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R Djukanovic

Publications and source records attributed to R Djukanovic.

At least 55 records · Page 3Linked to original sources
PubMed1997

Identification of neutrophil chemotactic factors in bronchoalveolar lavage fluid of asthmatic patients.

BACKGROUND: Although neutrophils have been implicated in bronchial asthma, the mechanism(s) which bring these cells into the airways is poorly understood. OBJECTIVE: To investigate the presence and identity of neutrophil chemotactic factors in bronchoalveolar lavage (BAL) fluid from atopic asthmatic subjects. METHOD: BAL fluid was obtained from 13 subjects (seven asthmatics and six normals), aged 19 to 60 yr, at bronchoscopy. Separation of neutrophil chemotactic activity (NCA) was achieved by FPLC cation exchange chromatography. Fractions were collected and assayed for chemotaxis in multiwell micro-chemotaxes chambers using polycarbonate filters, for the complement peptide C5a/C5a des Arg by radioimmunoassay (RIA) and for interleukin-8 (IL-8) by ELISA. RESULTS: NCA was found in FPLC fractions of BAL samples in four out of seven asthmatics and each of these subjects had at least three similar peaks of NCA. The major peak of NCA was found to contain immunoreactive C5a/C5a des Arg and chemotaxis. In response to this NCA could be blocked by desensitization of the neutrophils with recombinant C5a. Purified serum derived C5a/C5a des Arg was found to have altered chromatographic properties when added to BAL fluid; this suggested that BAL fluid contained proteins which interacted with the C5a/C5a des Arg. Immunoreactive IL-8 (iIL-8) was also detected but its concentration or chemical form was insufficient to induce neutrophil chemotaxis. CONCLUSION: This study demonstrates that bronchial asthmatic lavage fluid contains C5a/ C5a des/Arg and IL-8, together with other as yet unidentified factors which may contribute to neutrophil recruitment in this disease.

Adolescent↗
PubMed1996

Alveolar tissue inflammation in asthma.

As physiologic and autopsy evidence suggests that peripheral airways and parenchyma are involved in asthma, we hypothesized that significant alveolar tissue inflammation is present in patients with stable, chronic asthma. Eleven patients with nocturnal asthma (NA) and 10 patients with non-nocturnal asthma (NNA) were studied. Each subject underwent two bronchoscopies with proximal airway endobronchial and distal alveolar tissue transbronchial biopsy in a random order at 4:00 P.M. and 4:00 A.M. Morphometric analysis was used to determine the number per volume (Nv) of inflammatory cells. Between-group comparisons showed that the Nv of eosinophils was greater in the NA alveolar tissue 4:00 A.M. compared with the subjects with NNA (40.2 x 10(3) [26.4-57.1 x 10(3), IQ] versus 15.7 x 10(3) [2.1-35.2 x 10(3), IQ], p = 0.05). In regard to the airway biopsies, no difference in the inflammatory and epithelial cells between the two groups was seen at either time. The NA group exhibited greater eosinophils and macrophages in the alveolar tissue at 4:00 A.M. compared with 4:00 P.M. (40.2 x 10(3) [26.4-57.1 x 10(3), IQ] versus 10.3 x 10(3) [2.7-16.8 x 10(3), IQ], p = 0.016 for eosinophils and 215.1 x 10(3) [129.9-356.1 x 10(3), IQ] versus 166.3 x 10(3) [150.7-212.6 x 10(3), IQ], p = 0.031 for macrophages). Only alveolar tissue eosinophils, not proximal airway tissue eosinophils, correlated with the nocturnal decrement in lung function (r = -0.54, p = 0.03). These findings suggest that eosinophils and macrophages accumulate to a greater extent in the alveolar tissue and these changes contribute more to the variation in lung function compared with inflammation in the more proximal tissue.

Adult↗
PubMed1996

T-cell cytokine profile evaluated at the single cell level in BAL and blood in allergic asthma.

