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1.
Health Sci Rep ; 7(5): e2082, 2024 May.
Article in English | MEDLINE | ID: mdl-38736472

ABSTRACT

Introduction: Across the globe, nurses and midwives play a crucial role in providing care to patients in healthcare facilities. They often contact the patient, providing direct care as directed by medical doctors or clinical officers. Traditionally, the role of nurses and midwives in the clinical diagnosis process is to coordinate the clinical diagnosis process-which includes laboratory diagnosis requests-from diagnosticians to the clinical laboratory. In these settings, these diagnosticians are general or specialist medical doctors. However, in some regions in sub-Saharan Africa (SSA), nurses and midwives are primary diagnosticians in healthcare facilities. Methodology: We present a perspective on the role of nurses and midwives in medical laboratory investigations in SSA. We highlight how, on top of nursing and midwifery roles, nurses take up the role of diagnosticians in facilities where doctors are few or are absent and what key issues are worth consideration. Furthermore, we present how efficient collaboration between nursing midwifery and medical laboratory diagnostic systems facilitates effective patient management. Conclusion: Emphasizing training on laboratory test utilization for nurses and midwives in SSA is vital for enhancing healthcare outcomes.

5.
Malawi Med J ; 31(4): 233-237, 2019 12.
Article in English | MEDLINE | ID: mdl-32133052

ABSTRACT

Background: Trypanosomes are protozoan flagellates that cause human African trypanosomiasis (HAT) and African animal trypanosomiasis (AAT). HAT is caused by Trypanosoma brucei rhodesiense in East and Central Africa and T.b. gambiense in West Africa, whereas AAT is caused by a number of trypanosome species, including T. brucei brucei, T. evansi, T. vivax, T. congolense, T. godfreyi and T. simiae. The aim of this study was to establish if tsetse flies at Liwonde Wild Life Reserve (LWLR) are infected with these trypanosomes and thus pose a risk to both humans and animals within and surrounding the LWLR. Methods: A total of 150 tsetse flies were caught. Of these, 82 remained alive after capture and were dissected such that the mid-gut could be examined microscopically for trypanosomes. DNA extractions were performed from both mid-guts and the 68 dead flies using a Qiagen Kit. Amplification techniques involved the Internal Transcriber Spacer 1 (ITS 1) conventional polymerase chain reaction (PCR) with primers designed to identify trypanosome species, and Repetitive Insertion Mobile Element - Loop Mediated Isothermal Amplification (RIME LAMP), a sequence specific to T. brucei. Results: Analysis showed that 79/82 (96.3%) of the mid-guts examined microscopically were positive for trypanosomes and that 75/150 (50%) of the DNA extracts (from the mid-gut, and tsetse fly carcasses) were positive for T. brucei, as determined by the RIME LAMP method. ITS1 PCR further showed that 87/150 (58.0%) flies were positive for trypanosomes, of which 56/87 (64.4%) were T. brucei, 9/87 (10.3%) were T. vivax; 7/87 (8.1%) were T. simiae; 6/87 (6.9%) were T. congolense, and 6/87 (6.9%) were T. godfreyi. Ten samples had a mixture of infections. Conclusion: Our analysis demonstrated a mixture of infections from trypanosome species in tsetse flies at LWLR, and that T. brucei, the species that causes HAT, was the most common. Our study successfully used molecular techniques to demonstrate the presence of T. b. rhodesiense at LWLR, a species that causes HAT in both East and Central Africa.


Subject(s)
Insect Vectors/parasitology , Polymerase Chain Reaction/methods , Trypanosoma/classification , Trypanosoma/genetics , Tsetse Flies/parasitology , Animals , DNA, Ribosomal/genetics , DNA, Ribosomal Spacer/genetics , Humans , Malawi , Molecular Epidemiology , Molecular Sequence Data , Trypanosoma/isolation & purification , Trypanosomiasis, African/epidemiology , Trypanosomiasis, African/parasitology , Trypanosomiasis, African/transmission
6.
Clin Exp Allergy ; 47(7): 890-899, 2017 Jul.
Article in English | MEDLINE | ID: mdl-28493293

