Biomedical subjects

P S Hiemstra

Publications and source records attributed to P S Hiemstra.

At least 19 recordsLinked to original sources

PubMed1998

Cytokine release and its modulation by dexamethasone in whole blood following exercise.

Glucocorticoids (GC) play an important role in the treatment of inflammatory diseases like asthma. However, in selected patients a relative resistance to GC has been reported. Recently, it has been suggested that GC sensitivity of peripheral blood leucocytes may be regulated in a dynamic fashion during exercise, in association with activation of the hypothalamic-pituitary-adrenal (HPA) axis. The aim of the present study was to explore changes in the GC sensitivity of cytokine production by leucocytes following strenuous exercise by well trained oarsmen. These changes were studied using lipopolysaccharide (LPS)-induced and anti-CD2/anti-CD28 MoAb-stimulated cytokine release in whole blood and its modulation by dexamethasone. Following exercise, significant decreases in LPS-induced release of IL-6, tumour necrosis factor-alpha (TNF-alpha) and IL-10 and anti-CD2/anti-CD28 MoAb-stimulated secretion of interferon-gamma (IFN-gamma) were observed. In addition, the inhibitory effect of dexamethasone on both IL-6 and TNF-alpha secretion was significantly reduced following exercise, whereas that on IL-10 and IFN-gamma release was not affected. These exercise-induced changes were accompanied by activation of the HPA axis, as indicated by an increase in circulating adrenocorticotropic hormone (ACTH) levels immediately following exercise. The results from the present study suggest that GC sensitivity of whole blood cytokine release can be regulated in a dynamic fashion and that this can be assessed using an ex vivo stimulation assay. Moreover, since dexamethasone responsiveness of anti-CD2/anti-CD28 MoAb-induced IFN-gamma secretion in whole blood is not affected by exercise, it may suggest that exercise differentially affects monocytes and lymphocytes. The dynamic regulation of steroid responsiveness of leucocytes, as observed in the present study, could have important consequences for the effectiveness of GC treatment in inflammatory diseases.

Adolescent↗

PubMed1997

Effect of neutrophil serine proteinases and defensins on lung epithelial cells: modulation of cytotoxicity and IL-8 production.

Neutrophil accumulation in the lung may contribute to tissue injury as observed in inflammatory diseases. Both oxidative and non-oxidative mechanisms are involved in neutrophil-mediated tissue injury. Non-oxidative mechanisms include the release of neutrophil granule proteins such as the serine proteinases elastase and cathepsin G, and the non-enzymatic defensins. Because stimulated neutrophils are thought to release their products simultaneously, we investigated possible interactions between purified defensins and serine proteinases with respect to induction of cellular injury and their ability to induce interleukin-8 (IL-8) synthesis in cells of the lung epithelial cell line A549. Whereas defensins induced cell lysis, elastase and cathepsin G induced detachment of A549 cells. Co-incubation of elastase and cathepsin G revealed an additive effect on detachment, whereas defensins inhibited serine proteinase-induced detachment. Vice versa, both serine proteinases reduced defensin-induced cell lysis. Furthermore, elastase and cathepsin G prevented defensin-induced IL-8 synthesis. In contrast, no inhibitory interaction between cathepsin G and defensins was observed with respect to their antibacterial activity. The results from this study indicate that, at sites of inflammation, neutrophil-mediated injury might be regulated by interactions between released defensins and serine proteinases.

Blood Proteins↗

PubMed1997

An enzyme immunoassay for polymorphonuclear leucocyte-mediated fibrinogenolysis.

