The Effect of Lasso® Herbicide on Human Immune Function as Measured by In Vitro Assays

Abstract

Using in vitro assays, this study was undertaken to determine whether the components of Lasso® herbicide formulation had an effect on the human immune system. Mononuclear cells from human peripheral blood were exposed to analytical alachlor, alachlor conjugated to human serum albumin or Lasso formulation over a concentration range from. 01 uM-1.0 uM. The effects of the test materials on the following immunological functions were determined: lymphocyte proliferation induced by mitogen or antigen; antibody synthesis of IgG and IgM isotypes in pokeweed stimulated monouclear cell cultures; cytotoxic T cell proliferation; lysis of target cells by natural killer cells and lymphokine activated killer cells. The data demonstrated that the test compounds had no significant, dose related effect on the function of immunocompetent cells. Hence, the data suggest that the components of the Lasso formulation have no effect on the human immune system.     

Immunological Consequences of In Vitro Exposure to Lysergic Acid Diethylamide (LSD)

The ability of lysergic acid diethylamide (LSD) to alter immune function after direct in vitro exposure was examined. It was demonstrated that LSD is able to suppress the proliferation of B-lymphocytes; the production of the cytokines IL-2, IL-4, and IL-6; and the induction of cytotoxic T-lymphocytes at a concentration of 100 μM. In vitro exposure to LSD had differential effects on natural killer (NK) cell activity, with significant enhancement of both basal and IL-2-augmented NK cell function at concentrations between 0.0001 and 0.1 μM, and suppression of NK response at 100 μM. These results demonstrate that LSD may have a direct effect on components of the immune system at concentrations that may be reached upon human exposure.     

Cell wall of Candida albicans and host response

Modulation in chemistry and organization of cell wall macromolecules play decisive roles in the morphogenic processes and virulence of Candida albicans. Cell wall components also have a diversified range of effects on the host's immune system, including immunopotentiating or immunodepressing activities. Mannan, mannan-protein, and glucan fractions have been especially studied in this context. In in vitro cultured human peripheral blood mononuclear cells, a mannan-protein fraction (GMP) from the cell wall of the yeast form acted as a strong antigenic activator by stimulating lymphokine production and lymphocyte proliferation. Cytolytic effectors active against several tumor targets were also generated. In the mouse, GMP was a strong inducer of natural killer lymphocytes. Other cell wall components, mostly the insoluble beta-glucan, modulated the activity of macrophages and monocyte precursors. Some of the immunomodulating properties of artificially extracted components were shared, even with greater potency, by antigens which were released from C. albicans during its growth and hyphal morphogenesis. Altogether, the range of the immunoresponses elicited and the intensity of the observed effects are such as to individuate in this human indigenous fungus a microorganism capable of profoundly affecting the host's immune system.     

Effect of morphine and beta-endorphin on human Fc receptor-dependent and natural killer cell functions.

Interactions between opiates and the human immune system have important clinical implications. To further evaluate these interactions, we studied in vitro and in vivo effects of morphine sulfate (morphine) and beta-endorphin (Bend) on antibody-dependent cell cytotoxicity (ADCC), natural killer cell cytotoxicity (NKCC), and effector cell expression of antibody Fc receptors. Morphine and Bend had no potent in vivo or in vitro effect on FcR expression nor did they have a significant in vitro effect on ADCC by monocytes or polymorphonuclear cells. Bend enhancement of NKCC in vitro was inhibited by coincubation of effector cells with morphine. After taking 90 to 150 mg of oral morphine, study volunteers demonstrated a significant decrease in ADCC by peripheral blood mononuclear cells. The same individuals demonstrated a consistent increase in NKCC and no change in the expression of Fc receptors. Effector cells from these individuals responded normally to in vitro incubation with interferon-gamma (IFN-gamma).     

Inhibition of Natural Killer Cell Activity by Antigen-Antibody Complexes

The natural killer (NK) cell activity of human peripheral blood mononuclear cells was found to be inhibited by precipitated tetanus toxoid anti-tetanus toxoid complexes (Te/aTe) as well as soluble Te/aTe. Preincubation of the immune complexes with protein A decreased the inhibition of NK cell activity. When mononuclear cells were preincubated with interferon (IF) or interleukin 2 (Il-2) before incubation with Te/aTe, the immune complex-induced inhibition was decreased, while IF or Il-2 added after incubation with the immune complexes had no effect. Using NK cell-enriched suspensions in a single cell agarose assay, the immune complexes were shown to inhibit NK cell activity by inhibiting the formation of effector/target cell conjugates.     