ABSTRACT

BACKGROUND: Patients with severe asthma appear relatively corticosteroid resistant. Corticosteroid responsiveness is closely related to the degree of eosinophilic airway inflammation. The extent to which eosinophilic airway inflammation in severe asthma responds to treatment with systemic corticosteroids is not clear. OBJECTIVE: To relate the physiological and inflammatory response to systemic corticosteroids in asthma to disease severity and the baseline extent of eosinophilic inflammation. METHODS: Patients with mild/moderate and severe asthma were investigated before and after 2 weeks of oral prednisolone (Clintrials.gov NCT00331058 and NCT00327197). We pooled the results from two studies with common protocols. The US study contained two independent centres and the UK one independent centre. The effect of oral corticosteroids on FEV1 , Pc20, airway inflammation and serum cytokines was investigated. Baseline measurements were compared with healthy subjects. RESULTS: Thirty-two mild/moderate asthmatics, 50 severe asthmatics and 35 healthy subjects took part. At baseline, both groups of asthmatics had a lower FEV1 and Pc20 and increased eosinophilic inflammation compared to healthy subjects. The severe group had a lower FEV1 and more eosinophilic inflammation compared to mild/moderate asthmatics. Oral prednisolone caused a similar degree of suppression of eosinophilic inflammation in all compartments in both groups of asthmatics. There were small improvements in FEV1 and Pc20 for both mild/ moderate and severe asthmatics with a correlation between the baseline eosinophilic inflammation and the change in FEV1 . There was a ~50% reduction in the serum concentration of CXCL10 (IP-10), CCL22 (MDC), CCL17 (TARC), CCL-2 (MCP-1) and CCL-13 (MCP-4) in both asthma groups after oral corticosteroids. CONCLUSIONS AND CLINICAL RELEVANCE: Disease severity does not influence the response to systemic corticosteroids. The study does not therefore support the concept that severe asthma is associated with corticosteroid resistance. Only baseline eosinophilic inflammation was associated with the physiological response to corticosteroids, confirming the importance of measuring eosinophilic inflammation to guide corticosteroid use.


Subject(s)
Anti-Asthmatic Agents/administration & dosage , Asthma/drug therapy , Asthma/etiology , Eosinophils/immunology , Prednisolone/administration & dosage , Administration, Oral , Adrenal Cortex Hormones/administration & dosage , Adult , Asthma/diagnosis , Biomarkers , Cohort Studies , Cytokines/blood , Cytokines/metabolism , Eosinophils/metabolism , Eosinophils/pathology , Exhalation , Female , Humans , Male , Middle Aged , Nitric Oxide/metabolism , Respiratory Function Tests , Severity of Illness Index , Treatment Outcome , Young Adult
7.
Clin Exp Allergy ; 35(12): 1572-80, 2005 Dec.
Article in English | MEDLINE | ID: mdl-16393323

ABSTRACT

BACKGROUND: Chemokine receptors (CR) play an important role in T cell migration, but their contribution to lung trafficking is unclear. OBJECTIVE: We hypothesized that if a particular CR was involved in T cell homing its expression would be enriched on lung T cells compared with peripheral blood T cells (PBT). METHODS: We have measured the CR expression on BAL T cells from patients with sarcoid, other interstitial lung diseases (ILD), asthma and healthy volunteers. RESULTS: Of 14 CR studied in sarcoid, CXCR6 expression was the most markedly increased in the lung compared with the blood, a finding that was also seen in ILD patients. A striking although lesser increase was also seen in asthmatics and healthy controls. Analysis of expression of the CXCR6 ligand, CXCL16, by immunohistochemistry suggested that alveolar macrophages (AM) were the major source of CXCL16 in the lung. AM expressed mRNA for CXCL16 and released nanogram quantities after adhesion to plastic as shown by RT-PCR, Western blotting and ELISA. Bronchoalveolar lavage (BAL) fluid from all subjects contained large amounts of CXCL16. The full-length CXCL16 was the predominant isoform in AM lysates, supernatants and BAL. CONCLUSION: This data suggests that CXCR6 and CXCL16 may play a role in T cell recruitment to the lung.


Subject(s)
Chemokines, CXC/analysis , Lung Diseases/immunology , Lung/immunology , Receptors, Cytokine/analysis , Receptors, G-Protein-Coupled/analysis , Receptors, Scavenger/analysis , Receptors, Virus/analysis , T-Lymphocytes/chemistry , Adult , Asthma/immunology , Biomarkers/analysis , Blotting, Western , Bronchoalveolar Lavage Fluid/chemistry , Case-Control Studies , Chemokine CXCL16 , Chemokines, CXC/blood , Chemokines, CXC/genetics , Female , Flow Cytometry , Humans , Immunohistochemistry/methods , Lymphocyte Count , Macrophages, Alveolar/immunology , Male , Pulmonary Fibrosis/immunology , RNA, Messenger/analysis , Receptors, CXCR6 , Receptors, Chemokine , Receptors, Cytokine/blood , Receptors, Cytokine/genetics , Receptors, G-Protein-Coupled/blood , Receptors, G-Protein-Coupled/genetics , Receptors, Scavenger/blood , Receptors, Scavenger/genetics , Receptors, Virus/blood , Receptors, Virus/genetics , Sarcoidosis/immunology
8.
Thorax ; 58(12): 1066-70, 2003 Dec.
Article in English | MEDLINE | ID: mdl-14645977

