The effects of the immunologic release of histamine upon human lung cyclic nucleotide levels and prostaglandin generation.

The effect of the antigen-induced, immunoglobulin (Ig)E-dependent release of mediators from human lung tissue was analyzed for coincident changes in the tissue levels of cyclic nucleotides. Simultaneously with the appearance of mediators, lung cyclic guanosine 3',5'-monophosphate (GMP) increased from 0.9+/-0.2 to 12.63+/-4.5 pmol/mg protein and cyclic AMP increased threefold from the initial levels of 5.1+/-1.4 pmol/mg protein. The release of histamine and prostaglandin (PG)F(2alpha), as well as the associated increases in cyclic nucleotides, peaked within 10 min of anaphylaxis. Antagonists of histamine's H-1 receptor prevented anaphylaxis-associated increases in cyclic GMP, whereas H-2 antagonists prevented the cyclic AMP response. Neither of these antagonists influenced the pattern or quantity of histamine or slow-reacting substance of anaphylaxis release. Prevention of PGF(2alpha) synthesis with acetylsalicylic acid failed to influence histamine or slow-reacting substance of anaphylaxis release or the concomitant increases in cyclic nucleotides. Histamine, added exogenously, produced a prompt increase in the cyclic AMP and cyclic GMP levels of human lung. As was seen after anaphylaxis, H-1 anatagonists prevented the cyclic GMP response to histamine, whereas H-2 antagonists prevented the cyclic AMP response.H-1 antagonists prevented 50% of the PGF(2alpha) synthesis accompanying anaphylaxis; H-2 antagonists had no effect. Exogenous histamine induced PGF(2alpha) synthesis; this synthesis was prevented by H-1 but not H-2 antagonists, and was reproduced by 2-methylhistamine (H-1 agonist) but not by dimaprit (H-2 agonist). Arachidonic acid generation of PGF(2alpha) was not influenced by antihistamines. Therefore, histamine interactions with human lung result in the synthesis of both PGF(2alpha) and cyclic GMP in response to H-1 stimulation, and of cyclic AMP through H-2 stimulation.

A monoclonal antibody against the sulfidopeptide leukotrienes LTC4, LTD4 and LTE4.

A monoclonal antibody (1A-LDR1) against sulfidopeptide leukotrienes (LT) is described. The mAb shows a nearly identical detection limit of about 0.04 ng for LTC4, LTD4, LTE4 and NacLTE4 in standard fluid phase RIA. Steric modifications, however, diminish the sensitivity, as determined for the examples 5-epi-LTC4, 6-epi-LTC4, 5,6-epi-LTC4 and 11-trans-LTC4. No crossreactivity could be observed for LTB4. Crossreactions with components of the LT peptide chain such as L-cysteine or glutathione, as well as with arachidonic acid, were not detectable. In assessing the accuracy of the LT-RIA, recovery experiments with supernatants of mouse peritoneal macrophages and incubates of gastric mucosa showed a good correlation of r = 0.993 and 0.990, respectively. Results of an inhibition experiment with mouse peritoneal macrophages, incubated with several concentrations of indomethacin and nordihydroguaiaretic acid (NDGA), support the reliability of RIA and ELISA. The new LT-mAB allows an almost complete detection of peptide leukotrienes in one assay.

The diversity of mast cell-derived mediators: implications for acute, subacute, and chronic cutaneous inflammatory disorders.

The mast cell in tissues represents an effector cell capable of elaboration of all the essential mediators of inflammation. The effects of uncontrolled activation may be divided into pharmacologic and inflammatory phases with attendant implications for the initiation of both acute and subacute pathologic processes. The elaboration of chemical mediators by the mast cell makes it possible to recruit blood cells and proteins essential to host defense by a controlled physiologic process that can proceed without significant local tissue damage. When uncontrolled, the same potentiality can be injurious, with the nature of the clinical problem depending upon the location of the cells, the intensity of activation, and the ratio of newly generated and preformed mediators released. The evidence that the mast cell can participate in each form of immunologic reaction--immediate, immune complex, and delayed- as a primary or secondary effector cell and the diversity of its products foretell an evolving recognition of its role in host defense and tissue injury. It is pertinent to develop further methods and criteria to define the nature and extent of mast cell participation in disease processes.

A sensitive and specific radioimmunoassay for leukotriene C4.