Complement inhibits immune responses: C3 preparations inhibit the generation of human cytotoxic T lymphocytes

C3 fragments have been shown to inhibit mitogen- and antigen-induced human lymphocyte blastogenesis. In this study, C3 preparations and a small fragment of C3 (contained in a preparation called Fraction 2) were examined for their capacity to regulate cytotoxic T lymphocytes (CTL) and proliferative mixed lymphocyte responses (MLR). We found that both preparations inhibited the generation of allogeneic human CTL as well as MLR in a dose-related manner. In contrast, Fraction 1, which contained native C3 and C3b, did not inhibit the generation of CTL nor did it inhibit the MLR. The kinetics of inhibition of proliferation were divergent from the kinetics of inhibition of CTL generation; the active preparations inhibited proliferation significantly when added as late as day 5 of a 6-day culture, whereas no inhibition of CTL generation was seen when these preparations were added after day 3 of culture. Cultures in which C3 preparations caused complete inhibition of CTL generation had normal levels of the nonspecific, anomalous killer cell activity, as assayed on K 562 target cells. Furthermore, C3 preparations and Fraction 2 had no effect on the lytic function of differentiated CTL, on “spontaneous” natural killer cell activity or on interferon-induced augmentation of natural killer cell activity. These findings indicate that C3 fragments may play a negative role in the regulation of CTL responses.     

Endogenous transforming growth factor-beta inhibits toll-like receptor mediated activation of human uterine natural killer cells

PROBLEM: Toll-like receptors (TLRs) recognition is an important means for the innate immune system to rapidly respond to pathogen invasion. Our aim was to determine whether uterine natural killer (uNK) cell cytokine production induced by stimulation through TLRs could be regulated by endogenous transforming growth factor (TGF)-beta in human endometrium. METHOD OF STUDY: Single cells were isolated from human endometrium, and interferon (IFN)-gamma production by endometrium cells and uNK cells was determined after stimulation by TLR agonists. The role of TGF-beta in regulating this response was tested by blocking TGF-beta function using antibodies or a specific inhibitor, SB431542. RESULTS: TGF-beta blockade increased TLR agonist induced IFN-gamma by uNK cells. The regulation of uNK cell cytokine production was observed when uNK cells were incubated with agonists for TLR2 (PGN) or TLR3 (polyI:C). Blockade of TGF-beta or TGF-beta receptor signaling had no effect on constitutive cytokine production in the absence of TLR agonists. CONCLUSION: The results indicate that endogenous TGF-beta alters cytokine responses of uNK cells in human endometrium in response to TLR agonists. These data suggest that uNK cell responses to microbial pathogens in the endometrium are regulated by the amount of biologically active TGF-beta present within the human endometrium.     

Cytogenetic effects of alachlor and/or atrazine in vivo and in vitro.

The purpose of this study was to assess the cytogenetic effects of two commonly used herbicides, alachlor and atrazine, which are often found together in groundwater. Chromosome damage was examined in bone marrow cells of mice drinking water containing 20 ppm alachlor and/or 20 ppm atrazine, with an immunosuppressive dose of cyclophosphamide used as a positive control. Chromosome damage was also quantified in human lymphocytes exposed in culture to 1.0, 0.1, or 0.01 microgram/ml alachlor and/or atrazine. The in vitro study demonstrated dose related cytogenetic damage not associated with mitotic inhibition or cell death, with damage due to the alachlor-atrazine combination suggesting an additive model. The in vivo study also suggested additive damage due to the alachloratrazine combination after 30 days of treatment, but, unexpectedly, demonstrated less cytogenetic damage and fewer cells with multiple aberrations after 90 days. Also, at 90 days, all treated mice had elevated mitotic indices compared to controls. The fact that the elevated mitotic index was associated with immune suppression in the cyclophosphamide group suggests that death of cells with accumulated chromosomal aberrations resulted in increased bone marrow proliferation, so a higher fraction of cells examined were newer with less damage. Since the alachlor-atrazine combination treated mice showed little systemic toxicity despite bone marrow mitotic indices similar to the cyclophosphamide treated animals, as well as a similar decrease in cytogenetic damage at 90 days compared to 30 days, cell death and replacement must also be involved but cannot completely explain the results.     

