Effect of short-term systemic glucocorticoid treatment on human nasal mediator release after antigen challenge.

The effect of systemic glucocorticoid treatment on early- and late-phase nasal allergic reactions after allergen challenge was determined in a double-blind, cross-over study in 13 allergic individuals. The subjects were pretreated for 2 d before challenge with 60 mg prednisone per day or a matching placebo. A previously described model using repeated nasal lavages for measuring mediator release in vivo was utilized. Symptom scores obtained repeatedly before, during, and after the challenge and the number and timing of sneezes were recorded. The mediators measured were histamine. N-alpha-p-tosyl-L-arginine methyl ester (TAME)-esterase activity, kinins, PGD2, and LTC4/D4. Albumin was also measured as a marker of plasma transudation. Blood samples were taken for determination of total number of white blood cells, differential count, and total blood histamine content. No effect of steroid therapy was found on the appearance of symptoms or any of the mediators, except a reduction in kinins, in the early phase of the allergic reaction. However, in the late phase, the prednisone reduced the number of sneezes (P less than 0.01), as well as the level of histamine (P less than 0.05), TAME-esterase activity (P less than 0.05), kinins (P less than 0.05), and albumin (P less than 0.05). Only low levels of leukotrienes were found in the late phase, but the quantities of these mediators seemed to be decreased by the glucocorticoid treatment (P = 0.06). PGD2 did not increase during the LPR and thus was not affected by glucocorticosteroids. The immediate response to a second challenge 11 h after the first was also evaluated. Whereas the appearance of mediators was enhanced over the initial response to the same challenge dose in placebo-treated subjects, this enhancement was abrogated after prednisone treatment. As this dose of drug is known to be clinically effective in treating hay fever, the present study confirms the earlier findings of others that short-term systemic glucocorticoid treatment inhibits the late phase but not the immediate phase of antigen challenge. Furthermore, secondary enhancement of immediate responses is inhibited. This study shows that glucocorticoids inhibit the generation or release of inflammatory mediators during the late reaction and the physiologic response.

A method for the preparation of imprints from the nasal mucosa.

Important immunological reactions take place on the surface of mucosal membranes. Improved methods for the sampling and quantitative study of the cells taking part in these reactions are therefore desirable. We here describe a new technique for the preparation of imprints from the nasal mucosa. The method utilizes a plastic film coated with a thin layer of an albumin-glycerol mixture to improve cell adherence to the surface. The membrane is gently pressed onto a defined portion of the mucous membrane. Fixation and staining procedures are performed on the plastic film, which is then mounted on a slide and covered by a coverslip. The preparations have excellent optical properties and specific cell types can be easily studied, quantified and related to the specific area of the mucosa from which the imprint was taken.

A brush method to harvest cells from the nasal mucosa for microscopic and biochemical analysis.

A method is described for the sampling of epithelial cells and other effector cells from the human airway mucosa for structural and biochemical analysis. The cell samples are obtained from the nasal mucosa using a small nylon brush which is rotated over the epithelium and soaked and shaken in a small volume of a balanced salt solution. Morphological evaluation using light microscopy and transmission electron microscopy revealed excellently preserved cytological detail. In asymptomatic individuals the cells harvested were as follows: 45 +/- 5.9% (mean +/- SEM) epithelial cells, 38 +/- 7.1% granulocytes, 16 +/- 2.3% large mononuclear cells (monocytes), and 1.3 +/- 2.3% eosinophils. Repeated measurements in the same individual revealed a coefficient of variation of the order of 40% for the proportions of cells harvested. In comparison with nasal airway lavage, a higher proportion of epithelial cells and monocytes were obtained with the brush method. The cells harvested could also be used for biochemical analysis. The histamine content of the cell pellets was found to be strongly correlated with the mast cell count (r = 0.93) and was estimated to about 10 pg/cell, which is higher than previously reported for mast cells obtained from human lung tissue dispersed by an enzymatic method. The present method appears to be appropriate for the study of cellular events in the nasal mucosal epithelium.

