Action of hydrocortisone on the cytotoxic reactions in delayed hypersensitivity to microbial antigens

A study was made of the effect of hydrocortisone (HC) injected to animals with delayed hypersensitivity (DH) to BCG antigens on the cytotoxic activity of lymphocytes and production of lympho- and macrophage toxins. The cytotoxic test with the use of sensitized lymphocytes and preparation of lympho- and macrophage toxins were performed in vitro in the presence of specific microbial antigens. It was shown that HC exerts the most intense inhibitory action on the production of macrophage toxin. High doses of the hormone also inhibited the production of lymphotoxin. At the same time the cytotoxic activity of lymphocytes of the lymph nodes in DH was not inhibited by the employed doses of HC. No reduction was seen either in the sensitivity of autologous adhesive cells (macrophages) used as target cells for studying the cytotoxic activity of lymphocytes.     

A novel regulatory mechanism of naringenin through inhibition of T lymphocyte function in contact hypersensitivity suppression

Naringenin, a flavonoid in grapefruits and citrus fruits, has been reported to exhibit anti-inflammatory and anti-oxidative activities. Contact hypersensitivity (CHS) is a T cell-mediated immune reaction, and the factors released from macrophages also contribute to this response. Previous studies showed that naringenin suppressed CHS by inhibiting activation and migration of macrophages. However, little is known about naringenin’s effects on T lymphocytes. Our study indicated that naringenin potently suppressed picryl chloride (PCl)-induced contact hypersensitivity by inhibiting the proliferation and activation of T lymphocytes. In vitro, both of the activated hapten-specific T cells and the T cells stimulated with anti-CD3/anti-CD28 showed growth arrest after naringenin treatment. Furthermore, naringenin reduced CD69 (the protein level) and cytokines such as IL-2, TNF-α, and IFN-γ (the mRNA level) expressions which highly expressed by activated T cells. Meanwhile, naringenin also induced T cell apoptosis by upregulation of Bax, Bad, PARP, cleaved-caspase 3 and downregulation of phosphorylated Akt, Bcl-2. These findings suggest that, besides its anti-inflammatory activities in macrophages, naringenin also showed inhibitory effects on the activation and proliferation of T cells to alleviate symptoms of contact hypersensitivity.     

Desensitization V: Suppression of MIF production by lymphokine-activated macrophages

The systemic injection of high doses of antigen into a previously immunized animal results in a state of transient anergy with respect to cell-mediated immune reactions. This phenomenon is known as desensitization. We have previously shown that desensitization is a multistage process. The initial 24-hr period is characterized by excessive lymphokine production with a failure to express delayed hypersensitivity reactions due to abolition of local chemotactic gradients. Subsequent stages of desensitization involve failure of lymphokine production in vivo. The results presented here demonstrate that lymphocytes obtained from immunized and desensitized animals later than 24 hr after desensitization are markedly suppressed in their ability to produce MIF. In addition, it was found that lymphokine-activated macrophages can suppress in vitro MIF production by lymphocytes from immune, nondesensitized animals. In vitro and in vivo activation of macrophages were equally effective. Thus, it is likely that at least one mechanism for the inhibition of lymphokine production in the post-24-hr period of desensitization, involves activation of a population of suppressor macrophages by lymphokines produced during the initial 24-hr period.     

Immunomodulating properties of salmozan and its effect on the functional activity of macrophages

The effect of salmozan on the resistance of mice to Listeria monocytogenes infection, the formation of delayed hypersensitivity (DH) to sheep red blood cells in the animals, as well as changes in some functional activity characteristics of macrophages have been studied. The study has revealed that salmozan enhances anti-infectious resistance, suppresses the dermal manifestations of DH, and decreases the level of 5'-nucleotidase in peritoneal macrophages, stimulating their phagocytic activity. The intensity of the drug action depends on the time of its administration. The most pronounced immunomodulating action and maximal changes in the function of macrophages have been registered simultaneously after the treatment of the animals with salmozan.     