Dietary Curcumin Enhances Antibody Response in Rats

The effects of dietary curcumin on three major types of immune function were examined in rats. Antibody (IgG) production, delayed-type hypersensitivity and natural killer cell activity were evaluated after 5 weeks of dietary exposure to 1, 20 or 40 mg/kg curcumin. The highest dose of curcumin significantly enhanced IgG levels. Rats receiving lower dietary concentrations (1 or 20 mg/kg) of curcumin were not different in IgG production from rats receiving no curcumin in their diet. Neither delayed-type hypersensitivity nor natural killer cell activity was different from control values at any dietary concentration of curcumin. In vitro incubation of YAC-1 and EL4 tumor cells and normal splenocytes in varying concentrations of curcumin for varying times revealed differences between cell types in curcumin's effects on cell proliferation and viability. No cytotoxic effect was seen in EL4 cells at 12S μg/ml curcumin at 4, 24 and 48 hrs incubations, however, cell proliferation was reduced by almost 50 % at 24 hrs. YAC-1 cell viability and cell numbers were diminished at longer incubations. A lower curcumin concentration (1.25 μg/ml) enhanced cell growth in the YAC-1 cells at 24 and 48 hr. This enhancement was not seen in spleen or EL4 cells.     

Homeostatic migration and distribution of innate immune cells in primary and secondary lymphoid organs with ageing

Ageing of the innate and adaptive immune system, collectively termed immune senescence, is a complex process. One method to understand the components of ageing involves dissociating the effects of ageing on the cells of the immune system, on the microenvironment in lymphoid organs and tissues where immune cells reside and on the circulating factors that interact with both immune cells and their microenvironment. Heterochronic parabiosis, a surgical union of two organisms of disparate ages, is ideal for this type of study, as it has the power to dissociate the age of the cell and the age of the microenvironment into which the cell resides or is migrating. So far, however, it has been used sparingly to study immune ageing. Here we review the limited literature on homeostatic innate immune cell trafficking in ageing in the absence of chronic inflammation. We also review our own recent data on trafficking of innate immune subsets between primary and secondary lymphoid organs in heterochronic parabiosis. We found no systemic bias in retention or acceptance of neutrophils, macrophages, dendritic cells or natural killer cells with ageing in primary and secondary lymphoid organs. We conclude that these four innate immune cell types migrate to and populate lymphoid organs (peripheral lymph nodes, spleen and bone marrow), regardless of their own age and of the age of lymphoid organs.     

In Vitro Effects of Various Metals on Natural Killer Cell Activity in Cultured Human Lymphocytes

Abstract

Heavy metals have been shown to have a differential effects on various aspects of immune response. Recently natural killer cells have been widely investigated due to their purported role in immune surveillance. To ascertain the immunotoxic effects of lead, cadmium, nickel and chromium on natural killer (NK) cell activity in vitro, peripheral blood lymphocytes from normal donors were examined in the presence of different concentrations (10^?5-10^?8 M) of four selected metal salts (cadmium sulphate, lead nitrate, chromium nitrate and nickel sulphate). NK cell activity was evaluated in a 4-h chromium release assay against K562 target cells. All of the metal salts were found to exert no effect on NK cell function in the human concentration range.     

Role of cytokines and chemokines in the regulation of innate immunity and HIV infection

The earliest defense against microbial infection is represented by the responses of the innate (or natural) immune system, that also profoundly regulates the adaptive (or acquired) T- and B-cell immune responses. Activation of the innate immune system is primed by microbial invasion in response to conserved structures present in large groups of microorganisms (LPS, peptidoglycan, double-stranded RNA), and is finely tuned by different cell types (including dendritic cells, macrophages, natural killer cells, natural killer T cells, and gammadelta T cells). In addition, several soluble factors (complement components, defensins, mannose-binding lectins, interferons, cytokines and chemokines) can play a major role in the regulation of both the innate and adaptive immunity. In this review, we will briefly overview the regulation of some cellular subsets of the innate immune system particularly involved in human immunodeficiency virus (HIV) infection and then focus our attention on those cytokines and chemokines whose levels of expression are more profoundly affected by HIV infection and that, conversely, can modulate virus infection and replication.     

