Histologic studies of human skin test responses to ragweed and compound 4880: II. Effects of corticosteroid therapy

In a controlled study, a 1-week course of a moderate dosage of corticosteroids given to ragweed-sensitive subjects was associated with a statistically significant decrease in blood eosinophil levels and tissue eosinophil, but not mast cell, responses to ragweed antigen and compound $\text{48}\text{80}$, This therapy did not affect IgE-class anti-ragweed antibody levels or gross whealing responses to antigen or compound $\text{48}\text{80}$. While these findings suggest indirectly that a major component of the corticosteroid effect is likely an alteration of the mobilisation of eosinophils to the inflammatory site, additional studies will be required to more directly characterize the mechanisms involved.     

Histamine suppression of in vivo eosinophil accumulation and histamine release in human allergic reactions

Although it has been shown that histamine inhibits antigen-induced in vitro histamine release from basophils, it is unclear whether histamine inhibits in vivo mediator release in human allergic reactions. We report effects of exogenous histamine on histamine release and inflammatory cell responses in antigen-challenged skin sites in eight ragweed-sensitive individuals. Four heat-suction blisters in each subject were unroofed, and a collection chamber was appended to each blister base. Chamber A contained 1000 PNU/ml ragweed extract; chamber B contained buffered saline (control fluid); chamber C contained 1000 PNU/ml ragweed and 50 ng (5 x 10(-7) M) of histamine; and chamber D contained histamine alone (50 ng). Comparative analyses of chamber histamine levels in individual subjects showed that (1) histamine levels in chamber A were significantly greater than those in chamber B (p less than 0.01) and that histamine levels in chamber C were not significantly different than those in chamber D (p less than 0.5). Likewise, comparison of eosinophils attaching to membrane filters appended to the chamber bases for 2 hr showed that there were significantly more eosinophils in chamber A than in chamber B (p less than 0.01) and that there was no significant difference in eosinophil numbers on filters appended to chamber C vs chamber D. In three of four subjects studied, addition of exogenous histamine (50 ng/ml) to ragweed before intradermal injection inhibited the ultrastructural mast cell alterations seen within 10 min after injection of ragweed alone. In the one subject in which mast cell alterations were not prevented, exogenous histamine also did not inhibit antigen-induced histamine release or subsequent eosinophil accumulation in the skin chambers.     

Inhibitory effect of cetirizine 2HCl on eosinophil migration in vivo

The effect of a potent antihistamine, cetirizine, was studied on allergic patients and normal subjects by means of an in-vivo 'skin window' technique. All subjects showed significant inhibition of skin-test responses to grass pollen, compound 48/80, histamine and methacholine, after administration of a single dose (10 mg) of cetirizine. Compared to placebo, cetirizine significantly decreased the eosinophils attraction at skin sites challenged with grass pollen and compound 48/80. In allergic patients no change in eosinophil migration pattern was noted with histamine and methacholine skin-tested sites. In normal subjects, compound 48/80 and histamine did not induce eosinophil accumulation and cetirizine did not modify cellular patterns as compared to placebo. These results suggest that cetirizine acts on eosinophil migration by inhibiting the release of mast cell mediators or inhibiting the eosinophilotactic mediators themselves.     

Dopamine inhibition of histamine-mediated cutaneous responses

Several patients receiving dopamine for hypotension were skin tested for possible penicillin sensitivity. Not only were the penicillin skin tests negative but also the histamine control. On the possibility that dopamine might affect cutaneous histamine responses, we examined the effect of dopamine on histamine, antigen, morphine, and compound 48/80 skin responses. Both intradermal and intravenous dopamine selectively inhibited histamine but not antigen, morphine, or compound 48/80 skin responses, and the inhibition was in a dose-related fashion. This observation indicates that histamine should not be used to demonstrate dermal reactivity in patients receiving dopamine. The results of this study also suggest that histamine may not be the sole mast cell-derived mediator involved in the wheal-and-flare reaction characteristic of immediate-type skin tests since dopamine did not affect skin reactions caused by endogenous mast cell degranulation. Finally, the possible use of dopaminergic drugs in diseases with histamine-associated symptoms is discussed.     