Macrophage-lymphocyte interaction in the formation of delayed-type hypersensitivity

Macrophage-lymphocyte interaction was studied on 121 CBA mice during a 2-hour contact of lymph-node cells of non-immune mice with a monolayer of peritoneal macrophages of BCG-immunized mice and subsequent intravenous administration of 4.10(7) pre-incubated lymphocytes to syngenic recipients. Sensitivity to tuberculin was demonstrated in the recipients by means of blast-transformation reaction of spleen cells (stimulation index was evaluated according to incorporation of 3H-thymidine--SI = 1.32 +/- 0.40) using administration of tuberculin into the paws (Mantoux reaction--MR = 0.11 +/- 0.02 mm) and the cytotoxic effect (CTE) of the lymphocytes on tuberculin-loaded sheep-cell erythrocytes whose disintegration was assessed according to discharge of iron from the target cells (CTE = 13.98 +/- 2.73%). At transfer of intact lymphocytes after contact with non-immune macrophages, SI = 1.046 +/- 0.019; MR = 0.014 +/- 0.002 mm; CTE = 0.214 +/- 0.048%. The treatment of lymphocytes with indomethacin during the contact with macrophages induced idvere changes in the indices of delayed-type hypersensitivity (DTHS). The conclusion has been drawn that the antigen-presenting ability of macrophages can materialize in vitro.     

The development of granulomatous pulmonary inflammation in rabbits by aerosol challenge: I. Release of plasminogen activator by alveolar macrophages

A rabbit model of hypersensitivity pneumonitis (HP) was employed to evaluate the release of plasminogen activator (PA) as a method for monitoring the degree of pulmonary inflammation. PA release from alveolar macrophages (AM) was shown to coincide with inflammation and was maximal at approximately 2 weeks of aerosol challenge. PA release could also be induced in normal AM by peripheral lymphocytes obtained from sensitized animals after incubation with antigen. Unseparated peripheral blood mononuclear cells from experimental animals also exhibited antigen-induced PA release. These results suggest that the measurement of PA release using several different cell populations can be used to evaluate pulmonary inflammation in HP.     

甲状腺激素对小鼠免疫功能的调节作用

正常昆明种成年小鼠每天给予一定量的甲状腺激素可使其抗体生成能力、淋巴细胞转化能力、迟发型超敏反应和巨噬细胞的吞噬功能明显增强,外周血成熟 T 细胞数量亦明显增加。而每天给予抗甲状腺药物则使其抗体生成能力、淋巴细胞转化能力和迟发型超敏反应明显降低。切除甲状腺亦使抗体生成能力下降。这些结果提示甲状腺激素参与免疫功能的生理性调节。     

Molecular and cellular analysis of histamine H1 receptors on cultured smooth muscle cells

Histamine is an important mediator of immediate hypersensitivity for both animals and humans. The action of histamine on target tissues is believed to be mediated by specific cell surface receptors, especially H1 and H2 receptors for hypersensitivity and inflammatory reactions, which involve stimulation of smooth muscle contractility, alterations in vascular permeability, and modifications in the activities of macrophages and lymphocytes. Although the nature of histamine receptors in the brain and peripheral tissues has been studied extensively by many laboratories, the molecular mechanism of histamine receptor-mediated reactions is not fully understood, mainly because histamine receptors are incompletely characterized from the biochemical point of view. In previous studies, we have found that the cultured smooth muscle cell line DDT1MF-2, derived from hamster vas deferens, expresses low-affinity histamine H1 receptors and responds biochemically and functionally to H1-specific stimulation (Mitsuhashi and Payan, J Cell Physiol 134:367, 1988). This cell line provides a model for analyzing the biochemical responses of H1 receptor-mediated reactions in peripheral tissues. In this review, we summarized our recent progress in the study of low-affinity H1 receptors on DDT1MF-2 cells.     

Activated lymphocytes depress phagocytosis of latex particles by human monocyte-macrophages

In this study, monocyte-macrophages of normal human donors were cultured with and without lymphocytes and antigen in order to define the effect of antigen-stimulated lymphocytes on phagocytosis by macrophages. Phagocytosis was assessed by uptake of radio-labeled latex particles by macrophage monolayers. Although no effect was noted when purified macrophage monolayers were cultured in the presence of antigen, marked inhibition of phagocytosis was observed when macrophages were cultured in the presence of autochthonous antigen-stimulated lymphocytes. The degree of phagocytic depression correlated with the delayed cutaneous hypersensitivity response of the donor and with the concentration of antigen present in the system.     