Vascular effects of topically applied bradykinin on the human nasal mucosa.

To evaluate the vascular effects of topically applied bradykinin on the human nasal mucosa, 13 asymptomatic hay fever patients and 11 non-allergic subjects were challenged with diluent or bradykinin in three increasing doses. Mucosal blood flow was determined with the 133Xenon wash-out method and expiratory peak flow measurements used to assess nasal airway resistance before and after challenge. Nasal symptoms were recorded. Nasal secretion quantity was measured from preweighed paper handkerchiefs. Bradykinin induced a slight increase in nasal airway resistance which was similar in both allergic and non-allergic subjects. Nasal secretion was clearly increased after challenge with bradykinin compared with challenge with diluent in both allergic and non-allergic subjects. Bradykinin did not, however, induce any change in mucosal blood flow in either group. The present findings could be explained by direct effects of bradykinin on the vascular bed without reflex activity. Bradykinin would then induce an increase in vascular permeability with subsequent oedema formation and increased amounts of fluid on the mucosal surface. In contrast to allergen challenge, bradykinin challenge had no effect on the resistance vessels, changes of which had previously been shown to be largely reflex-mediated.

Long-term treatment with budesonide in vasomotor rhinitis.

Twelve patients entered a long-term study of budesonide treatment in vasomotor rhinitis. The efficacy of the treatment was studied. Possible systemic side effects were studied via an ACTH stimulation test prior to and during treatment. The local effect in the nasal mucosa was studied by means of a nasal biopsy before and after one year of treatment. The treatment has proved effective, while producing no systemic or local side effects during a one-year treatment period.

Topical terlipressin (Glypressin) gel reduces nasal mucosal blood flow but leaves ongoing nose-bleeding unaffected.

Nasal bleeding where the lesion cannot clearly be localized is today usually treated with different forms of nasal packings which are traumatizing to the nasal mucosa and often causes the patient discomfort. In an attempt to develop a more convenient form of treatment the effect of two vasoconstrictor gels on nasal mucosal blood flow was evaluated. Terlipressin gel was shown to reduce nasal blood flow in a dose-dependent way. Therefore, this gel was chosen in a double-blind comparison with placebo in the treatment of 44 patients with posterior epistaxis. Although 50% of the patients did stop bleeding after topical gel administration into the nose there was no statistically significant difference in effect between those who received terlipressin gel and those who received placebo gel. We cannot disregard the fact, however, that the gel in itself has a beneficial effect on nose-bleeding.

Allergen-induced specific and non-specific nasal reactions. Reciprocal relationship and inhibition by topical glucocorticosteroids.

The correlation between allergen-induced specific and non-specific (histamine) nasal reactions was studied, especially as regards topical glucocorticosteroid effects on the non-specific reactions. Thirteen patients with strictly seasonal allergic rhinitis participated. A nasal challenge with histamine and three increasing doses of allergen was performed on the first day. The patients were rechallenged 24 h later with the same histamine dose and the lowest allergen dose from the previous day. The same 2-day challenge was also performed after pretreatment for one week with budesonide in 8 of these patients. Symptom scores were recorded. The N-alpha-tosyl-L-arginine-methyl-esterase (TAME esterase) activity in nasal lavages was determined. The nasal symptoms and the TAME-esterase activity increased at rechallenge for both histamine and allergen, compared with the initial challenge. The mean ( +/- SE) composite nasal symptom score after histamine increased from 3.5 +/- 0.49 to 4.9 +/- 0.35 (p less than 0.01) and after allergen from 1.62 +/- 0.3 to 3.2 +/- 0.5 (p less than 0.01) at rechallenge. Similar increases were recorded for the TAME-esterase activity. A close correlation between the allergen-induced increase in specific (allergen) and non-specific (histamine) reactivity was found (r = 0.7, p less than 0.01, composite nasal symptom score). Treatment with topical glucocorticosteroids abolished the allergen-induced increase in both specific (p less than 0.001) and non-specific (histamine) (p less than 0.01) nasal reactivity.