Upon stimulation, polymorphonuclear leucocytes (PMNs) release potent serine proteases, i.e. elastase, cathepsin G and proteinase 3, which contribute to the degradation of tissue and plasma components. Here, we describe the development of a plasma test to assess PMN-mediated fibrinogenolysis as a biochemical marker for actual PMN-derived proteolysis in vivo, useful for monitoring therapeutic efficacy, i.e. of elastase inhibitors. We generated a monoclonal antibody (MAb), designated 1-1/B3, with a high affinity for elastase-degraded fibrinogen (EDF). The epitope for 1-1/B3 becomes exposed in a time-dependent manner during digestion of fibrinogen with purified PMN-derived serine proteases and with isolated PMNs in vitro. However, 1-1/B3 does not react with plasma fibrinogen or with fibrin(ogen) degradation products generated by plasmin or by other active proteases that may occur locally, i.e. metalloproteases and lysosomal cathepsins. On the basis of MAb 1-1/B3, we developed a plasma test for the assessment of PMN-mediated fibrin(ogen) degradation products (PMN-FDP). In a panel of control plasmas, we observed concentrations of PMN-FDP of 8.2 +/- 0.9 ng mL-1 (n = 18). These values were increased twofold in patients with alpha 1-proteinase inhibitor deficiency (18.6 +/- 3.3 ng mL-1; n = 12; P < 0.0001) and even more in patients with sepsis (365.7 +/- 97.7 ng mL-1; n = 16; P < 0.0001). Furthermore, synovial tissue extracts from patients with rheumatoid arthritis contained increased levels of PMN-FDP, compared with synovial tissue extracts (P < 0.005) from patients with osteoarthritis.

Antibodies, Monoclonal↗

PubMed1997

Difference in binding of killed and live Streptococcus pneumoniae serotypes by C-reactive protein.

The binding and opsonic properties of C-reactive protein (CRP) for various species of bacteria were investigated. CRP bound more avidly to killed than to live Streptococcus pneumoniae, the binding varying among various serotypes; CRP hardly bound to a number of other bacterial species studied. CRP enhanced complement-dependent phagocytosis of live S. pneumoniae by granulocytes but did not enhance the phagocytosis of live Staphylococcus aureus or group B streptococci. We suppose that CRP may serve as an opsonin for killed bacteria and bacterial debris but is probably not an important opsonin for live bacteria other than S. pneumoniae.

Adult↗

PubMed1997

Antileukoprotease: an endogenous protein in the innate mucosal defense against fungi.

Previous studies have suggested that endogenous protease inhibitors may participate in the mucosal host defense. Antileukoprotease (ALP) is an important protease inhibitor found on various mucosal surfaces, including those of the respiratory and genital tracts. This study reports on the antimicrobial activity of recombinant (r) ALP toward the human fungal pathogens Aspergillus fumigatus and Candida albicans. rALP expressed pronounced fungicidal activity toward metabolically active A. fumigatus conidia and C. albicans yeast cells; however, metabolically quiescent A. fumigatus conidia were totally resistant. In contrast with the protease inhibitory activity of rALP, the fungicidal activity was localized primarily in the NH2-terminal domain. On a molar base, the fungicidal activity of rALP was comparable with that of human defensins and lysozyme. In addition, rALP caused inhibition of C. albicans yeast cell growth. By exhibiting antifungal activity, ALP may play an important role in the innate mucosal defense against human pathogenic fungi.

Antifungal Agents↗

PubMed1997

Proteases from Aspergillus fumigatus induce release of proinflammatory cytokines and cell detachment in airway epithelial cell lines.

Aspergillus fumigatus is a pathogen causing diverse respiratory disorders. Several studies have suggested that fungal proteases may play a role in the pathogenicity of fungi. Since the airways are the most common route for entry of A. fumigatus, this study focused on the ability of fungal proteases to induce the release of proinflammatory cytokines and to cause cell detachment in human pulmonary epithelial cell lines. It was shown that fungal serine protease activity induced the production of interleukin (IL)-8 and IL-6 and monocyte chemotactic protein-1 and caused cell detachment in a dose-dependent fashion. Chymostatin, antipain, phenylmethylsulfonyl fluoride, and heat treatment completely inhibited fungal protease activity, cytokine production and cell detachment; antileukoprotease partially inhibited these activities. By causing cell detachment, fungal proteases may decrease the physical barrier function of the epithelium; however, by eliciting a cytokine response, the epithelium may signal the mucosal inflammatory response against A. fumigatus.