Differential effect of transforming growth factor beta on the synthesis of Th1- and Th2-like lymphokines by human T lymphocytes.

(TGF)-beta is a pluripotent cytokine exerting differential effects on distinct components of the immune response. The present report, based on lymphokine determination in culture supernatants and Northern blot analysis of lymphokine mRNA, demonstrates that TGF-beta 2 markedly inhibits interleukin (IL)-4 and IL-5 synthesis by polyclonally activated human T cells in the absence of any significant effect on (IFN)-gamma, lymphotoxin or IL-2, suggesting a modulatory effect of TGF-beta 2 on the interferon Th1/Th2) balance of immune responses. The inhibitory effect of TGF-beta on IFN-gamma production by unfractionated peripheral blood mononuclear cells is likely to reflect the blunting of natural killer cell activation by TGF-beta.     

Effect of human endometrial stromal cell-derived conditioned medium on uterine natural killer (uNK) cells' proliferation and cytotoxicity

Citation Chen Y, Zhuang Y, Chen X, Huang L. Effect of human endometrial stromal cell‐derived conditioned medium on uterine natural killer (uNK) cells’ proliferation and cytotoxicity. Am J Reprod Immunol 2011; 65: 589–596 Problem Human endometrial stromal cells are involved in the regulation of immune cell proliferation, apoptosis, differentiation, and function. In the endometrium, uNK cells are in close contact with stromal cells. The aim of the study was to investigate the effects of human endometrial stromal cells on uNK‐cell proliferation and uNK‐cell cytotoxicity. Method of study The conditioned medium was derived from the endometrial stromal cells in the proliferative phase, secretory phase, and early pregnancy. The effects of stromal cell‐derived conditioned medium on uNK‐cell proliferation and cytotoxicity were detected by mitochondrial lactate dehydrogenase‐based MTS staining and flow cytometry. Results The stromal cell‐derived conditioned medium in both secretory phase and early pregnancy significantly promoted uNK‐cell proliferation. Compared with the control group, the uNK‐cell cytotoxicity were significantly reduced by conditioned medium in the proliferative, secretory, and decidua groups, but there were no significant differences among these different physiological stages in the inhibiting ability. Conclusion Human endometrial stromal cells may be involved in the regulation of uNK‐cell functions through influencing proliferation and cytolytic activity.     

Cell walls, peptidoglycans, and teichoic acids of Gram-positive bacteria as polyclonal inducers and immunomodulators of proliferative and lymphokine responses of human B and T lymphocytes.

In this study the mitogenic and immunomodulating effects of bacterial cell wall preparations were investigated. Cell walls, peptidoglycans, and teichoic acids from Bacillus subtilis and Staphylococcus aureus Wood 45 activated both human T cells (supplemented with 10% monocytes) and B cells to proliferate and to produce leukocyte-inhibitory factor. Similar results were obtained with adult and umbilical cord blood cells, suggesting that these bacterial preparations were acting as mitogens. Cell walls and peptidoglycans had a modulating effect on purified protein derivative- or protein A-induced proliferation. In the presence of suboptimal concentrations of these stimulants, bacterial components enhanced the proliferative response. However, at optimal concentrations of purified protein derivative or protein A, bacterial components suppressed lymphocyte proliferation. Peptidoglycans solubilized by lysozyme activated B lymphocytes but not T cells. Solubilization had no effect on the immunomodulating capacity.     