ABSTRACT

BACKGROUND: We have recently reported a strong association between organ specific autoimmune disease and idiopathic chronic cough and have suggested that cough may be caused by airway inflammation secondary to aberrant homing of activated lymphocytes to the lung. An immunopathological study was undertaken to test the hypothesis that idiopathic chronic cough is associated with lymphocytic airway inflammation. METHODS: Bronchoscopy, bronchial biopsies, bronchoalveolar lavage (BAL), and peripheral blood and BAL flow cytometry were performed in 19 patients with idiopathic chronic cough, 14 with explained chronic cough, and 11 normal subjects. RESULTS: Organ specific autoimmune disease or positive autoantibodies were present in eight of the 19 patients with idiopathic cough, in one of the 14 patients with explained cough, and in one of the 11 normal subjects. Median BAL fluid differential lymphyocyte counts were significantly higher in patients with idiopathic cough (10.0%) than in normal subjects (6.3%, 95% confidence interval of difference 1.5 to 11.9, p = 0.01) or patients with explained cough (5.2%, 95% CI of difference 2.0 to 10.4, p = 0.001). There were no differences in bronchial biopsy T lymphocyte counts between the groups. The mean (SE) proportion of CD3+ peripheral blood mononuclear cells expressing CD4 was significantly higher in normal subjects than in patients with idiopathic cough (69 (3)% v 58 (3)%, mean difference 11%, 95% CI of difference 2 to 20, p<0.02) but not than those with explained chronic cough (63 (2)%). There were no differences in BAL T lymphocyte phenotype between groups. CONCLUSION: BAL fluid lymphocytosis occurs in some patients with idiopathic chronic cough. The association of idiopathic chronic cough with organ specific autoimmune disease raises the possibility that this might be caused by lymphocyte homing from the primary site of autoimmune inflammation or the result of an autoimmune process in the lung.


Subject(s)
Autoimmune Diseases/immunology , Bronchitis/immunology , Cough/immunology , Lymphocytosis/immunology , Autoantibodies/analysis , Autoimmune Diseases/pathology , Bronchitis/pathology , Bronchoalveolar Lavage Fluid/cytology , Bronchoalveolar Lavage Fluid/immunology , Chronic Disease , Cough/pathology , Female , Flow Cytometry , Humans , Immunohistochemistry , Lymphocyte Count , Male , Middle Aged , Sputum/cytology
9.
Clin Exp Allergy ; 33(12): 1711-6, 2003 Dec.
Article in English | MEDLINE | ID: mdl-14656359

ABSTRACT

BACKGROUND: Airway smooth muscle infiltration by mast cells is a feature of asthma and not eosinophilic bronchitis. In asthma, Th2 cytokines have been implicated as playing a critical role in the development of airway inflammation and hyper-responsiveness. Whether inflammatory cells within the airway smooth muscle release these cytokines is unknown. METHODS: We have undertaken a comparative immunohistochemical study in bronchial biopsies from 14 subjects with asthma, 10 with eosinophilic bronchitis and eight normal controls recruited from two centres. RESULTS: The median number of IL-4+ cells/mm2 smooth muscle was significantly higher in subjects with asthma than eosinophilic bronchitis and normal controls for both the anti-IL-4 mAb 3H4 (2.4, 0, 0, respectively; P=0.001) and anti-IL-4 mAb 4D9 (1.6, 0, 0, respectively; P=0.02). There were no group differences in the number of IL-5+ cells (P=0.31). In six subjects with asthma, IL-13 expression by cells within the airway smooth muscle was studied. The median (range) of IL-13+cells was 2 (0.9-2.7). Ninety-four percent of the cells expressing IL-4 (3H4), 92% of those expressing IL-4 (4D9) and 100% expressing IL-13 in the airway smooth muscle were mast cells. Fifty-five percent of the mast cells within the airway smooth muscle co-localized to IL-4 (3H4), 29% to IL-4 (4D9) and 17% to IL-13. CONCLUSIONS: In asthma, IL-4+ and IL-13+ cells were present within the airway smooth muscle and were expressed predominantly by mast cells, suggesting that IL-4 and IL-13 may play an important role in mast cell-airway smooth muscle interactions.