Atopic asthma is characterized by bronchial mucosal inflammation, involving eosinophils, mast cells, and lymphocytes. It has been suggested that the development and maintenance of this allergic inflammation is due to T-lymphocyte activation with predominant production of the cytokines interleukin 4 (IL-4) and IL-5. To address the ability of peripheral blood and bronchoalveolar lavage T-cells to generate IL-2, IL-4, or interferon gamma (IFN-gamma), we have employed a flow cytometric method which permits analysis of cytokine production at the single cell level within 5 h of obtaining cell samples. When stimulated with PMA and ionomycin, there was a greatly increased percentage of IFN-gamma-producing cells among bronchoalveolar lavage (BAL) T-cells from the subjects with asthma (median 74%), compared with atopic and nonatopic controls (35 and 43%, respectively; P>0.01). The proportion of BAL T-cells producing IL-4 was small (median 1.7%, range 0 to 7.8% in the asthmatic group). In all three groups, the proportion of BAL T-cells producing IL-2 or IFN-gamma was increased compared with T-cells from peripheral blood. There was no significant difference between the three groups in the percentage of BAL T-cells producing IL-2, or in the percentage of peripheral blood T-cells producing IFN-gamma, IL-2 or IL-4. These findings indicate that IL-4 production is confined to a relatively small proportion of airway and blood T-cells and that there is selective enhancement of IFN-gamma production by airway T-cells in asthma.

Adult↗
PubMed1996

Effects of theophylline on inflammatory cells and cytokines in asthmatic subjects: a placebo-controlled parallel group study.

The anti-inflammatory effects of oral theophylline on cells in bronchial biopsies of symptomatic atopic asthmatic subjects were investigated. Following a 2 week run-in period, asthmatic subjects were randomly assigned to either placebo (n=11) or theophylline (n=15). Bronchial biopsies were taken at fibre-optic bronchoscopy at the beginning and end of a 6 week period, during which subjects took placebo or theophylline medication at a dose intended to produce therapeutic concentrations. Nine of the placebo subjects and 12 of the theophylline subjects completed the study. Improvement in asthma control was seen in the theophylline-treated group. The mean (SD) theophylline blood level at the end of the study was 10.9 (6.0) microg x mL-1. A significant decrease in interleukin (IL)4 expression from 1.38 to 1.04 cells x mm-2 (<0.05) and a trend to a reduction in IL-5 from 1.29 to 0.48 cells x mm-2 (NS) were seen in biopsies from the theophylline-treated group compared with placebo, although there was no change in mast cell numbers (judged by tryptase expression). A decrease in epithelial CD8+ cells from 2.60 to 0.53 cells x mm-1 of surface (<0.05) was noted. This study shows an anti-inflammatory effect of theophylline in asthmatic bronchi, both in cell numbers and in the expression of IL-4, believed to be an important cytokine in the pathophysiology of asthmatic inflammation. We speculate that theophylline induces downregulation in vivo of cytokine production, accounting for the known inhibitory effect of theophylline on the late asthmatic reaction.

Administration, Inhalation↗
PubMed1995

15-lipoxygenase immunoreactivity in normal and in asthmatic airways.

Products of the 15-lipoxygenase (15-LO) pathway of arachidonic acid metabolism such as the mono- and di-hydroxyeicosatetraenoic acids (HETEs) may contribute to the pathophysiology of allergic airway inflammation through the recruitment and activation of inflammatory cells and stimulation of glandular secretion. In this study we have examined the expression of 15-LO and its cellular localization in the asthmatic and normal bronchial mucosa. Bronchial mucosal biopsies were obtained by fiberoptic bronchoscopy from 10 patients with symptomatic allergic asthma and six normal control subjects and processed into glycolmethacrylate resin. Sections 2 microns thick were immunostained using a specific rabbit polyclonal antihuman 15-LO antibody. Strong immunoreactivity for 15-LO was present throughout the epithelium in both the asthmatic and the normal subjects, with no difference between the two groups. Cells expressing 15-LO immunoreactivity were also present in the submucosa of both groups, with a significantly greater number present in the asthmatic group (median, 15.3 cells/mm2) than in the normal group (median, 6.9 cells/mm2) (p = 0.01). The majority (85%) of the submucosal 15-LO+ cells were eosinophils. Patchy 15-LO immunoreactivity was also seen in the vascular endothelium in both groups. These findings demonstrated increased 15-LO expression in the bronchial submucosa of asthmatic subjects, and they suggest that 15-LO products in asthma originate from both bronchial epithelium and infiltrating eosinophils.

Adult↗
PubMed1995

Neuropeptide-containing nerves in endobronchial biopsies from asthmatic and nonasthmatic subjects.