Aspergillus fumigatus↗

PubMed1997

Bactericidal/permeability-increasing protein release in whole blood ex vivo: strong induction by lipopolysaccharide and tumor necrosis factor-alpha.

In this study, the release of bactericidal/permeability-increasing protein (BPI), which is stored in polymorphonuclear leukocytes (PMNL), was analyzed in a whole blood ex vivo system. Of the microbial products tested, lipopolysaccharide (LPS) most potently induced BPI release; FMLP, serum-treated zymosan (STZ), and lipoteichoic acid (LTA) also induced BPI release. In addition, the inflammatory mediator tumor necrosis factor (TNF)-alpha potently activated PMNL in whole blood, via TNF receptor p55, to release BPI, whereas interleukin (IL)-1, IL-8, platelet activating factor, and C5a were poor inducers of BPI release. STZ and phorbol myristate acetate, but not LPS, FMLP, or LTA, stimulated isolated PMNL to release BPI. BPI was released in comparable magnitude with the azurophilic granule protein elastase. Furthermore, both proteins were released with similar kinetics, which started within 30 min after onset of stimulation and lasted 1-4 h.

Antigens, CD↗

PubMed1997

Effect of defensins on interleukin-8 synthesis in airway epithelial cells.

Neutrophils play an important role in inflammatory processes in the lung and may cause tissue injury through, for example, release of proteinases such as neutrophil elastase. In addition to neutrophil elastase, stimulated neutrophils also release small nonenzymatic and cationic polypeptides termed defensins. The aim of the present study was to investigate whether defensins induce interleukin (IL)-8 expression in cells of the A549 lung epithelial cell line and in human primary bronchial epithelial cells (PBEC). Supernatants of defensin-treated A549 cells contained increased neutrophil chemotactic activity (16-fold) that was inhibited by antibodies against IL-8. Concurrently, within 3 and 6 h, defensins significantly increased the IL-8 levels in supernatants of both A549 cells (n = 6, P < 0.05 and P < 0.01, respectively) and PBEC (n = 4, P < 0.001 and P < 0.001, respectively). This defensin-induced increase was fully inhibited by the serine proteinase inhibitor alpha 1-proteinase inhibitor. In addition, defensins also increased IL-8 mRNA levels (12-fold); this increase was dependent on de novo mRNA synthesis and did not require protein synthesis. Furthermore, defensins did not affect IL-8 mRNA stability, indicating that the enhanced IL-8 expression was due to increased transcription. Our findings suggest that defensins, released by stimulated neutrophils, stimulate IL-8 synthesis by airway epithelial cells and thus may mediate the recruitment of additional neutrophils into the airways.

Aged↗

PubMed1997

Experimental rhinovirus 16 infection. Effects on cell differentials and soluble markers in sputum in asthmatic subjects.

Asthma exacerbations are often associated with respiratory virus infections, particularly with rhinovirus. In the present study we investigated the effect of experimental rhinovirus 16 (RV16) infection on airway inflammation as assessed by analysis of hypertonic saline-induced sputum. Twenty-seven nonsmoking atopic, mildly asthmatic subjects participated in a placebo-controlled parallel study. RV16 (n = 19) or its diluent (n = 8) was nasally administered. Sputum inductions were performed at entry and on Days 2 and 9 after inoculation, and airway responsiveness to histamine (PC20) was measured on Days 4 and 11. Cell differentials and levels of albumin, eosinophil cationic protein (ECP), IL-8, and IL-6 were determined. The cellular origin of IL-8 was investigated by intracellular staining. RV infection was confirmed by culture and/or by antibody titer rise in each of the RV16-treated subjects. There were no significant changes in the sputum differentials of nonsquamous cells (MANOVA, p > or = 0.40). In the RV16 group, there was a significant increase in the levels of ECP, IL-8, and IL-6 at Day 2 after infection (p < 0.05), whereas the albumin levels did not change (p = 0.82). The levels of IL-8 and IL-6 remained elevated for as long as 9 d after infection (p < 0.05). The increase in the percentage of IL-8 positive cells at Day 2 after infection could be attributed to the increase in IL-8 positive neutrophils (p < 0.02). There was a significant decrease in PC20 at Day 4 (p = 0.02), which was no longer significant at Day 11 (p = 0.19). The decrease in PC20 correlated significantly with the increase in ECP in the first week (r = -0.60) and with the change in the percentage eosinophils in the second week after inoculation (r = -0.58). We conclude that experimental RV16 infection in atopic asthmatic subjects increases airway hyperresponsiveness in conjunction with augmented airway inflammation, as reflected by an increase in ECP, IL-8, and IL-6 in sputum. Our results suggest that the RV16-enhanced airway hyperresponsiveness is associated with eosinophilic inflammation.