A sensitive and specific radioimmunoassay suitable for direct measurement of leukotriene C4 was developed. Acetylated leukotriene C4 was coupled to polyamino bovine serum albumin using 1-ethyl-3-(3-dimethylamino-propyl) carbodiimide hydrochloride as coupling agent. The conjugate in complete or incomplete Freund's adjuvant was injected into New Zealand white rabbits. At a final antiplasma dilution of 1 : 1050 the lowest detection limit of leukotriene C4 was 0.046 pmol. The antiplasma cross-reacted less than 1% with leukotrienes D4, E4 and F4, while high relative cross-reaction (86-100%) was obtained with leukotrienes C3, C5 and 11-trans leukotriene C4. In experiments where known amounts of leukotriene C4 were added to leukocyte suspensions, 67-100% of the added amount was recovered by the method. The radioimmunoassay was used to study leukotriene C4 formation after stimulation of leukocyte suspensions with the ionophore A23187.

Eosinophilic gastroenteritis--a complex disease entity.

Although usually regarded as merely a manifestation of a simple food allergy, eosinophilic gastroenteritis remains a poorly understood disorder. Characterized by peripheral eosinophilia, eosinophilic infiltration of the bowel wall to a variable depth and gastrointestinal complaints, the disease responds inconsistently to simple food withdrawal programs. Immunoglobulin E (IgE) antibodies to specific food substances have been found in a few patients leading to a postulated pathophysiologic mechanism involving tissue mast cells, release of slow-reacting substance of anaphylaxis (SRS-A) and chemotaxis of eosinophils. Oral steroids appear, in uncontrolled trials, to ameliorate the disease.

Lipid mediators of inflammation.

Many mediators of inflammation are derived from phospholipids and polyunsaturated fatty acids, including prostaglandins, leukotrienes, and platelet-activating factor. These mediators augment the vascular phase of inflammation and modify functions of inflammatory cells and cells of the immune system. Several growth factors or cytokines augment production of prostaglandin synthesis. The production of lipid-derived mediators may be inhibited by anti-inflammatory drugs and by modifying dietary polyunsaturated fatty acids.

Helminth-induced intestinal inflammation.

Gastrointestinal inflammation is a prominent feature of protective reactions in animals immune against helminths. Infiltration into the inflamed mucosa of various cells and their subsequent activation result in the elaboration of an array of pharmacologically and biologically active substances. The release of mediators is also associated with alterations in the epithelial layer. Furthermore, increased smooth muscle reactivity and enhanced secretory function of the mucosal tissue contribute to the development of an unfavourable environment and lead to worm expulsion. Mediators elaborated from inflammatory cells, whether associated with cell granules (i.e., preformed) or de novo-generated from membrane phospholipids, possess a number of potent vasoactive and spasmogenic properties which may contribute to events leading to worm elimination. The lipoxygenase metabolites of arachidonic acid (leukotrienes) derived from cell membranes probably contribute to the state of intestinal hypersensitivity against helminths. The measurement of elevated levels of these lipid mediators following worm challenge of immune, but not control, rats suggests that leukotrienes may play a role in amplifying and augmenting the inflammatory process associated with worm expulsion.

Immunoaffinity purification of prostaglandin E2 and leukotriene C4 prior to radioimmunoassay: application to human synovial fluid.

When human synovial fluid as such was subjected to radioimmunoassays of prostaglandin E2 (PGE2) and leukotriene C4 (LTC4), there was no linear increase in PGE2 and LTC4 as the amount of synovial fluid was raised. For removal of substances thus disturbing the assay we developed a method of immunoaffinity purification of PGE2 and LTC4. A monoclonal antibody against PGE2 or LTC4 was coupled to BrCN-activated Sepharose 4B. When synovial fluid mixed with radiolabelled PGE2 or LTC4 was applied to the column of immobilized antibody, the ligand was adsorbed to the column and eluted with a mixture of methanol/water in a recovery of about 80%. The purified material showed a linearity between the amount of the sample and the value of radioimmunoassay. The one-step method was applied to synovial fluid from patients with rheumatoid arthritis, osteoarthritis and other joint diseases.

Allergic mucociliary dysfunction.

Allergic rhinitis is typically associated with excessive nasal secretions and morphologic changes of the nasal mucosa; however, an impairment of mucociliary transport, the ultimate expression of mucociliary function, has so far not been clearly demonstrated. Tracheal mucociliary transport rates are decreased in patients with allergic asthma, and inhalation challenge with antigen causes a further impairment that appears to be related to the liberation of chemical mediators, notably slow reacting substance of anaphylaxis. Physiologic studies of mucociliary function in the nose similar to those that have been reported for the lower airways will be required to assess the role of mucociliary dysfunction in allergic rhinitis.