The presence and distribution of neuropeptide-containing nerves within endobronchial biopsies has been investigated in symptomatic asthmatics (n = 17) and in asymptomatic nonasthmatic control subjects (n = 17). Biopsies from large airways, obtained under local anesthesia by flexible fiberoptic bronchoscopy, were processed immediately and analyzed for nerves using specific indirect immunofluorescence with antisera to the neural marker protein gene product 9.5 (PGP 9.5) and the neuropeptides vasoactive intestinal peptide (VIP), substance P (SP), calcitonin gene-related peptide (CGRP), and neuropeptide tyrosine (NPY). PGP 9.5-positive nerves were present in all the biopsies from both subject groups, being identified in relationship to epithelium, glands, smooth muscle, and blood vessels. VIP- and NPY-immunoreactive nerves were equally present in the biopsies of both asthmatic and nonasthmatic subjects, being localized to smooth muscle and glands. Using well-substantiated antibodies, no nerves immunofluorescent for SP or CGRP were identified in any of the biopsies of the subjects. In the asthmatic patients, there were no significant correlations between the PGP 9.5, VIP, or NPY immunofluorescence scores and the resting spirometric values (FEV1) or the level of nonspecific bronchial responsiveness, as assessed by the provocative concentration of methacholine required to produce a 20% fall in FEV1. To verify the single biopsy findings, two further studies were undertaken, one in which biopsies were stained from two airway sites (proximal and distal) and a second in which the findings in carinal specimens obtained using biopsy forceps from freshly resected lung tissue were compared with those in a surrounding area of tissue.(ABSTRACT TRUNCATED AT 250 WORDS)

Adolescent↗
PubMed1994

Cytokine binding to CD4+ inflammatory cells: implications for asthma.

While LCF is present in BAL early after antigen challenge, we know little about its other potential effects beyond CD4+ T cell, monocyte, and eosinophil chemotaxis and monocyte and CD4+ T cell activation. The work described here focuses on the hypothesis that the secreted protein products of T cells participate in the airway inflammatory process that underlies human asthma, and in particular that LCF could play an early role because of the unusual responsiveness of LCF-producing T to histamine. To date, most studies have addressed the measurement of cytokines derived from CD4+ T cells (e.g., IL-2, IL-3, IL-4, IL-5, and GM-CSF) in the airways of asthmatics, and attempted to correlate the presence of protein or mRNA with the complexion of the inflammatory infiltrate. These studies have been based upon the reports that there are increased numbers of CD4+ T cells in the airways of asthmatics, and that the presence of eosinophils might correlate with the secretion of TH2-type cytokines like IL-3, -4, and -5. Using this information as a background, our work has approached the problem in an entirely different way. We have focused our attention on the early events in antigen-induced asthma that are responsible for CD4+ cell accumulation in the lung, including CD4+ T cells, eosinophils, and monocytes. We have attempted to identify mechanisms by which mast cell mediators, in particular histamine, might play a role in the secretion of chemotactic lymphokines that are selective for CD4+ cells by using CD4 itself as a chemotactic factor receptor.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1994

Interleukin-4, -5, and -6 and tumor necrosis factor-alpha in normal and asthmatic airways: evidence for the human mast cell as a source of these cytokines.

Asthma is characterized by the presence of an inflammatory cell infiltrate in the bronchial mucosa consisting of activated mast cells, eosinophils, and T cells. Several cytokines are considered to play a pivotal role in this response, particularly interleukin (IL)-4, IL-5, IL-6, and tumor necrosis factor-alpha (TNF-alpha). In this study, we have used immunohistochemistry applied to thin glycol methacrylate sections of bronchial mucosal biopsies to define the cellular provenance of these cytokines in normal and asthmatic airways. Both the asthmatic and normal mucosa contained numerous cells staining positively for all four cytokines, with the majority identified as mast cells by their tryptase content. Eosinophils also accounted for some IL-5 immunostaining in the asthmatic biopsies. By using two monoclonal antibodies directed to different epitopes of IL-4, we provide tentative evidence for enhanced IL-4 secretion in asthma. Similarly, a sevenfold increase in the number of mast cells staining for TNF-alpha in the asthmatic biopsies suggests that this cytokine is also up-regulated in this disease. These findings clearly identify human mast cells as a source of IL-4, IL-5, IL-6, and TNF-alpha and add to the view that, along with T cells, mast cells may play an important role in initiating and maintaining the inflammatory response in asthma.

Adult↗
PubMed1993

Skin responses to bradykinin, kallidin, and [desArg9]-bradykinin in nonatopic and atopic volunteers.

