens of old mice, suggesting a qualitative difference in LAK precursor cells with age and supporting the heterogeneity of the cells which are capable of developing LAK activity.

Suppression of murine natural killer cell activity by adherent cells from aging mice.

Natural killer (NK) cell activity declines with age in mice. The purpose of this study was to investigate the effect of peritoneal and splenic adherent cells from young and old mice on NK activity to determine whether adherent cell suppressor function might contribute to this decline. Peritoneal adherent cells from old mice suppressed NK activity of young splenic non-adherent indicator cells more than peritoneal cells from young mice. Splenic adherent cells from old but not from young mice also suppressed this activity. That (1) the suppressive activity of the adherent cell populations was not affected by treatment with anti-Thy-1 plus complement, and that (2) the adherent cell population contained 77-92% cells positive for alpha-naphthyl acetate esterase activity, suggests that the active adherent suppressor cell may be a macrophage. Therefore, the age-related decline in NK activity in mice can be explained, in part, by an increase in adherent cell suppressor function.

Immunological Responses of Aging Japanese A-Bomb Survivors

Immune response parameters were studied on 1341 A-bomb survivors residing in Hiroshima, Japan. Mononuclear cells were isolated from venous blood and tested for interleukin-2 production; lymphocytes were purified and tested for natural killer (NK) cell activity and interferon (IFN) production; and serum was tested for IFN and circulating immune complex (CIC) levels. Statistical analyses were performed for each type of assay using a linear models procedure including sex, age at the time of the bomb, radiation exposure, all the interaction variables, and the categorical variable day-of-assay in the model. The findings showed that (1) none of the immunologic variables were significantly affected by radiation exposure; (2) NK activity and CIC levels were positively associated with age; and (3) NK activity was on average higher for males than females. The data exemplify the difficulty in reaching firm conclusions concerning associations with radiation exposure when the dependent variable exhibits a large degree of interindividual and day-of-assay variability.