Reflex activation in allergen-induced nasal mucosal vascular reactions.

Subjects with allergic rhinitis were challenged unilaterally with diluent and increasing doses of allergen. Challenge with the highest dose of allergen was also carried out after topical anesthesia of the nasal cavity using lidocaine. In the contralateral, unprovoked nasal cavity the mucosal blood flow was determined using the 133Xenon wash-out technique and the nasal airway resistance was determined by rhinomanometry before and after challenge. Nasal symptom scores were estimated 15 min after each challenge. Blood flow in the nasal mucosa in the unprovoked right nasal cavity decreased in a dose-dependent manner for th two highest doses of allergen where a reduction of 21% (p less than 0.05) and 26% (p less than 0.01) was obtained. Nasal airway resistance increased somewhat after the highest dose (p greater than 0.05). Topical anesthesia in the provoked nasal cavity inhibited the decrease in blood flow in the unchallenged nasal cavity. These findings suggest that the changes in the tone of the resistance vessels, but not the capacitance vessels, which are induced by allergen, are largely reflex-mediated.

Allergen-induced changes in nasal secretory responsiveness and eosinophil granulocytes.

The release of toxic granule proteins from the eosinophil granulocytes is generally believed to play a crucial part in the development of allergen-induced lesions of the barrier function leading to such clinical features of continuous allergic airway disease as oedema, hypersecretion, changes in responsiveness to specific and non-specific stimuli and, in the case of the lower airways, bronchoconstriction. In the upper airways, a nasal challenge/rechallenge model has proved useful in the study of the allergic inflammatory response in hay fever patients both in experimental settings and during natural pollen exposure. Repeated nasal lavage procedures and challenges with methacholine following an initial challenge with different doses of allergen or placebo were performed in 16 hay fever patients. Following an immediate allergic reaction, a statistically significant increase in the secretory response to methacholine was seen 30 min after challenge with the higher doses of allergen (p less than 0.01) but not after the lowest dose or placebo. An influx of eosinophil granulocytes was seen within 30-60 min of the allergen challenge regardless of the dose (p less than 0.01). The activation of these cells was measured by the increased levels of ECP (eosinophil cationic protein) in the nasal lavage fluid. No relationship was found between individual changes in eosinophils or levels of ECP and changes in the secretory response to methacholine or nasal symptoms. This lends further support to our previous observations that eosinophil granulocytes are not necessarily linked to allergen-induced changes in nasal secretory responsiveness.

125I-albumin may not be used as a tracer of absorption across the human nasal airway barriers.

This study set out to examine the effects of histamine on airway absorption of macromolecules. By employment of a novel "nasal pool" technique instillates containing 125I-albumin, with or without histamine, were kept for 15 min on human nasal mucosa. Unaffected by the presence of histamine, the instillations produced significant levels of plasma radioactivity, increasing for 60 min. However, gel-filtration data showed that only 30% of the plasma radioactivity was still bound to albumin. Incubation experiments indicated that radioiodine did not dissociate from albumin in nasal liquids nor in the blood. Further experiments involved oral ingestion of the entire nasal instillate. Prompt gastrointestinal absorption of radioactivity occurred, giving rise to plasma levels about two orders of magnitude higher than those recorded after the nasal applications. Moreover, only 25% of the plasma radioactivity was now bound to albumin. It must be considered unavoidable that a small portion (less than 1%) of the nasal instillate is swallowed. Hence, the plasma radioactivity detected in this study may largely reflect gastrointestinal break-down of 125I-albumin and subsequent absorption of radioiodine. We conclude that 125I-albumin may not be employed in studies addressing macromolecular absorption across the human nasal mucosa and that previous work and conclusions based on nasal absorption of 125I-albumin are invalid.

Macrophages on the nasal mucosal surface in provoked and naturally occurring allergic rhinitis.