Subject(s)
Asthma/immunology , Bronchi/immunology , Interleukin-13/analysis , Interleukin-4/analysis , Mast Cells/immunology , Muscle, Smooth/immunology , Adult , Bronchitis/immunology , Case-Control Studies , Eosinophils/immunology , Female , Humans , Immunohistochemistry/methods , Interleukin-5/immunology , Male , Middle Aged , Statistics, Nonparametric , Th2 Cells/immunology
10.
Thorax ; 58(6): 528-32, 2003 Jun.
Article in English | MEDLINE | ID: mdl-12775868

ABSTRACT

BACKGROUND: Eosinophilic bronchitis is a condition characterised by a corticosteroid responsive cough, sputum eosinophilia, and normal tests of variable airflow obstruction and airway responsiveness. We performed a detailed comparative immunopathological study to test the hypothesis that the different airway function in patients with eosinophilic bronchitis and asthma reflects differences in the nature of the lower airway inflammatory response. METHODS: Exhaled nitric oxide was measured and induced sputum, bronchoscopy, bronchial wash (BW), bronchoalveolar lavage (BAL), and bronchial biopsy were performed in 16 subjects with eosinophilic bronchitis, 15 with asthma, and 14 normal controls. RESULTS: Both eosinophilic bronchitis and asthma were characterised by an induced sputum, BW and BAL eosinophilia, an increased number of epithelial and subepithelial eosinophils, and increased reticular basement membrane thickness. The median concentration of exhaled nitric oxide was higher in those with eosinophilic bronchitis (12 ppb) or asthma (8.5 ppb) than normal controls (2 ppb) (95% CI of the difference 5 to 16, p<0.0001 and 2 to 11.3, p=0.004, respectively). There were no group differences in epithelial integrity or the number of subepithelial T lymphocytes, mast cells or macrophages. CONCLUSION: With the exception of our previously reported association of smooth muscle mast cell infiltration with asthma, the immunopathology of eosinophilic bronchitis and asthma are similar which suggests that eosinophilic airway inflammation, increased exhaled nitric oxide, and increased basement membrane thickening are regulated independently of airway hyperresponsiveness.


Subject(s)
Asthma/pathology , Bronchitis/pathology , Eosinophilia/pathology , Basement Membrane/pathology , Bronchoalveolar Lavage Fluid , Eosinophils , Female , Humans , Immunohistochemistry , Male , Middle Aged , Nitric Oxide/analysis , Sputum/chemistry
11.
Thorax ; 57(12): 1054-9, 2002 Dec.
Article in English | MEDLINE | ID: mdl-12454301

ABSTRACT

BACKGROUND: The lung is an important tertiary lymphoid organ and many lung diseases are associated with disordered lung immunity. Unlike the gut (alpha4beta7 binding to MAdCAM-1) and skin (CLA+ve T cells binding to E-selectin) where the adhesion receptors involved in organ specific homing of T cells have been identified, the molecular pathways controlling lymphocyte migration to the lung are unclear. Using a modified version of the Stamper-Woodruff assay we have investigated the receptors mediating adhesion of peripheral blood lymphocytes to airway endothelium. METHODS: Longitudinal frozen sections of bronchus (8 micro m) obtained from lung resection specimens were incubated with T cell enriched peripheral blood mononuclear cells for 30 minutes under shaking conditions in the presence of a fluorescently labelled polyclonal anti-von Willebrand antibody to identify blood vessels. After fixation the percentage of blood vessels supporting adhesion was measured. Blocking monoclonal antibodies were used to determine the role of individual adhesion receptors in lymphocyte binding. RESULTS: Specific cation dependent binding of lymphocytes to bronchial endothelium was observed which was significantly inhibited by antibodies against P-selectin, PSGL-1, L-selectin, LFA-1, ICAM-1 and ICAM-2 but not E-selectin, VLA-4, VCAM-1 or Mac-1. This was consistent with the pattern of endothelial expression of these receptors with strong expression of P-selectin and ICAM-1, but negligible expression of E-selectin on bronchial endothelium. CONCLUSION: This study suggests an important role for PSGL-1/P-selectin in T cell migration into the bronchi and provides further evidence for a pattern of recirculation for respiratory tract homing T cells distinct from the gut and skin.