Patterns of mast cell alterations and in vivo mediator release in human allergic skin reactions

Patterns of in vivo histamine release in skin sites challenged with ragweed antigen were compared in five human subjects sensitive to this antigen and four nonallergic individuals, using a newly developed skin-chamber technique. These findings were compared with inflammatory cell responses in the reaction sites and patterns of ultramicroscopic mast cell alterations in biopsy specimens of skin tests in the same subjects. Definite mast cell alterations occurred within 15 sec and appeared maximal within 5 to 10 min after antigen injection. Histamine levels in appended chambers increased after a lag of 10 to 30 min and were elevated for at least 60 min after antigen challenge. Eosinophils accumulated only in antigen-induced reaction sites. However, there was no precise quantitative correlation among the degree of change in these three measurements. These appear to be promising approaches to further in vivo studies of human allergic reactions.     

Factors in the tissue eosinophil responses in human immediate hypersensitivity reactions.

Factors underlying the eosinophil responses in the tissue sites of immediate hypersensitivity reactions have been investigated. A quantitative assessment of the number of eosinophils appearing in skin test reactions to ragweed antigen and compound 48/80 in ragweed-sensitive humans has been compared with several other parameters. There was a moderate, statistically significant, correlation with serum levels of IgE-class antiragweed antibody; also, eosinophil responses were minimal or absent in minimally positive threshold dilution skin tests. Tissue eosinophil responses were generally limited to those with baseline blood eosinophil levels of at least 150 mm3; however, there was no correlation between blood and tissue eosinophil levels in individual subjects. These findings suggest that the pathogenesis of the observed tissue eosinophil responses may be multifactorial.     

Effect of Terbutaline, a β2-adrenergic Receptor Stimulating Compound, on Cutaneous Responses to Histamine, Allergen, Compound 48/80, and Trypsin

The beta-adrenoceptor stimulating agent terbutaline (2 ng-2 microgram) injected intradermally in eight atopic subjects produced a dose-dependent inhibition of the skin reactions induced by subsequently injected allergen. After injection of 0.5 microgram terbutaline inhibition of the flare and weal responses was demonstrable throughout the observation period of 90 min. The flare response induced by histamine, the histamine liberator compound 48/80 and the proteolytic enzyme trypsin was not inhibited by terbutaline in the doses used, suggesting a selective action of terbutaline on the allergen-induced response. The weal response elicited by histamine and compound 48/80 was slightly reduced by 2 microgram terbutaline. It is suggested that pretreatment of the skin with terbutaline interferes with the ability of the cutaneous mast cells to respond to challenge with allergen and that terbutaline produces this effect in doses lower than those needed to counteract the permeability increasing effect of released mediator substances.     

Increased compound 48/80 induced local histamine release from nonlesional skin of patients with chronic urticaria

Histamine released from skin mast cells in normal skin sites of patients with idiopathic chronic urticaria (CU) and normal volunteers was assessed with the skin chamber technique. Small amounts of histamine were spontaneously and continuously released during the 4-hour observation in both groups but were twofold greater in patients with CU. In addition, histamine levels were significantly more elevated in sites challenged with compound 48/80 than in unstimulated sites. Patients with CU differed from normal volunteers in that histamine release induced by 48/80 compound was significantly greater at 1 and 2 hours after challenge. The number of mast cells and the histamine content of the skin did not differ in the two groups. These observations could suggest a functional defect at the mast cell level rather than a difference in their numbers.     