Macrophages are the most common cell type residing in the lumen of the lower airways. However, very little is known about the presence and putative pathogenic implications of macrophages in the upper airways. Using specific immunohistochemical techniques, the presence of and changes in macrophage density were studied before and after allergen exposure in the laboratory and during natural allergen exposure of subjects with seasonal allergic rhinitis. The monoclonal antibody EBM 11 combined with the alkaline phosphatase-anti-alkaline phosphatase-technique was applied on cytospin-prepared slides. In the challenge experiment, 0.5 +/- 0.2% (mean +/- SEM; n = 10) of the total cell number were positive for the EBM 11 marker before challenge, thereby not differing from the controls (0.2 +/- 0.2%; mean +/- SEM; n = 3). Local allergen challenge induced an increase of these cells to a peak of 1.3 +/- 0.4% after 4 h (p less than 0.05). During seasonal exposure there was also a similar increase, from 0.7 +/- 0.2 to 1.3 +/- 0.3% (p less than 0.05; n = 11) in placebo-treated patients and from 0.7 +/- 0.2 to 1.6 +/- 0.4% (p less than 0.05; n = 11) in patients treated with topical glucocorticoids. There was, however, no direct relationship between nasal symptoms and number of macrophages present on the mucosal surface. The study indicates that macrophages are involved in the inflammatory processes of allergic rhinitis.

Effects of glypressin on human nasal mucosa.

The effect of intravenous and topical glypressin, a triglycyl hormonogen of vasopressin, on the nasal mucosa was evaluated in healthy subjects. A dose-dependent reduction of nasal blood flow resulted from both intravenous and topical glypressin. The effect of the latter in gel form lasted 2 hours. Glypressin was also found to decongest the nasal mucosa. Topical application of glypressin gel might be an alternative to conventional treatment with intranasal packing in nose-bleed.

Hay fever treatment with budesonide and beclomethasone dipropionate twice daily - a clinical comparison.

Fifty-two patients with seasonal allergic rhinitis were admitted to a randomized clinical comparison between budesonide (Rhinocort) and beclomethasone dipropionate (Becotide Nasal). All patients were sensitive to birch pollen, which was confirmed by a skin prick test. The drugs were administered intranasally 200 micrograms b.i.d. Symptoms were assessed over four weeks starting with a run-in period of one week. Daily pollen counts were recorded throughout the trial and showed a rather mild birch pollen season. The patients diary cards revealed a beneficial therapeutic effect of the two drugs. No statistically significant differences between the drugs were seen except with regard to sneezing symptoms, where the Rhinocort-treated patients showed less symptoms (p less than 0.05). Side effects were few and transient with both drugs.

Mucociliary transport in the human nose. Effect of topical glucocorticoid treatment.

The effect of topical budesonide on the human mucociliary transport in the nose was investigated utilizing the saccharin-dye test in 10 healthy volunteers. Measurements were made before treatment, after a single dose and after one week continuous treatment with either placebo or active substance. The design of the study was double blind, randomized and cross-over. A single dose of placebo or budesonide did not alter the mucociliary transport as compared to pretreatment values. A trend towards decrease of mucociliary transport was noted after one week treatment with the active drug, a trend that reached statistical significance in the comparison between the 1 h value and the 1 week value.

Budesonide - a new nasal steroid.

39 patients with seasonal rhinitis entered a double-blind study comparing nasal sprays of budesonide 400 micrograms/day and placebo. Symptoms were assessed over a treatment of of three weeks. There were statistically significant differences in favour of the active spray on all measures of assessment. Side effects were mild and the incidence was negligible.

Mast cells on the surface of the mucous membrane--a general feature of inflammatory reactions in the nose?