Guinea pig T-cell in vitro proliferative responses to tyrosine-azobenzenearsonate-pulsed and azobenzenearsonate-coupled macrophages have different specificities

The cell interactions involved in azobenzenearsonate-N-acetyl-tyrosine (ABA-tyr)-induced delayed hypersensitivity in the guinea pig were studied by in vitro blastogenesis. The ABA-sensitive lymphocyte was demonstrated to be a T lymphocyte and its presence in peritoneal exudate cells was shown to be much higher than spleen or lymph node populations. The secondary response of ABA-sensitized lymphocytes to ABA-tyr in culture is dependent on the presence of an accessory cell, with both splenic and peritoneal macrophages being equally effective. ABA coupled directly to macrophages as an immunogen induced strong responses to itself and not to ABA-tyr-pulsed macrophages or ABA-tyr in solution. The reverse was true in animals, immunized with ABA-tyr. ABA conjugated to thymocytes, L₂C leukemia cells, and guinea pig erythrocytes however, did not elicit significant responses. The results obtained in animals immunized with ABA- or ABA-tyr-modified cells was similar whether or not CFA was used. The difference in specificity shown between ABA-coupled and ABA-tyr-pulsed macrophages favors a single receptor hypothesis for T-cell recognition.     

Activity and Properties of Macrophage Migration Inhibitory Factor produced by Mixed Lymphocyte Cultures

THE macrophage migration test is an in vitro demonstration of delayed hypersensitivity. Supernatant fluids of sensitive lymphocytes cultured for 24 h in the presence of specific antigen contain migration inhibitory factor (MIF) that arrests the migration of macrophages of unsensitized animals in vitro¹,². In vivo, it induces delayed skin reactions³. The use of the macrophage migration test, based on differences of transplantation antigens in donor and recipient, to show histocompatibility has been suggested⁴. The test was also recommended as an indicator of immunological reactivity after organ transplantation, to demonstrate impending rejection⁵. It can demonstrate homograft sensitivity, for migration of peritoneal exudate cells (containing lymphocytes and macrophages) of CBA mice previously sensitized by grafts from A/Jax donors was inhibited when they were mixed with peritoneal exudate cells of the donor strain. However, histocompatibility was not demonstrated, for mixtures of peritoneal exudate cells of ungrafted CBA mice and A/Jax mice migrated regularly during the 24 h test⁶.     

Delayed Hypersensitivity in Relation to Suppression of Growth of Listeria monocytogenes by Guinea Pig Macrophages

Prototypes of delayed hypersensitivity (tuberculin allergy, graft rejection immunity, and contact dermatitis) were established in guinea pigs. The macrophages from peritoneal exudates of such animals were examined for their capacities to suppress the growth of Listeria monocytogenes in vitro. Only the macrophages from animals sensitized to BCG clearly exhibited this property.     

N-Acetylcysteine inactivates Migration Inhibitory Factor and Delayed Hypersensitivity Reactions

In vitro studies suggest that delayed hypersensitivity follows the production of migration inhibitory factor (MIF) by sensitive lymphocytes in the presence of specific antigen. This factor arrests the migration of macrophages in vitro and in vivo. After attraction, aggregation and activation in vivo, these bystander cells produce toxic substances which induce the local reaction¹. When lymphocytes from tuberculin (PPD) sensitized guinea-pigs were incubated with PPD, cell-free supernatant fluids of the cultures contained MIF². Such migration inhibitory fluids injected intradermally with PPD, into PPD-sensitive animals, enhanced the delayed hypersensitivity reaction³. Concentrated migration inhibitory supernatant fluids injected intradermally into unsensitized animals produced local reactions of induration and erythema within 6 h; reactions reached a maximum after 16 h. Histologically there was an infiltrate of mononuclear cells at the site of injection and neutrophils and eosinophils were also present¹.     