Effects of infused histamine on asthmatic and normal subjects: comparison of skin test responses

The effect of histamine infused intravenously at sequentially increasing concentrations (0.05, 0.1, 0.25, 0.5, and 1 μ/kg/min) on the wheat responses to intradermal histamine and compound 48/80 in eight normal and five asthmatic subjects and to allergen skin tests in five asthmatic subjects was measured. These measurements were repeated following pretreatment with the H-1 antagonist hydroxyzine or the H-2 antagonist cimetidine, either alone or in combination. Histamine infused in progressively increasing concentrations had no effect on histamine, compound 48/80, or allergen skin tests either before or after H-1 or H-2 antihistamine treatment. No significant differene was found in the concentration of histamine or compound 48/80 required to elicit a 10-mm wheat in normal or asthmatic patients. Pretreatment with the H-2 antagonist alone had no effect on histamine or compound 48/80 skin tests in either group. However, the H-1 antagonist significantly reduced the wheat response to histamine (p < 0.05 normal; p < 0.025 asthmatics) and compound 48/80 (p < 0.05 normal; p < 0.025 asthmatics) in both groups. The combination of H-1 and H-2 histamine antagonists was not significantly different from the H-1 antagonist alone. Antigen skin testing was suppressed 82% by the hydroxyzine alone; no significant suppression was induced by cimetidine alone, and the combination of hydroxyzine plus cimetidine was only slightly more effective than hydroxyzine alone. The results indicate that blockade of histamine H-2 receptors with cimetidine has little or no additive effect on H-1 antagonist-suppressed skin test responses to histamine, compound , or antigen. Furthermore, the capacity of histamine to suppress histamine release in vitro from basophils was not demonstrated in vivo assessing skin mast cell responses. This observation combined with earlier studies on the human lung mast cell, which also failed to demonstrate that histamine had an inhibitory action, suggests that the human mast cell may not respond to histamine like the basophil and that this discrepancy may represent a fundamental difference in the cell types.     

Inhibition of the Late Phase Reaction to Anti-IgE by Previous Mast Cell Activation with Compound 48/80

The concept of an obligate association between mast cell activation and development of a late phase reaction (LPR) to various agents in human skin was further elucidated. Skin sites were treated four times at 24 h intervals with non-LPR-inducing doses of the histamine liberator compound 48/80 in 10 volunteers. The previously compound 48/80-challenged sites responded with an approximately 40% attenuated early response ( P < 0.01) and a 70% reduction of the LPR ( P < 0.001) to subsequently injected anti-IgE as compared with simultaneous reactions at control sites The data suggest that mediators from the cutaneous mast cells are necessary for the final expression of an LPR.     

Leukocyte histamine release in Hymenoptera-allergic patients. Correlation with skin test reactivity and changes following hyposensitization therapy

We have studied 46 Hymenoptera-allergic patients and 11 nonallergic controls by grading the severity of their sting reaction, determining their skin test reactivity, and performing human leukocyte histamine release with a commercial mixed stinging insect extract. A significant correlation was found between increasing severity of sting reaction and increasing skin test reactivity (p < 0.001). In $\text{41}\text{46}$ allergic patients from 15 to 100 per cent release of total cellular histamine occurred, whereas in the 11 nonallergic controls no greater than 10 per cent release of total cellular histamine was observed. Skin test reactivity correlated significantly with cell sensitivity in the allergic patients (p < 0.001). Following hyposensitization therapy, cell sensitivity generally did not change, but significant increases in antigen-neutralizing capacity (A.N.C.) of allergic serum did occur in $\text{11}\text{15}$ Hymenoptera-allergic patients.     