Adolescent↗

PubMed1997

Effects of photochemical air pollution and allergen exposure on upper respiratory tract inflammation in asthmatics.

Asthma is an inflammatory disease of the airways, and exacerbations of this disease have been associated with high levels of air pollution. The objective of this study was to examine whether ambient air pollution and/or allergen exposure induces inflammatory changes in the upper airways of asthmatics. Sixty patients with intermittent to severe persistent asthma visited the Hospital's Out Patient Clinic every 2 wk for a period of 3 mo, and on each visit a nasal lavage was obtained. Associations between nasal inflammatory parameters and seasonal allergens and/or air pollution exposures were analyzed using linear regression analysis. The study ran from July 3 to October 6, 1995, during which period ozone (8-h mean: 80 micrograms/m3) and PM10 (24-h mean: 40 micrograms/m3) were the major air pollutants; the major aeroallergen was mugwort pollen (24-h mean: 27 pollen grains/m3). Effects on both cellular and soluble markers in nasal lavage were demonstrated for both ozone and mugwort pollen, but not for PM10. Ambient ozone exposure was associated with an increase in neutrophils (112% per 100 micrograms/m3 increase in 8-h average ozone concentration), eosinophils (176%), epithelial cells (55%), IL-8 (22%), and eosinophil cationic protein (ECP) (19%). Increases in environmental mugwort pollen counts were associated with an increase in nasal eosinophils (107% per 100 pollen/m3) and ECP (23%), but not with neutrophils, epithelial cells, or lL-8. This study demonstrated that both ambient ozone and allergen exposure are associated with inflammatory responses in the upper airways of subjects with asthma, although the type of inflammation is qualitatively different.

Adolescent↗

PubMed1997

Effect of experimental rhinovirus 16 colds on airway hyperresponsiveness to histamine and interleukin-8 in nasal lavage in asthmatic subjects in vivo.

BACKGROUND: Asthma exacerbations are closely associated with respiratory virus infections. However, the pathophysiological consequences of such infections in asthma are largely unclear. OBJECTIVE: To examine the effect of rhinovirus 16 (RV16) infection on airway hypersensitivity to histamine, and on interleukin-8 (IL-8) in nasal lavage. METHODS: Twenty-seven non-smoking atopic, mildly asthmatic subjects participated in a placebo-controlled, parallel study. A dose of 0.5-2.9 x 10(4) TCID50 RV16 or placebo was nasally administered. Cold symptoms were recorded by questionnaire throughout the study. Histamine challenges were performed at entry, and on days 4 and 11 after inoculation. Nasal lavages were obtained at entry, and on days 2 and 9. The response to histamine was measured by PC20 (changes expressed as doubling doses: DD) IL-8 levels were obtained by ELISA, and were expressed in ng/ml. RESULTS: RV infection was confirmed by culture of nasal lavage and/or by antibody titre rise in each of the RV16-treated subjects. Among the 19 RV 16-treated subjects, eight developed severe cold symptoms. Baseline FEV1, did not change significantly during the study in either treatment group (P = 0.99). However, in the RV16-treated subjects there was a decrease in PC20 at day 4, which was most pronounced in those with a severe cold (mean change +/- SEM: -1.14 +/- 0.28 DD, P = 0.01). In addition, IL-8 levels increased in the RV16 group at days 2 and 9 (P < 0.001). The increase in nasal IL-8 at day 2 correlated significantly with the change in PC20 at day 4 (r = -0.48, P = 0.04). CONCLUSION: We conclude that the severity of cold, as induced by experimental RV16 infection, is a determinant of the increase in airway hypersensitivity to histamine in patients with asthma. Our results suggest that this may be mediated by an inflammatory mechanism, involving the release of chemokines such as IL-8.