BACKGROUND: Kinins are potent vasoactive oligopeptides that may act as mediators in a variety of inflammatory diseases of the skin by interacting with specific receptors designated B1 and B2. In this study we have investigated the structure-activity relationship of intradermally injected bradykinin, kallidin (lysine-bradykinin), and [desArg9]-bradykinin in atopic (n = 8) and nonatopic (n = 8) subjects. METHODS: On two separate occasions, each separated by a week, subjects randomly underwent intradermal challenge with incremental doses (0.5, 5, and 50 nmol) of either the B1-agonist [desArg9]-bradykinin, the B2-agonists bradykinin or kallidin, or vehicle placebo. In a separate randomized double-blind study we have also examined the effect of an orally administered antihistamine, terfenadine, on kinin-induced wheal and flare responses and their repeatability in a group of nine volunteers. The skin responses were monitored objectively by measurement of wheal and flare areas. RESULTS: Both bradykinin and kallidin induced a dose-dependent increase in wheal and flare areas in all subjects studied. Although the effects of the two lowest doses (0.5 and 5 nmol) of [desArg9]-bradykinin on skin responses were indistinguishable from those of placebo, this kinin at the highest dose administered (50 nmol) caused a significant increase in wheal and flare areas in all subjects studied. No difference could be identified in the skin responses to kinins between atopic and nonatopic subjects. In addition kinin-induced cutaneous responses were not altered by pretreatment with terfenadine. CONCLUSIONS: These in vivo structure activity studies suggest that in human beings the skin responses to kinins may be compatible with the stimulation of B2 receptors, which is unrelated to histamine release from cutaneous mast cells.

Adult↗
PubMed1992

Intercellular adhesion molecule-1 (ICAM-1) and endothelial leucocyte adhesion molecule-1 (ELAM-1) expression in the bronchial mucosa of normal and asthmatic subjects.

Bronchial lavage and biopsy studies suggest the involvement of eosinophils and T-lymphocytes in allergic inflammation in asthma. There is evidence suggesting that the expression of adhesion molecules on endothelial cells and of their receptors on leucocytes is involved in this process. To investigate these mechanisms we have obtained bronchial mucosal biopsies from 10 normal subjects and from 10 symptomatic atopic asthmatics. Six of the asthmatics were re-biopsied after 6 weeks of inhaled beclomethasone dipropionate (BDP) during which time their clinical response was monitored. Frozen sections were stained by the immunoperoxidase method using monoclonal antibody (MoAb) 6.5B5 to identify expression of intercellular adhesion molecule (ICAM-1) and MoAb 1.2B6 for endothelial leucocyte adhesion molecule (ELAM-1). Araldite-embedded sections were also stained for eosinophils using MoAb EG2 to identify eosinophilic cationic protein (ECP). A significant mucosal eosinophilia was apparent in the asthmatic but not in the normal biopsies. Immunostaining for ICAM-1 was observed in both the epithelium and endothelium and ELAM-1 in endothelium, with no significant differences being apparent between the asthmatic and normal subjects. Topical BDP markedly reduced the mucosal eosinophilia without affecting the expression of either adhesion molecule. Using this method, we conclude that there is basal expression of ICAM-1 and ELAM-1 in normal human bronchial mucosa, which is not significantly different from that in asthmatics, and that it is insensitive to suppression with corticosteroids at an inhaled dose that causes clinical improvement.

Adult↗
PubMed1991

Endothelin immunoreactivity of airway epithelium in asthmatic patients.

There is extensive pharmacological and physiological evidence that endothelin-1 influences airway calibre. In mammals, endothelin receptor occur on airway smooth muscle, local storage and release of the peptide have been demonstrated, and inhalation of endothelin-1 induces bronchoconstriction. To investigate the relation between endothelins and asthma the expression of this peptide in endobronchial biopsy specimens was examined immunohistochemically with an antiserum against endothelin-1. Biopsy specimens from 17 asthmatic patients and 11 atopic and non-atopic healthy controls revealed striking differences, with endothelin expression being evident in airways epithelium and vascular endothelium in 11 of the 17 asthmatic patients but in only 1 of 11 controls. These results suggest that endothelins may play a part in the exaggerated bronchomotor tone of asthma.

Adolescent↗
PubMed1991

Airway inflammation and atopic asthma: a comparative bronchoscopic investigation.