Subject(s)
Bronchi/chemistry , Membrane Glycoproteins/metabolism , P-Selectin/metabolism , T-Lymphocytes/physiology , Bronchi/immunology , Cell Adhesion , Cell Movement , Endothelium/chemistry , Endothelium/immunology , Humans , Immunohistochemistry/methods , Intercellular Adhesion Molecule-1/metabolism , Receptors, Lymphocyte Homing/metabolism , T-Lymphocytes/immunology , Vascular Cell Adhesion Molecule-1/metabolism
12.
Thorax ; 57(1): 91-2, 2002 Jan.
Article in English | MEDLINE | ID: mdl-11809998

ABSTRACT

Chronic cough is a common reason for presentation to a respiratory clinic. In up to 20% of cases the cause remains unclear after investigations. We report one such case where there was bronchoscopic evidence of lymphocytic airway inflammation in association with newly diagnosed coeliac disease. All features improved markedly on a gluten free diet, suggesting a causal relationship between coeliac disease, cough, and lymphocytic bronchoalveolitis.


Subject(s)
Bronchitis/diet therapy , Celiac Disease/diet therapy , Cough/diet therapy , Lymphoproliferative Disorders/diet therapy , Pneumonia/diet therapy , Aged , Bronchitis/etiology , Celiac Disease/etiology , Chronic Disease , Cough/etiology , Diet, Protein-Restricted/methods , Glutens , Humans , Lymphoproliferative Disorders/etiology , Male , Pneumonia/etiology , Pulmonary Alveoli , Treatment Outcome
13.
J Immunol ; 166(4): 2842-8, 2001 Feb 15.
Article in English | MEDLINE | ID: mdl-11160352

ABSTRACT

The lung is an important tertiary lymphoid organ with constant trafficking of T cells through the lung in both health and disease. T cell migration is controlled by a combination of adhesion receptors and chemokines expressed on vascular endothelium and in the tissue, often in an organ-specific manner. This leads to selective accumulation of different T cell subsets, a process called lymphocyte homing. There is evidence for a distinct lung-homing pathway, but no specific lung-homing receptors have been described. We analyzed the chemokine receptor profile of lung T cells to determine the extent to which lung T cells shared homing pathways with other organs such as the gut and skin. In addition, we compared expression of receptors in normal and asthmatic individuals to determine whether different pathways were used in health and disease. We observed that lung T cells expressed a profile of chemokine and adhesion receptors distinct from that of gut- and skin-homing T cells although no chemokine receptor specific for the lung was found. In particular, lung T cells expressed CCR5 and CXCR3, but not CCR9 or cutaneous lymphocyte Ag, and only low levels of CCR4 and alpha(4)beta(7). No differences were observed between lung T cells from normal vs asthmatic subjects. This study provides added support for the concept of a lung-homing pathway separate from other mucosal organs such as the gut and suggests that the chemokine pathways that control T cell migration in normal homeostasis and Th2-type inflammatory responses are similar.


Subject(s)
Asthma/immunology , Lung/immunology , Lung/metabolism , Receptors, Chemokine/biosynthesis , T-Lymphocyte Subsets/immunology , T-Lymphocyte Subsets/metabolism , Antigens, Differentiation, T-Lymphocyte/biosynthesis , Asthma/metabolism , Bronchoalveolar Lavage Fluid/cytology , Bronchoalveolar Lavage Fluid/immunology , CD4-Positive T-Lymphocytes/immunology , CD4-Positive T-Lymphocytes/metabolism , CD8-Positive T-Lymphocytes/immunology , CD8-Positive T-Lymphocytes/metabolism , Cell Movement/immunology , Cell Separation , Humans , Immunophenotyping , Lung/pathology , Lung/surgery , Pneumonectomy , Receptors, Lymphocyte Homing/biosynthesis , Th2 Cells/immunology , Th2 Cells/metabolism
14.
Blood ; 95(10): 3146-52, 2000 May 15.
Article in English | MEDLINE | ID: mdl-10807781