Adolescent↗

PubMed1997

Chronic obstructive pulmonary disease: role of bronchiolar mast cells and macrophages.

Chronic obstructive pulmonary disease (COPD) is considered to be caused in part by smoking-induced inflammation, but it is unknown which inflammatory cells within the small airways are associated with the obstruction. We investigated the inflammatory infiltrate in the small airways of 16 current or ex-smokers with COPD (FEV1 < or = 75% predicted) and 15 without COPD (FEV1 > or = 85% predicted) in pneumectomy specimens that were removed for lung cancer. Mast cells, macrophages, neutrophils, eosinophils, T cells, and B cells were identified using immunohistochemistry on formalin-fixed, paraffin-embedded specimens. These cells were quantified in the epithelium and the remainder of the airway wall. The number of mast cells and macrophages in the epithelium, but not in the remainder of the airway wall, was significantly increased in patients with COPD. Neutrophil and T cell numbers did not differ between the groups. Only few B cells and eosinophils were present in both groups. Smoking history, perioperative steroid usage, tumor localization, or reversibility in the FEV1 to salbutamol could not account for the observed differences. We conclude that the number of epithelial mast cells and macrophages is increased in the bronchioli in smokers with airflow limitation, suggesting a role in development of COPD.

Adult↗

PubMed1996

Colony growth of human hematopoietic progenitor cells in the absence of serum is supported by a proteinase inhibitor identified as antileukoproteinase.

Serum contains many growth factors and nutrients that stimulate colony formation of hematopoietic progenitor cells (HPC) in semisolid cultures. In the absence of serum, no proliferation of HPCs could be obtained in semisolid medium cultures of partially purified bone marrow cells in the presence of multiple hematopoietic growth factors, insulin, cholesterol, and purified clinical-grade human albumin. This appeared to be due to a suppressive activity induced by monocyte- and T lymphocyte-depleted accessory cells on CD34+ HPCs. Serum-free conditioned medium from the bladder carcinoma cellline 5637 could replace serum to support the growth of HPCs in these cultures. After gel filtration and reverse-phase high-performance liquid chromatography of 5637 supernatants, this activity could be attributed to a 15-kD protein that was further identified by NH2-terminal sequence analysis as the serine proteinase inhibitor antileukoproteinase (ALP). The growth-supportive activity from the 5637 conditioned medium and the (partially) purified fractions could be completely neutralized by a polyclonal rabbit IgG antibody against human ALP (huALP). Similar supportive effects on the growth of HPC could be obtained in the presence of recombinant huALP. We demonstrated that the COOH-terminal domain of ALP containing the proteinase inhibitory activity was responsible for this effect. alpha-1 proteinase inhibitor was capable of similar support of in vitro HPC growth. These results illustrate that proteinase inhibitors play an important role in the in vitro growth of hematopoietic cells by the neutralization of proteinases produced by bone marrow accessory cells. This may be of particular relevance for in vitro expansion of human hematopoietic stem cells in serum-free media.

Amino Acid Sequence↗

PubMed1996

N-acetylcysteine pretreatment of cardiac surgery patients influences plasma neutrophil elastase and neutrophil influx in bronchoalveolar lavage fluid.