Cytotoxic action of immune lymphocytes on "adherent" cells (macrophages) of autologous system lymph nodes during delayed hypersensitivity]

A study was made in the autologous system of the cytotoxic action of immune lymphocytes on the "adherent" cells (macrophages) of the lymph nodes of guinea pigs in delayed hypersensitivity (DH) to the streptococcus antigens and tuberculoproteins. Death of a considerable number of the "adherent" cells in cultivation of a suspension of cells of the lymph nodes of animals sensitized with the culture of streptococcus or BCG in the presence of specific antigen (thermo-stable streptococcus fraction or tuberculin, respectively) alone. Detection of death of the "adherent" cells in the autologous system can be used as one of specific and sensitive tests in studying the DH.     

Stimulation of anaphylaxis in the mouse footpad by dietary fish oil fatty acids

The effect of fish oil-derived omega-3 (omega-3) fatty acids on anaphylaxis, Arthus and delayed type hypersensitivity reactions in mice has been investigated. Mice on a normal chow diet were fed eicosapentaenoic acid and docosahexaenoic acid at a dose of 500 and 333 mg/kg/day, respectively, by a gastric tube over a period of 61 days. Control groups were given water, safflower oil or oleic acid. Anaphylactic and Arthus type reactions were induced in the mouse footpad using bovine serum albumin as an antigen. Carrageenin was utilized to produce a delayed type hypersensitivity reaction. The animals fed omega-3 fatty acids induced a more anaphylactic foodpad reaction. There was no significant effect of the diet on Arthus and delayed type hypersensitivity responses. There was no effect of the fish oil-supplemented diet on production of antibodies to bovine serum albumin. Synthesis of prostaglandin E2 by peritoneal macrophages was significantly inhibited in the animals fed omega-3 fatty acid-enriched fish oil, while leukotriene B4 production was not affected. These results suggest that a diet enriched in omega-3 fatty acids modulates production of arachidonic acid metabolites and this may influence anaphylaxis, but not Arthus and cellular mediated hypersensitivity responses.     

Interactions between heterochronic cells in the formation of delayed-type hypersensitivity.

The effect of ageing on lymphocyte and macrophage functional activity in the induction of tuberculin sensitivity was studied in experiments on 440 CBA mice of three age groups. The capacity of old animals to generate delayed-type hypersensitivity following administration of BCG cells was found to be diminished and their lymphocytes caused decreased hypersensitivity on transplantation. The capacity of recipients to reproduce delayed-type hypersensitivity after thymocyte and bone marrow cell transplantation was influenced by the age of transplanted thymocytes. This capacity was markedly suppressed on the recipients of old cells. The antigen presentation function of macrophage did not after significantly as a function of age.     

Production of tumor necrosis factor in cells of hibernating ground squirrels Citellus undulatus during annual cycle

TNF production has been studied in peritoneal macrophages and splenic T cells of Arctic Yakutian ground squirrel (Citellus Undulatus Pallas) in hibernating and awake animals in winter and in prehibernating autumn as well as in active euthermic spring-summer animals. A high level of TNF production in macrophages of ground squirrel is observed over the active period and during arousals in winter. There are no significant season variations in TNF production in splenic T lymphocytes of ground squirrels. This suggests the major role of activated macrophages in the arousals of hibernating animals. T lymphocyte proliferation in ground squirrels in the active period is higher than in winter, and the most significant seasonal variations are found in T cell mitogenic response, which increases in spring-summer period. Evidence is presented that functional activity of macrophages of squirrel in autumn has much in common with that in winter rather than in spring-summer period.     