ABSTRACT

Selective eosinophil accumulation is a hallmark of diseases such as asthma. In a model of chronic eosinophilic inflammation, we have previously shown that the tethering step in eosinophil adhesion is mediated by PSGL-1 binding to P-selectin. The Th2-associated cytokine IL-13 is of potential importance in allergic disease. We have therefore investigated whether IL-13 can mediate eosinophil binding to human umbilical vein endothelial cells (HUVEC) through P-selectin. IL-13 caused dose- and time-dependent increases of P-selectin expression, as assessed by flow and laser scanning cytometry. A similar degree of expression was observed with IL-4. There was no effect on E-selectin or ICAM-1 expression. Tumor necrosis factor-alpha induced the expression of VCAM-1, E-selectin, and ICAM-1 but had no effect on P-selectin expression. IL-13 increased the production of mRNA for surface and soluble variants of P-selectin. Under flow conditions, eosinophils, but not neutrophils, showed enhanced binding to IL-13 and to IL-4-stimulated HUVEC compared to medium-cultured cells. Eosinophil adhesion was completely inhibited by a blocking monoclonal antibody against PSGL-1 and P-selectin. Anti-VLA-4 and anti-VCAM-1 antibodies inhibited binding to a lesser extent. Thus, at physiologic levels of expression induced by Th2 cytokines, P-selectin/PSGL-1 supported eosinophil but not neutrophil adhesion. This mechanism is likely to be a key event leading to the selective accumulation of eosinophils in allergic inflammation.


Subject(s)
Endothelium, Vascular/cytology , Endothelium, Vascular/physiology , Eosinophils/physiology , Interleukin-13/pharmacology , Membrane Glycoproteins/physiology , Neutrophils/physiology , P-Selectin/physiology , Animals , Cell Adhesion/drug effects , Cell Adhesion/physiology , Cell Differentiation , Cells, Cultured , Dose-Response Relationship, Drug , Eosinophils/cytology , Humans , Interleukin-13/physiology , Ligands , Mice , Neutrophils/cytology
15.
Clin Exp Allergy ; 30(2): 255-63, 2000 Feb.
Article in English | MEDLINE | ID: mdl-10651778

ABSTRACT

BACKGROUND: Eosinophil-bronchial epithelial cell interactions are thought to be central to the pathogenesis of asthma, both in terms of the epithelium as a source of pro-inflammatory mediators and as a target for eosinophil-mediated damage. We have therefore investigated adhesion interactions between these two cell types. OBJECTIVES: To determine the role of eosinophil and epithelial activation on eosinophil adhesion to bronchial epithelium and to characterize the adhesion receptors mediating eosinophil adhesion. METHODS: Eosinophils were purified from human peripheral blood by immunomagnetic selection and adhesion to confluent cultures of the airway epithelial cell lines A549 and BEAS-2B was studied. RESULTS: Stimulation of A549 cells with TNFalpha, IFNgamma or a combination of 50 ng/mL of TNFalpha, IFNgamma and IL-1 (cytomix) did not effect eosinophil binding despite an increase in ICAM-1 expression. Similarly stimulation of eosinophils with PAF or IL-5 had no effect on eosinophil binding to medium- or cytokine-treated A549 cells. In contrast stimulation of BEAS-2B cells with cytomix caused a significant increase in eosinophil adhesion. This was associated with an increase in expression of ICAM-1 and induced expression of VCAM-1. Treatment of eosinophils with Mn2+ and IL-5 but not eotaxin, RANTES or PAF also significantly enhanced eosinophil adhesion to medium-treated BEAS-2B cells. Using blocking mAbs we were able to demonstrate that the increased adhesion resulting from stimulation of eosinophils or BEAS-2B cells was in both cases mediated by a combination of CD18 and alpha4 integrins. CONCLUSIONS: This study demonstrates a selective role for IL-5 in mediating integrin-dependent eosinophil adhesion to airway epithelium and once again emphasizes the importance of this cytokine in controlling eosinophil activation in diseases such as asthma.


Subject(s)
Bronchi/cytology , Cell Adhesion/drug effects , Eosinophils/physiology , Interleukin-5/pharmacology , Respiratory Mucosa/cytology , Antigens, CD/metabolism , Bronchi/physiology , Cell Adhesion Molecules/metabolism , Cell Line , Chemotactic Factors/pharmacology , Enzyme-Linked Immunosorbent Assay , Flow Cytometry , Humans , Respiratory Mucosa/physiology , Tumor Cells, Cultured , Vascular Cell Adhesion Molecule-1/metabolism
16.
Am J Respir Cell Mol Biol ; 20(6): 1251-9, 1999 Jun.
Article in English | MEDLINE | ID: mdl-10340944