OBJECTIVE: Study of leukocyte activation and release of toxic mediators during extracorporeal circulation (ECC). ECC can be used to study the potential protective effect of a pharmacon against neutrophil-mediated lung injury. Clinical studies have indicated that N-acetylcysteine (NAC) may improve systemic oxygenation and reduce the need for ventilatory support when given to patients with acute lung injury. DESIGN: Cardiac surgery patients were pretreated with high-dose NAC in order to assess the potential role of NAC to interfere with neutrophil-mediated inflammation and lung injury. PATIENTS: 18 patients who underwent ECC: group 1 (n = 8) no premedication (only placebo); group 2 (n = 10) NAC (72 mg/kg i.v. as a bolus, later 72 mg/kg over 12 h). MEASUREMENTS AND RESULTS: In group 2, the partial pressure of oxygen in arterial blood/fractional inspired oxygen 4 h after surgery was significantly higher than in group 1 (213 +/- 31 vs 123 +/- 22; p = 0.044). NAC pretreatment prevented an increase in plasma neutrophil elastase activity (18.9 +/- 6.9 vs 49.9 +/- 5.6 ng/ml in group 1 at the end of ECC; p = 0.027). Release of myeloperoxidase (MPO) was not affected (group 1:1105 +/- 225 ng/ml vs group 2:1127 +/- 81 at the end of ECC; p = 0.63). At the end of ECC, total antigenic human neutrophil elastase (group 1:671 +/- 72 ng/ml vs group 2:579 +/- 134; p = 0.37) and complex formation between elastase and alpha 1-proteinase inhibitor were no different in the two groups. There were no significant difference in cellular composition and mediators in the lavage fluid, although values for total number of neutrophils, elastase, MPO and interleukin-8 were lower in group 2. CONCLUSION: Pretreatment with NAC may prevent lung injury by diminishing elastase activity. Since the release of mediators, especially MPO, is not affected, this diminished activity of elastase may be achieved by enhanced inactivation by antiproteases after initial treatment.

Acetylcysteine↗

PubMed1996

Regulation and production of IL-8 by human proximal tubular epithelial cells in vitro.

A number of inflammatory kidney diseases are associated with interstitial nephritis and influx of leucocytes in the renal interstitium. Potentially the influx of neutrophils in the interstitium may be induced by the chemotactic cytokine IL-8. In the present study we have analysed the production of IL-8 by cultured human proximal tubular epithelial cells (PTEC) in response to a number of proinflammatory cytokines. Primary cell lines of proximal tubular epithelium obtained from ten different kidneys, and cultured under serum-free conditions, were found to produce IL-8 to different degrees from not detectable levels up to 10.8 +/- 1.5 ng IL-8 per 1 x 10(5) cells in 72 h. Gel filtration chromatography of PTEC supernatant indicated that the size of IL-8 of PTEC is 15.1 and 8.1 kD, and is chemotactically active for polymorphonuclear neutrophils (PMN). Addition of 0.5 ng/ml rIL-1 alpha or 1000 U/ml recombinant tumour necrosis factor-alpha (rTNF-alpha) to the culture media of PTEC induced an up-regulation of IL-8 production up to 6.3-fold and 3.0-fold, respectively. The up-regulation by IL-1 alpha and TNF-alpha was dose- and time-dependent. In contrast, 500 U/ml recombinant interferon-gamma (rIFN-gamma) down-regulated the production of IL-8 3.4-fold. Northern blot analysis showed that IL-1 alpha and TNF-alpha increased the expression of IL-8 mRNA, whereas IFN-gamma reduced IL-8 mRNA expression. Taken together, these experiments indicate that human PTEC are a potential source of IL-8 in the kidney, and that IL-8 produced in the proximal tubule can be induced by various proinflammatory cytokines.

Cells, Cultured↗

PubMed1996

Antibacterial activity of antileukoprotease.

Antileukoprotease (ALP), or secretory leukocyte proteinase inhibitor, is an endogenous inhibitor of serine proteinases that is present in various external secretions. ALP, one of the major inhibitors of serine proteinases present in the human lung, is a potent reversible inhibitor of elastase and, to a lesser extent, of cathepsin G. In equine neutrophils, an antimicrobial polypeptide that has some of the characteristics of ALP has been identified (M. A. Couto, S. S. L. Harwig, J. S. Cullor, J. P. Hughes, and R. I. Lehrer, Infect. Immun. 60:5042-5047, 1992). This report, together with the cationic nature of ALP, led us to investigate the antimicrobial activity of ALP. ALP was shown to display marked in vitro antibacterial activity against Escherichia coli and Staphylococcus aureus. On a molar basis, the activity of ALP was lower than that of two other cationic antimicrobial polypeptides, lysozyme and defensin. ALP comprises two homologous domains: its proteinase-inhibitory activities are known to be located in the second COOH-terminal domain, and the function of its first NH2-terminal domain is largely unknown. Incubation of intact ALP or its isolated first domain with E. coli or S. aureus resulted in killing of these bacteria, whereas its second domain displayed very little antibacterial activity. Together these data suggest a putative antimicrobial role for the first domain of ALP and indicate that its antimicrobial activity may equip ALP to contribute to host defense against infection.