Ligation of IgE receptors causes an anaphylactic response and neutrophil infiltration but does not induce eosinophilic inflammation in mice

Background: Eosinophils are selectively recruited into the tissues during chronic allergic inflammation. IgE is considered an initiator of the allergic reaction; however, the roles of IgE in allergic inflammation are not fully understood. Objective: We tested the hypothesis that antigen interaction with specific IgE antibody provokes eosinophilic inflammation. Methods: BALB/c mice were actively sensitized with ragweed extract and passively sensitized with anti-dinitrophenyl (anti-DNP) mouse IgE and challenged intraperitoneally by injecting either ragweed extract or DNP-ovalbumin (OVA). Immediate anaphylactic responses were examined by monitoring vascular permeability and by measuring histamine content in peritoneal lavage fluids. Late-phase allergic responses were examined by total cell counts and cell differentials. Results: Mice sensitized and challenged with ragweed showed immediate anaphylactic responses followed by temporal increases in neutrophils at 3 to 12 hours and sustained ncreases in eosinophils in their peritoneal cavities after 24 hours. Double-sensitized mice (ie, sensitized actively for ragweed and passively for DNP-OVA) challenged with ragweed showed immediate anaphylactic responses and peritoneal eosinophilia at 48 hours. Double-sensitized mice challenged with DNP-OVA showed comparable immediate anaphylactic responses but no peritoneal eosinophilia. Furthermore, at 8 hours, ragweed-challenged animals recruited both eosinophils and neutrophils, but DNP-OVA–challenged animals recruited only neutrophils. Finally, after active sensitization and challenge with ragweed, mast cell–deficient mice (WBB6F1-W/W^v) lacked the immediate response but showed comparable eosinophil accumulation as their litter mate controls (WBB6F1-+/+). Conclusion: Interaction of antigen with IgE antibody is insufficient to provoke eosinophilic inflammation in mice. (J Allergy Clin Immunol 2000;105:1202-10.)     

Cutaneous responses to histamine, compound 48/80, and codeine in patients with chronic renal failure

Pruritus is a common symptom associated with chronic renal failure (CRF). But increased plasma histamine levels and skin mast cell proliferation previously reported in these patients did not correlate with the intensity of the pruritus. Since increased mast cell releasability was described in chronic idiopathic urticaria, we attempted to examine whether this mechanism could explain pruritus in patients with CRF. Twenty-five patients with end stage renal failure were skin tested with histamine, codeine, and compound 48/80. There were nine patients on continuous ambulatory peritoneal dialysis, eight patients on hemodialysis, (tested both before and after dialysis) and eight patients with advanced CRF. Wheal area after intradermal injection of three concentrations of the above substances was measured. In general, the wheal areas in all patients with CRF were either similar to or smaller than those of the control group who were without renal impairment. In conclusion, patients with CRF with or without dialysis therapy demonstrated unchanged or decreased skin test responses to histamine, codeine, and compound 48/80. Increased mast cell releasability cannot explain the pruritus in patients with CRF.     

Histologic responses in human skin test reactions to ragweed. IV. Effects of a single intravenous injection of steroids.

A controlled study has been carried out dealing with the early effects of a single intravenous dose of either methylprednisolone or placebo or newly developed and ongoing cellular inflammatory responses in immediate hypersensitivity skin test reactions. There was no significant difference in the tissue eosinophil responses to ragweed injected 2 hr before and 2 hr after placebo; there was a significant rise (101% +/- 39) from the second to fourth hour after antigen injection. By contrast, there was a marked decrease in the tissue eosinophil response to antigen injected 2 hr after steroids as compared to the pattern seen in the presteroid reaction. In addition, the eosinophil numbers not only did not increase from the second to fourth hour when steroids were injected at the second hour but decreased markedly. These findings suggest early suppressive effects on tissue eosinophil responses within 2 hr after steroid were administered intravenously. Also, there may be trafficking of eosinophils both into and out of these inflammatory sites during the first hours after intradermal antigen injection.     