Mast cell-mediated reactions of host defense and tissue injury: the regulatory role of eosinophil polymorphonuclear leukocytes.

Immunological stimulation of mast cells, by way of either IgE- or IgG-directed reactions, initiates the rapid release of an array of chemical mediators. The predominant local tissue effects of these mediators collectively constitute a defensive response of the host. The early humoral phase of defense is exemplified by the alterations in microvascular permeability induced by histamine which provide a local concentration of immunoglobulins and complement components. The later cellular phase of defense is composed of the PMN leukocytes that accumulate in response to mast cell-derived chemotactic principles and which phagocytose and degrade opsonized foreign material, thus eliminating the inciting stimulus. Of the several endogenous regulatory mechanisms which act to contain the immediate hypersensitivity reaction, the eosinophil has a special role since it is specifically attracted to sites of mast cell activation and has selective concentrations of several enzymes which degrade the mast cell-derived chemical mediators. Failure of the local regulatory processes can permit the mast cell responses of host defense to become pathological reactions leading to tissue injury by virtue of persistence of high levels of humoral mediators and/or increasing infiltration with PMN leukocytes.

A rapid, simple method for calculating equilibrium constants and antibody site concentrations from dilution curves alone.

A rapid new practical method for calculating both the antibody-antigen equilibrium constant and the antibody concentration from antibody dilution curve data alone is described. This method is faster than the inhibition curve method for evaluating a humoral immune response. It is particularly suitable for monitoring the immune response of an immunization program. The response is assessed as an immunization index, Abi*Ka. This index is more exact than the antibody titer obtained from dilution curves and independent of the specific activity of the labelled molecule and total activity used in the assay. The method was used to monitor the production of a monoclonal antibody to the sulphide peptide leucotriene including immunization, cloning and purification.

The role of acupuncture in asthma: changes in airways dynamics and LTC4 induced LAI.

A cohort of nine extrinsic asthma patients were treated by means of acupuncture. Patients were followed up for changes in medical treatment, spirometry, skin reactivity to immediate type reactions, total serum IgE levels and reactivity of their leukocytes to leukotriene C4 challenge (LTC4 induced leukocyte adherence inhibition (LAI) assay). Our results show that after acupuncture, treated patients were able to reduce bronchodilator and taper completely corticosteroid therapy. No change in skin reactivity or in IgE levels were noted. However, acupuncture treatment was able to negate, in 66.6%, the positive LTC4 induced responses.

Roflumilast inhibition of pulmonary leukotriene production and bronchoconstriction in ovalbumin-sensitized and -challenged Guinea pigs.

This study investigated the effects of roflumilast, a PDE4 inhibitor, on slow-reacting substance of anaphylaxis (SRS-A)-mediated bronchoconstriction and pulmonary leukotriene (LT) release in ovalbumin (OVA)-sensitized and -challenged guinea pigs. Animals were treated with roflumilast orally (0.04, 0.12, 0.4, or 4 mg/kg) or placebo 1 hour before OVA challenge. Bronchoconstriction was quantified by measuring airway conductance (Gaw) and dynamic lung compliance (Cdyn). Roflumilast significantly attenuated the decrease in Gaw (50% inhibitory dose [ID50] = 0.33 mg/kg) and Cdyn (ID50 = 0.25 mg/kg) in a dose-dependent manner and significantly inhibited Cys-LT (ID50 = 0.06 mg/kg) and LTB4 (ID50 = 0.05 mg/kg) release versus placebo-treated animals. Roflumilast did not affect LTD4-induced bronchoconstriction. These findings support the role of roflumilast as an anti-inflammatory treatment for asthma.

Measurement of immunoreactive leukotriene C4 in blood of asthmatic children.

Peptide leukotriene (LT) such as LTC4, LTD4, LTE4 have been considered to be major mediators of immediate type hypersensitivity reaction such as asthma. We have developed a rapid and simple extraction method using a Sep-Pak C18 cartridge for the measurement of LTC4 by radioimmunoassay (i-LTC4). In this extraction method, 91% LTC4 was recovered in a final methanol fraction. The identity was confirmed by the recovery test and by the dilution method. The amount of i-LTC4 in plasma from asthmatic patients was determined by radioimmunoassay after the extraction. The order of the plasma level of i-LTC4 was; severe asthma greater than slight or moderate asthma greater than asthmatic patient without attack greater than healthy adult. The highest level of LTC4 was 0.27 +/- 0.11 pmol/ml in severe asthmatic plasma.