Anti-Bacterial Agents↗

PubMed1996

Role of YadA in resistance to killing of Yersinia enterocolitica by antimicrobial polypeptides of human granulocytes.

The virulence plasmid pYVe of Yersinia enterocolitica codes for the production of the outer membrane protein YadA and the secretion of several proteins, called Yops, which may protect this bacterium against killing by human granulocytes. Granulocytes kill ingested microorganisms by oxygen-dependent and oxygen-independent mechanisms, the latter including antimicrobial polypeptides. The aim of this study was to determine whether virulent (pYVe+) Y. enterocolitica and plasmid-cured avirulent (pYVe-) Y. enterocolitica differ in susceptibility to antimicrobial polypeptides extracted from granules of human granulocytes. The acetic acid granule extract contained several polypeptides with antimicrobial activity against Y. enterocolitica as determined by gel overlay and radial diffusion assays. Two of these polypeptides were identified as lysozyme and defensins. pYVe+ Y. enterocolitica was less susceptible than pYVe- Y. enterocolitica to the antimicrobial activity of granule extract, lysozyme, and defensins as determined in a suspension assay, which indicated that the pYVe plasmid mediates a reduced susceptibility to these polypeptides. The role of YadA in the resistance to antimicrobial polypeptides was analyzed by using mutants of Y. enterocolitica that specifically lack or express YadA. The results demonstrated that YadA conferred resistance to the killing of Y. enterocolitica by the granule extract. Together, these results indicate that the plasmid-encoded factor YadA contributes to the resistance of Y. enterocolitica to the killing by antimicrobial polypeptides of human granulocytes.

Adhesins, Bacterial↗

PubMed1996

Repeatability of cellular and soluble markers of inflammation in induced sputum from patients with asthma.

Sputum induced by inhalation of nebulized hypertonic saline is increasingly used to monitor airways inflammation in asthma. The aim of this study was to assess the repeatability of measuring cellular and soluble markers of inflammation in whole sputum samples as obtained by sputum induction in patients both with mild and moderate-to-severe asthma. Twelve patients with mild, atopic asthma without inhaled steroid treatment and nine patients with moderate-to-severe, atopic asthma treated with inhaled steroids were studied on two separate days at least 2 days apart. Whole sputum samples, induced by inhalation of hypertonic (4.5%) saline, were homogenized, and analysed for differential cell counts and for concentrations of albumin, fibrinogen, interleukin-8 (IL-8), and eosinophil cationic protein (ECP). Repeatability was expressed as intraclass correlation coefficient (Ri), and as coefficient of repeatability (CR) in percentage cells or in doubling concentration. Samples from two patients with mild asthma contained more than 80% squamous cells and were excluded from analysis. The repeatability for cell differential counts in both groups combined was: for neutrophils, Ri = 0.57 and CR = 31.0; for eosinophils, Ri = 0.85 and CR = 12.4; and for lymphocytes, Ri = 0.76 and CR = 6.9. The repeatability of the fluid phase measurements was: for albumin, Ri = 0.71 and CR = 3.2; for fibrinogen, Ri = 0.88 and CR = 2.8; for IL-8, Ri = 0.66 and CR = 2.2; and for ECP, Ri = 0.82 and CR = 1.1. We conclude that the repeatability of cellular and soluble markers of inflammation in induced sputum from patients with mild and moderate-to-severe asthma is satisfactory. Hence, induced sputum, processed by using the whole expectorated sample, seems to be a valuable method to monitor airway inflammation in asthma.

Adult↗