Chronic idiopathic urticaria: Profiles of skin mast cell histamine release during active disease and remission

Chronic idiopathic urticaria (CIU) is characterized by the increased releasability of histamine by mast cells in normal-appearing skin. In active CIU, this abnormality is consistently present. To determine if this finding subsides when the disease goes into remission phase, we analyzed the histamine secretion in patients with CIU in remission (CIUR) compared with that of patients with CIU and in normal control (NC) subjects, with the skin-chamber technique. The profiles of histamine release in sites challenged with compound 48/80 were significantly different in the groups with CIU and CIUR. Furthermore, patients with CIUR did not differ from NC subjects in terms of histamine releasability under compound 48/80 stimulation (p greater than 0.1). These data suggest that the state of excessive skin mast cell sensitivity is a reversible and transient phenomenon in CIU disease.     

The biologic activity of mast cell granules in rat skin: effects of adrenocorticosteroids on late-phase inflammatory responses induced by mast cell granules

Mast cell degranulation in rat skin in response to Compound $\text{48}\text{80}$, anti-IgE or to the intradermal injection of purified rat peritoneal mast cell granules (MCG) produces a late-phase response (LPR) characterized by polymorphonuclear infiltrates at 2 to 8 hr followed by mononuclear cell infiltrates at 24 to 48 hr. MCG generate LPR in a dose-related fashion; 1 μg is sufficient to produce discernible LPR, whereas 5 μg attract ≥ 150 cells/high-powered field (3+ to 4+ response) in rat skin after 8 and 24 hr. Infiltrates producing 3+ responses are also induced by 5 μg of two partially purified factors obtained from solubilized MCG by ultrafiltration: a high molecular weight (HMW) fraction containing molecules >10,000 daltons and a low molecular weight (LMW) fraction with a molecular weight of 500 to 10,000. The effect of depletion of endogenous corticosteroids on LPR was studied in adrenalectomized, sham-operated, and nonoperated littermates. LPRs were induced by 5 μg each of MCG, HMW and LMW fractions, and anti-IgE. Adrenalectomized, sham-operated, and normal nonoperated littermates expressed equivalent LPRs. Thus adrenalectomy has no detectable effect on LPR after mast cell degranulation. The dose-response effects of exogenous corticosteroids on LPR were assessed by pretreating rats with hydrocortisone, methylprednisolone, or dexamethasone. Normal rats injected with methylprednisolone (0.5 to 500 μg), hydrocortisone (0.4 to 400 μg), or dexamethasone (7.6 μg) per 24 hr for 3 days had significantly suppressed LPRs after 8 or 24 hr in response to anti-IgE, MCG, HMW, or LMW fractions. Within 1 day of hydrocortisone (400 μg) pretreatment, the LPR induced by anti-IgE and MCG were considerably reduced. Thus there are at least two separate constituents of MCG capable of eliciting late-phase inflammatory responses in rat skin, and corticosteroids may prevent the LPR thereby elicited. The capacity of corticosteroids to suppress LPR may contribute to their clinical usefulness in treating allergic diseases.     

Passive cutaneous anaphylaxis in the rat, induced with two homologous reagin-like antibodies, and its specific inhibition with disodium cromoglycate

Rats immunized with egg albumin and Bordetella pertussis organisms produce a `mast cell sensitizing' antibody (MCSAb) which is thermolabile, a potent skin sensitizer and reagin in character. Similarly the immune response to Nippostrongylus brasiliensis in rats is closely associated with the formation of antibodies which also resembles human reagins. Homologous passive cutaneous anaphylactic (PCA) reactions induced by N. brasiliensis serum were found to be similar to those produced using the adjuvant induced antibody in that both were completely inhibited by, combined treatment with mepyramine and 2-bromo-D-lysergic acid diethylamide (BOL148), cyproheptadine or pretreatment with compound 48/80. In contrast, skin reactions involving passive sensitization of rats with rabbit hyperimmune antiserum were much less affected. Studies on mast cell disruption at the site of PCA reactions showed that such reactions using N. brasiliensis serum were accompanied by degranulation of mast cells, and confirmed that mast cell damage occurs in PCA induced with MCSAb. Both the PCA and the mast cell disruption were maximal 5 minutes after antigen challenge in both rat reagin systems. The skin reaction produced using rabbit hyperimmune antiserum was not primarily dependent on, or associated with, mast cell disruption, since it was still possible to induce skin reactions when the mast cells had been disrupted by compound 48/80, and skin reactions could be obtained without significant mast cell disruption.