A redistribution of mast cells towards the epithelial lining of the nasal mucous membrane has been shown to be a part of the allergic inflammatory reaction in hay fever. This results in an increased number of metachromatically stained cells on the surface of the mucous membrane. The involvement of mast cells in other inflammatory reactions in the human nose is not clarified and this may partly be due to methodological difficulties. Utilizing a recently developed imprint technique, specimens were taken from patients with infectious rhinosinusitis in acute and chronic stages. The total number of mast cells on 2 cm2 of the imprint area were counted. Mast cells in extremely low numbers were found in 5 out of 26 patients. Our results indicate that mast cell migration is not present in patients with infectious inflammatory reactions of the nasal mucous membrane.

IgE-positive mast cells on the human nasal mucosal surface in response to allergen exposure.

IgE-bearing mediator cells are suggested to be the effector cells in type I allergic rhinitis. These cells can be demonstrated by their granular constituents or by the surface-bound IgE antibodies. We developed immunohistochemical techniques in order to stain the cell-bound IgE using polyclonal or monoclonal anti-human IgE antibodies. These techniques can be applied to nasal biopsies as shown previously or to cytospin specimen harvested by a brush method. They deliver excellent staining results with well-preserved morphological details. Brush samples were taken from 24 grass pollen allergic subjects before season, after a nasal allergen provocation and two weeks after the onset of season. There was a statistically significant increase in toluidine blue positive and IgE-positive cells 24 hours after nasal provocation (app. 12-fold, p less than 0.05) and more pronounced within the season (app. 58-fold, p less than 0.001) compared to preseasonal values. These cells appeared to be mast cells rather than blood basophils judged by morphological criteria. There was a striking correlation between the number of toluidine blue cells and that of IgE-positive cells (r = 0.98). The number of eosinophils also increased due to the seasonal allergen exposure (p less than 0.001), but less pronouncedly compared to the mast cells. These data re-emphasize the migration of IgE-bearing mast cells and eosinophils into the epithelial lining of the nasal mucosa due to allergen interaction and point to a possible role of mast cells as a carrier for IgE-molecules.

Effect of topical glucocorticoid treatment on nasal mucosal mast cells in allergic rhinitis.

Glucocorticoids were previously considered not to affect the immediate allergic reaction. However, in a nasal allergen challenge, an inhibitory effect on the nasal symptoms induced at the challenge has been shown to occur in patients treated with a recently developed glucocorticoid, budesonide, for 1 week prior to the challenge. This treatment was also found to reduce tissue histamine levels in the nasal mucosa. Mast cells in the mucosa were therefore studied with a view to finding out whether this reduction could be due to a reduction of mast cells. A double-blind study was performed in 14 asymptomatic patients. Nasal biopsies were made before and after 1 week's treatment with either budesonide or placebo. The number of mast cells was counted in two Epon sections after the specimens had undergone specific staining with toluidine blue. No quantitative or qualitative morphological changes in the mast cells were found as a result of treatment.

The effect of budesonide on the immediate reaction to allergen challenge - a rhinomanometric study.

In a nasal allergen challenge test, the effect of the glucocorticosteroid budesonide on the pollen allergic type I reaction was investigated. Placebo and two different doses of budesonide were given intranasally for one week before the challenge. The study was designed as a double-blind cross-over trial. Treatment for one week with the active steroid significantly reduced the nasal secretion compared with placebo, as measured with the aid of a symptom score, and significantly reduced the induced nasal blockage measured objectively by rhinomanometry. No difference was found between the two doses of budesonide, 400 micrograms and 100 micrograms daily, used in the study.

Budesonide and nasal histamine challenge.

The possible mode of action of the lately demonstrated steroid effect on the immediate type allergic reaction was investigated. The influence of a topical steroid, budesonide, on the effects that released mediators have on the nasal mucosa was also studied. A nasal histamine challenge study was performed in a double-blind, cross-over fashion, a 1-week pretreatment with budesonide or placebo preceding the challenge. Symptoms were recorded by means of symptom score as well as objectively via rhinomanometry. In contrast to a previous allergen challenge study, the steroid was found to have minimal effect on the histamine-induced nasal symptoms. It is therefore concluded that other modes of steroid action must also be involved in the steroid effect on the immediate type allergic reaction.