Delayed hypersensitivity and nonspecific cellular immunity. The role of mono- and polynuclear phagocytes

Differences in the influence produced by sensitization with BCG vaccine and Staphylococcus aureus and by the reaction of delayed hypersensitivity (DH) induced, respectively, by the injection of old tuberculin and staphylococcal phagolysate on the phagocytic activity of peritoneal macrophages and blood leukocytes in different animals were experimentally demonstrated. A considerable activation of the bactericidal and ingesting functions of macrophages was observed in animals showing a pronounced DH reaction (rabbits, guinea pigs and mice), while in Wistar rats no such activation was noted. The latter showed no DH reaction after sensitization with BCG vaccine and the injection of the specific antigen. Among different strains of mice, the activation of macrophages occurred in the animals with the most pronounced DH reaction. Sensitization with BCG vaccine led to an insignificant sensitization of macrophages, and sensitization with S. aureus even suppressed the phagocytic activity of macrophages. The treatment of mice with antimacrophagal preparations (carrageenan, silica and trypan blue, but T-lymphocyte antiserum) before and after the injection of the specific antigen into the sensitized animals abolished the stimulation of anti-infection immunity.     

In vitro delayed hypersensitivity granuloma formation: development of an antigen-coated bead model

Previously, we have described an in vitro model of granulomatous hypersensitivity around Schistosoma mansoni eggs in both the murine model of schistosomiasis and in human schistosomiasis. These studies describe a new model of in vitro granuloma formation that complexes soluble egg antigen from S. mansoni eggs, a partially purified protein derivative of Mycobacterium tuberculosis (PPD), or bovine serum albumin to carrier beads. Ultrastructural and morphologic evaluations demonstrate that there are initial macrophage interactions, followed by the recruitment of antigen-specific T cells that interact with and recruit macrophages, lymphocytes, granulocytes, and fibroblasts. Finally, there is a stage of granulomatous organization involving fibroblast proliferation and collagen deposition. The in vitro reactivity, defined by a quantitative granuloma index, correlates with in vivo granulomas around S. mansoni eggs in the livers of infected cell donor animals. In vitro granuloma formation against PPD-coated beads correlated with delayed cutaneous hypersensitivity against PPD, which was judged by footpad swelling. The reactions demonstrate antigenic specificity and were intrinsically modulated in a manner that is analogous to that previously shown with the in vitro egg granuloma model. This model of in vitro granuloma formation promises to be a useful tool for elucidating mechanisms of cellular immunity and regulation.     

Macrophage infiltration in tumors and tumor-surrounding tissue: influence of serotonin and sensitized lymphocytes

Summary In a delayed-type hypersensitivity reaction serotonin released from mast cells plays an important role in the induction of a cellular infiltrate at the site of antigen challenge. In analogy, we have studied whether it is possible to enhance the number of intratumoral macrophages by injecting serotonin into a s.c. SL2 lymphosarcoma. The vessels in the tissue surrounding the tumor responded well to serotonin, as there was an influx of i.v. injected ⁵¹Cr-labeled sensitized spleen cells in this tissue during the first 4 h after intratumoral injection of serotonin. At 24 h after serotonin injection there was an influx of macrophages into this tumor-surrounding tissue. No influx of cells was detected in the tumor itself during the first hours after injection of serotonin. In the tumor, similar phenomena occurred as in the surrounding tissue, but with a delay of about 24 h. This suggests that lymphocytes leave the blood circulation in the tumor-surrounding tissue and migrate to the tumor. The influx of macrophages into the tumor after intratumoral injection of serotonin is probably due to an immunological reaction as the lymphocyte influx preceeds the macrophage influx into tumors. In addition, transfer of sensitized lymphocytes, as well as lymphocytes from a tumor-bearing host caused an enhanced influx of macrophages into the tumor. To test the specificity and serotonin dependency of the phenomenon of infiltrating cells in tumors we have used a footpad swelling assay in which the serotonin dependency and the antigen specificity of the response against syngeneic tumor cells was shown. The following picture emerged: an intratumoral serotonin injection enables lymphocytes to leave blood vessels in the tumor-surrounding tissue. These lymphocytes with specificity for tumor antigens migrate to the tumor. After contact with the antigenic tumor cells, these lymphocytes secrete chemoattractive factors for monocytes/macrophages. Also these monocytes/macrophages leave the circulation in the tumor-surrounding tissue. Subsequently the macrophages invade the tumor. We conclude that the number of intratumoral macrophages can be enhanced by serotonin.