Immunomodulatory effects of curcumin

Curcumin (diferuloylmethane), found in the spice turmeric, exhibits anti-inflammatory, antioxidant, and chemopreventive activities. However, the effect of curcumin on the immunological responses largely remains unknown. In this study we have investigated the effect of curcumin on mitogen (phytohaemagglutinin; PHA) stimulated T-cell proliferation, natural killer (NK) cell cytotoxicity, production of cytokines by human peripheral blood mononuclear cells (PBMCs), nitric oxide (NO) production in mouse macrophage cells, RAW-264.7. Furthermore, we have carried out an electromobility shift assay to elucidate the mechanism of action of curcumin at DNA protein interaction level. We observed that curcumin inhibits PHA-induced T-cell proliferation, interleukin-2 production, NO generation, and lipopolysachharide-induced nuclear factor-κB (NF-κB) and augments NK cell cytotoxicity. Our results suggest that curcumin most likely inhibits cell proliferation and cytokine production by inhibiting NF-κB target genes involved in the induction of these immune parameters.     

Simian immunodeficiency virus (SIV)/immunoglobulin G immune complexes in SIV-infected macaques block detection of CD16 but not cytolytic activity of natural killer cells

Natural killer cells are components of the innate immune system that play an important role in eliminating viruses and malignant cells. Using simian immunodeficiency virus (SIV) infection of macaques as a model, flow cytometry revealed a gradual loss of CD16+ NK cell numbers that was associated with disease progression. Of note, the apparent loss of NK cells was detected in whole-blood samples but not in isolated peripheral blood mononuclear cells (PBMC), suggesting that an inhibitor(s) of the antibody used to detect CD16, the low-affinity immunoglobulin G (IgG) receptor, was present in blood but was removed during PBMC isolation. (Actual decreases in CD16+ cell numbers in PBMC generally were not detected until animals became lymphopenic.) The putative decrease in CD16+ cell numbers in whole blood correlated with increasing SIV-specific antibody titers and levels of plasma virion RNA. With the addition of increasing amounts of plasma from progressor, but not nonprogressor, macaques to PBMC from an uninfected animal, the apparent percentage of CD16+ cells and the mean fluorescence intensity of antibodies binding to CD16 declined proportionately. A similar decrease was observed with the addition of monomeric IgG (mIgG) and IgG immune complexes (IgG-ICs) purified from the inhibitory plasma samples; some of the ICs contained SIV p27(gag) antigen and/or virions. Of interest, addition of purified IgG/IgG-ICs to NK cell lytic assays did not inhibit killing of K562 cells. These results indicate that during progressive SIV and, by inference, human immunodeficiency virus disease, CD16+ NK cell numbers can be underestimated, or the cells not detected at all, when one is using a whole-blood fluorescence-activated cell sorter assay and a fluorochrome-labeled antibody that can be blocked by mIgG or IgG-ICs. Although this blocking had no apparent effect on NK cell activity in vitro, the in vivo effects are unknown.     

Natural killer cells and exercise training in the elderly: a review.

Consistent reports of the positive relationship between regular physical activity and immunosenescence have generated much excitement in the field of exercise immunology. It is generally accepted that natural killer (NK) cell activity per NK cell decreases with age; decreases in NKCA have been associated with infection and death in the aged. The effects of exercise and training on natural killer cells, components of the innate immune system, have been studied extensively in young people. However, the published research on the elderly population is limited. Generally it has been found that training increases or does not change natural killer cell activity or counts in the elderly. The clinical relevance of these results is yet to be fully explored. In addition, the limitations of these studies on immune function have been many, and studies are often difficult to compare due to differences in their methods and presentation of results.     

The interactions between human dendritic cells and microbes; possible clinical applications of dendritic cells

The dendritic cells comprise several subsets that induce and regulate the immune responses against foreign and self-antigens, and that can therefore function as initiators of protective immunity and inducers of central or peripheral tolerance. The different subpopulations of dendritic cells interact with and also influence other cell populations of the immune system, such as T and B lymphocytes and natural killer cells. The factors that determine the given dendritic cell functions depend on the state of maturation and the local microenvironment. The interactions between dendritic cells and microorganisms are rather complex, but progress in the past few years has shed light on several aspects of these interactions. This review lays emphasis on the interactions between human dendritic cells, important components of the intima of arterial specimens at areas predisposed to atherosclerotic lesions, and Chlamydia pneumoniae and cytomegalovirus, the human pathogens most strongly implicated in the development of atherosclerosis. In addition, several examples of the potential clinical applications of dendritic cells are described.Received 13 January 2004; accepted by A. Falus 22 March 2004