Flexible fibre-optic bronchoscopy under local anaesthesia has been used to investigate the cellular airway events in atopic asthma. The findings have been compared to those from atopic individuals without asthma and non-atopic healthy controls, in an attempt to discern those changes relevant to clinical disease expression. Immunohistochemical and electron-microscopic analyses of airway biopsies identified that an atopic diathesis is associated with tissue eosinophil infiltration and mast cell degranulation. The eosinophilia was greatest in those atopic individuals with asthma. Flow-cytometric analysis of airway lavage revealed significantly enhanced T lymphocyte activation in clinical asthma. These findings are consistent with the hypothesis that T lymphocyte activation, through cytokine release, amplifies the tissue eosinophilia in asthma and that this combination is associated with clinical disease expression.

Asthma↗
PubMed1991

Mucosal nerves in endobronchial biopsies in asthma and non-asthma.

To investigate neural events within the airways in asthma, endobronchial biopsies were obtained by fibre-optic bronchoscopy from 8 atopic asthmatic subjects and 8 non-atopic healthy controls. The biopsies were immediately fixed on sampling and subsequently analysed for nerves using specific indirect immunofluorescence with antisera to the neural marker PGP 9.5 and to the neuropeptides vasoactive intestinal peptide (VIP), substance P (SP) and calcitonin gene-related peptide (CGRP). Nerves were present in all the biopsies from both subject groups, with no significant difference between the asthmatic and non-asthmatics. VIP-immunoreactive nerves were equally present in both subject groups, being localized to smooth muscle and glandular sites. No immunoreactive nerves to SP or CGRP could be identified in any biopsy at any location. These in vivo findings do not identify an anatomical neuronal imbalance in asthma.

Adult↗
PubMed1990

Myofibroblasts and subepithelial fibrosis in bronchial asthma.

A thickened bronchial epithelial basement membrane has long been regarded as a histopathologic characteristic of bronchial asthma. As we had previously demonstrated that this phenomenon is due to the deposition of interstitial collagens and fibronectin, we have now sought to determine the nature of the cell responsible for this process by studying endobronchial biopsies from eight normal and seven asthmatic volunteers by immunohistochemistry and electron microscopy. Biopsies were stained with PR 2D3, a monoclonal antibody to myofibroblasts of the pericrypt sheath of the colon and a monoclonal antibody to alpha-smooth muscle actin. The thickness of the subepithelial collagen and the organelle content of the cells therein were determined by electron microscopy. The subepithelial collagen thickness in the normal subjects ranged from 2.16 to 6.26 microns, while that in the asthmatic subjects ranged from 3.75 to 11.1 microns (Mann-Whitney test; P = 0.05). Elongated cells in the collagen layer were identified by staining with PR 2D3. As this antibody also stains smooth muscle, consecutive frozen sections were stained for alpha-smooth muscle actin and the number of positive cells per millimeter of basement membrane was subtracted from the count for PR 2D3. This yielded a count of 4.9 to 9.4 cells/mm in the normal subjects and 11.9 to 20.6 cells/mm in the asthmatics (P = 0.001). There was a highly significant correlation between the depth of subepithelial collagen and the number of PR 2D3-positive, alpha-smooth muscle actin-positive cells (Spearman rank correlation; r = 0.764 and P = 0.006). Electron microscopy confirmed the myofibroblastic nature of these cells.(ABSTRACT TRUNCATED AT 250 WORDS)

Actins↗
PubMed1989

Wheal-and-flare responses to intradermally injected adenosine 5'-monophosphate, hypertonic saline, and histamine: comparison of atopic and nonatopic subjects.

Adenosine 5'-monophosphate (AMP) in increasing concentrations, and saline solutions of corresponding tonicity, were injected intradermally in seven atopic and seven normal subjects. Skin wheal-and-flare responses were elicited in a dose-dependent fashion in all subjects, and no difference was found between responses produced by AMP and responses produced by saline of corresponding tonicity. Also, no difference in response to AMP and saline was found between atopic and nonatopic subjects. We further investigated, in seven atopic subjects, whether the skin wheal-and-flare response to the single, highest dose of AMP, saline, and histamine could be inhibited by preadministration of 180 mg of terfenadine, a potent H1 antagonist. A significant inhibition of the wheal-and-flare response to histamine and no significant inhibition to AMP were found. There was a significant inhibition of the flare response caused by hypertonic saline but no inhibition of the wheal response. We interpret these findings as indicating that AMP does not specifically lead to mast cell degranulation in the skin and that there are functional differences between cutaneous and lung mast cells. The observation that terfenadine significantly inhibited the flare response to hypertonic saline suggests that this stimulus produced histamine release.

Adenosine Monophosphate↗