Disodium cromoglycate, a new compound introduced for the treatment of asthma, was shown to inhibit both the PCA and mast cell disruption induced using both rat reagin antibodies but not the skin reactions produced with rabbit anti-serum. It was possible to obtain substantial inhibition of mast cell disruption induced by rat reagin, even when the PCA was inhibited only slightly. At higher doses the discharge of the mediators from mast cells was also prevented. This interference with mast cell permeability was not unspecific since disodium cromoglycate did not prevent the skin reaction or mast cell disruption produced by compound 48/80. As expected mepyramine was able to partially inhibit the skin reaction without affecting mast cell disruption induced by rat reagin or compound 48/80. It is suggested that disodium cromoglycate acts at some critical pathway in the events after the union of antigen with reagin antibody and that this critical pathway might be common to both the human and rat reagin systems.

     

The Effect of Aspirin during Aspirin "Desensitisation" on Compound 48/80 and Histamine-Induced Skin Responses in Aspirin-Sensitive Asthmatics

In a group of aspirin-sensitive asthmatics we studied skin weal and flare responses to intradermal injections of compound 48/80 and histamine during oral aspirin (ASA) provocation and after ASA "desensitisation". During provocation (bronchospasm accompanied by naso-ocular symptoms) the mean weal area after compound 48/80 increased to about 42.4% (P less than 0.05). Neither the threshold (provocative) doses of ASA nor 600 mg ASA, when given after ASA-desensitisation, significantly influenced the weal reactions to compound 48/80 (mean changes of area were -1.8% and -16.5% respectively). Aspirin did not change flare reactions to compound 48/80 and weal and flare reactions to histamine on any of the three study occasions. Initial (pre-aspirin) weal reactions to compound 48/80 after desensitisation to the threshold ASA doses were significantly reduced, but after desensitisation to 600 mg ASA were significantly increased as compared with the reactions before. These data suggest that ASA-"desensitisation" may influence the skin reactivity to non-specific mast cell degranulating stimulus in ASA-sensitive asthmatics.     

Human skin target cells: nature and fate in the immediate hypersensitivity reaction

Localisation of antibody and/or antigen to target cells in human skin was studied by autoradiography. Full-thickness biopsies were obtained 24, 48, and 72 hours following passive local sensitization with ¹²⁵I-labeled E-myeloma protein. Other biopsies were obtained 5, 15, and 30 minutes following challenge with goat anti-E-myeloma. Sites passively sensitized with ragweed-allergic sera were challenged with ¹²⁵I-labeled ragweed antigen E, and biopsies were obtained at 5, 15, and 30 minute intervals. Forty-eight and 72 hours following passive sensitization the E-myeloma protein was found to be exclusively associated with skin mast cells. In some biopsies, obtained 24 hours following passive sensitization in addition to mast cells, E-myeloma was localized to other cell types as well. Biopsies of the wheal-and-flare reaction sites revealed that the antigen-induced release of histamine from the human skin mast cells results in the degranulation of the reacting cells.     

Diagnostic tests in ragweed-allergic asthma. A comparison of direct skin tests, leukocyte histamine release, and quantitative bronchial challenge

Thirty-six patients with a history of seasonal ragweed asthma were tested. Direct intradermal skin tests, leukocyte histamine release, and quantitative inhalation bronchial challenge correlated significantly. Data reported suggest that quantitative skin tests are as diagnostically valuable as quantitative inhalation bronchial challenge or leukocyte histamine release. Data also suggest that lung and skin mast cells and circulating basophils respond as a single population of cells to ragweed antigens.