ABSTRACT

We have used the Stamper-Woodruff frozen-section assay (FSA) to characterize the integrin and activation steps involved in adhesion of peripheral blood eosinophils and neutrophils to nasal polyp endothelium (NPE). Eosinophil and neutrophil adhesion was significantly inhibited by monoclonal antibodies (mAbs) against CD18 (beta2) and CD11a-c. Eosinophil adhesion was also inhibited to a lesser extent by mAbs against CD29 (beta1), CD49d (alpha4), and vascular cell adhesion molecule-1. The involvement of integrins raised the possibility of an activation step being involved in the adhesion process. Although stimulation of the cells with granulocyte macrophage colony-stimulating factor (GM-CSF) before the assay failed to modulate adhesion, binding was inhibited by up to 50% by treatment of the leukocytes with azide. In addition, neutrophil adhesion was completely abrogated by pertussis toxin (PT) and inhibited by about 50% by the platelet-activating factor antagonist WEB 2086 and antibodies against interleukin (IL)-8 and the two IL-8 receptors IL8RA and IL8RB (C-X-CR1 and -CR2). In contrast, eosinophil adhesion was unaffected by PT, WEB 2086, or anti-IL8R mAbs. mAbs against CCR-3, IL-3, IL-5, and GM-CSF also had no effect. This study demonstrates that eosinophil and neutrophil adhesion to NPE in the FSA conforms to the multistep paradigm for leukocyte adhesion and can be used to model the molecular basis for adhesion to endothelium in the context of chronic inflammatory disease. Using this assay, we have observed significant differences in integrin usage between eosinophils and neutrophils and a striking difference in the mechanism of integrin activation. These differences could explain, in part, the preferential accumulation of eosinophils in diseases such as asthma.


Subject(s)
Eosinophils/metabolism , Epithelium/physiology , Inflammation , Integrins/immunology , Integrins/metabolism , Neutrophils/metabolism , CD11 Antigens/metabolism , CD18 Antigens/metabolism , Cell Adhesion , Endothelium/metabolism , Eosinophils/immunology , GTP-Binding Proteins/metabolism , Granulocyte-Macrophage Colony-Stimulating Factor/immunology , Humans , Interleukin-3/immunology , Interleukin-5/immunology , Interleukin-8/pharmacology , Nasal Polyps/metabolism , Neutrophils/immunology , Pertussis Toxin , Platelet Activating Factor/pharmacology , Receptors, CCR3 , Receptors, Chemokine/immunology , Virulence Factors, Bordetella/pharmacology
17.
Thorax ; 54(2): 124-30, 1999 Feb.
Article in English | MEDLINE | ID: mdl-10325916

ABSTRACT

BACKGROUND: Sputum induction is an important non-invasive technique for measuring airway inflammation in asthma. Cell numbers are often too low for flow cytometric analysis. Laser scanning cytometry (LSC) is a novel technique that allows objective multicolour fluorescence analysis of cells on a microscope slide. METHODS: LSC was used to determine sputum eosinophil and bronchial epithelial cell counts. We first confirmed that we could measure eosinophil counts accurately in peripheral blood using alpha-major basic protein (MBP) immunofluorescent staining. Sputum induction was performed according to standard protocols. Sputum samples from eight normal controls and 12 asthmatic patients were analysed by LSC and manual counting by two independent observers. Octospot cytospins were fixed and stained with mouse-alpha-human-MBP monoclonal antibody or mouse-alpha-human-cytokeratin antibody and goat-alpha-mouse Oregon Green conjugated second antibody. RESULTS: Sputum induction provided a mean (SE) of 0.99 (0.2) x 10(6) cells per donor. More than 3000 cells on three cytospins per slide were analysed per cell type. The intraclass correlation coefficient (R) and standard deviation (SD) of differences in eosinophils determined by manual counting and LSC were 0.9 and 2.1, respectively, and for bronchial epithelial cell counts they were 0.7 and 2.0. Selective detection of labelled cells was confirmed visually after relocation. CONCLUSION: Eosinophils and bronchial epithelial cells can be accurately and reproducibly counted in an objective manner. LSC is therefore a potentially powerful new method for immunophenotyping leucocytes and epithelial cells objectively in induced sputum in patients with asthma.


Subject(s)
Asthma/pathology , Eosinophils/pathology , Sputum/cytology , Bronchi/pathology , Cell Count , Flow Cytometry/methods , Humans , Lasers , Microscopy, Fluorescence
18.
Eur J Immunol ; 29(4): 1324-33, 1999 04.
Article in English | MEDLINE | ID: mdl-10229100

ABSTRACT

The inflammatory process that underlies allergic diseases such as asthma is characterized by tissue infiltration of eosinophils and T cells. We have used the Stamper-Woodruff frozen-section assay to characterize the receptors involved in adhesion of human peripheral blood T cells to nasal polyp endothelium (NPE) as a model of T cell migration in allergic disease. T cells bound specifically to NPE in a temperature-, cell concentration- and shear stress-dependent fashion. Adhesion was inhibited by approximately 70% by antibodies against P-selectin and its counter-receptor P-selectin glycoprotein-1 (PSGL-1). In addition, a blocking monoclonal antibody (mAb) against L-selectin caused significant although lesser inhibition. Cells adhering to NPE were primarily of the CD45RO+ memory subset. Although only a minority subset of peripheral blood T cells expressed functional PSGL-1, as determined by binding of a P-selectin Fc chimera, the majority of the P-selectin chimera-binding cells were found to be CD45RO+. This is consistent with the observation that memory T cells bind to NPE via P-selectin. Using blocking mAb we also investigated which integrins and their counter-structures were involved in T cell binding. A combination of anti-beta1 and beta2 mAb was able to inhibit adhesion by almost 50%. An antibody against intercellular adhesion molecule (ICAM)-2 gave an inhibition similar to that by anti-CD18 mAb, suggesting ICAM-2 was the major counter-receptor involved for the beta2 integrin component. This study suggests that P-selectin, and to a lesser extent L-selectin, may be acting as specific homing receptors for the airway mucosa in the context of chronic allergic disease.


Subject(s)
L-Selectin/physiology , Nasal Mucosa/immunology , Nasal Polyps/immunology , P-Selectin/physiology , T-Lymphocytes/physiology , Cell Adhesion , Chronic Disease , Endothelium/immunology , Humans , Integrins/physiology , Leukocyte Common Antigens/analysis , Membrane Glycoproteins/physiology , Receptors, Lymphocyte Homing/physiology
19.
Thorax ; 51(11): 1155-7, 1996 Nov.
Article in English | MEDLINE | ID: mdl-8958901

ABSTRACT

Selectins are adhesion receptors expressed by leucocytes, platelets, and endothelial cells. They mediate the initial binding of leucocytes to vascular endothelium in the post-capillary venules. This is an essential first step in leucocyte migration into tissue. The selectin family of adhesion receptors consists of three C-type lectins (E, P, and L selectin). Their ligands (counter structures) are sialylated and fucosylated carbohydrate molecules which, in most cases, decorate mucin-like glycoprotein membrane receptors. Studies using blocking monoclonal antibodies have shown that inhibition of selectin function can ameliorate a range of inflammatory processes, offering the possibility that antagonists of selectin function may be useful in the treatment of inflammatory lung diseases such as asthma.


Subject(s)
Lung Diseases/therapy , Selectins/physiology , Animals , Antibodies, Monoclonal , Humans , Leukocytes/physiology , Lung Diseases/metabolism , Mice , Mice, Knockout
20.
Ann N Y Acad Sci ; 796: 124-37, 1996 Oct 31.
Article in English | MEDLINE | ID: mdl-8906219

ABSTRACT

Considerable progress has been made in our understanding of the molecular mechanisms involved in eosinophil migration into sites of allergic inflammation. A number of differences between eosinophils and neutrophils have been observed in their pattern of adhesion interactions. These include expression of alpha 4 beta 1, alpha 4 beta 7, and alpha 6 beta 1 by eosinophils but not neutrophils and structural differences between eosinophil and neutrophil PSGL-1 associated with increased eosinophil binding to P-selectin. On the basis of current evidence the various receptors and mediators involved are summarized in FIGURE 1. Once in the tissues eosinophils may persist for several days or weeks, surviving under the influence of locally generated cytokines and this persistence may also partly explain selective tissue accumulation of eosinophils. Understanding the molecular mechanisms involved in leucocyte migration offers the possibility of selective and effective antagonists to treat allergic disease by preventing cell migration. Results in a number of animal models already offer the hope that this approach may be successful. The development of drugs that can be tested in the clinic are awaited with considerable interest.


Subject(s)
Endothelium, Vascular/metabolism , Eosinophils/metabolism , Animals , Asthma/metabolism , Cell Adhesion , Chemotaxis, Leukocyte , Endothelium, Vascular/immunology , Eosinophils/immunology , Extracellular Matrix/metabolism , Integrins/physiology , Intercellular Adhesion Molecule-1/metabolism , Mice , Models, Biological , Selectins/metabolism , Vascular Cell Adhesion Molecule-1/metabolism
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