Journal Article Interaction of Nontypable Haemophilus influenzae with Human Respiratory Mucosa In Vitro Get access Robert C. Read, Robert C. Read Host Defence Unitand Department of Lung Pathology, National Heart and Lung Institute, Royal Brompton and National Heart Hospital; Institute of Otolaryngology; and Electron Microscopy Unit, Kings College, London Search for other works by this author on: Oxford Academic PubMed Google Scholar Robert Wilson, Robert Wilson Host Defence Unitand Department of Lung Pathology, National Heart and Lung Institute, Royal Brompton and National Heart Hospital; Institute of Otolaryngology; and Electron Microscopy Unit, Kings College, London Search for other works by this author on: Oxford Academic PubMed Google Scholar Andrew Rutman, Andrew Rutman Host Defence Unitand Department of Lung Pathology, National Heart and Lung Institute, Royal Brompton and National Heart Hospital; Institute of Otolaryngology; and Electron Microscopy Unit, Kings College, London Search for other works by this author on: Oxford Academic PubMed Google Scholar Valerie Lund, Valerie Lund Host Defence Unitand Department of Lung Pathology, National Heart and Lung Institute, Royal Brompton and National Heart Hospital; Institute of Otolaryngology; and Electron Microscopy Unit, Kings College, London Search for other works by this author on: Oxford Academic PubMed Google Scholar Howard C. Todd, Howard C. Todd Host Defence Unitand Department of Lung Pathology, National Heart and Lung Institute, Royal Brompton and National Heart Hospital; Institute of Otolaryngology; and Electron Microscopy Unit, Kings College, London Search for other works by this author on: Oxford Academic PubMed Google Scholar Anthony P. R. Brain, Anthony P. R. Brain Host Defence Unitand Department of Lung Pathology, National Heart and Lung Institute, Royal Brompton and National Heart Hospital; Institute of Otolaryngology; and Electron Microscopy Unit, Kings College, London Search for other works by this author on: Oxford Academic PubMed Google Scholar Peter K. Jeffery, Peter K. Jeffery Host Defence Unitand Department of Lung Pathology, National Heart and Lung Institute, Royal Brompton and National Heart Hospital; Institute of Otolaryngology; and Electron Microscopy Unit, Kings College, London Search for other works by this author on: Oxford Academic PubMed Google Scholar Peter J. Cole Peter J. Cole Host Defence Unitand Department of Lung Pathology, National Heart and Lung Institute, Royal Brompton and National Heart Hospital; Institute of Otolaryngology; and Electron Microscopy Unit, Kings College, London Reprints or correspondence: Dr. Peter Cole, Host Defence Unit, National Heart & Lung Institute, Manresa Rd., London SW3 6LR, UK. Search for other works by this author on: Oxford Academic PubMed Google Scholar The Journal of Infectious Diseases, Volume 163, Issue 3, March 1991, Pages 549–558, https://doi.org/10.1093/infdis/163.3.549 Published: 01 March 1991 Article history Received: 06 March 1990 Revision received: 24 August 1990 Published: 01 March 1991
Pseudomonas aeruginosa colonizes the lower respiratory tracts of patients with severe bronchiectasis, including cystic fibrosis, a condition associated with increased airway mucus output. We have shown that an extract containing chloroform-soluble extracellular products of P. aeruginosa releases glycoconjugates into the cat trachea in vivo. This activity was not related to pyocyanin, a major component of the extract, but was associated with the rhamnolipids. Purified monorhamnolipid (100 µg/ml) released radiolabeled and periodic acid-Schiff (PAS)-reactive glycoconjugates (Δ3H = +490 ± 70%, Δ35S = +170 ± 40%, ΔPAS = +86 ± 1.7 µ/min; n = 6, P < 0.02 for each). Dirhamnolipid (200 µg/ml) was also effective (Δ3H = +640 ± 70%, Δ35S = +130 ± 20%, ΔPAS = +9.3 ± 1.5 µg/min; n = 6, P < 0.02 for each). Monorhamnolipid (100 µg/ml) also released 35S-labeled and PAS-reactive glycoconjugates from human bronchial tissue in vitro (Δ35S = +189 ± 47%, ΔPAS = +26.3 ± 8.5 µg/min; n = 7, P < 0.001 versus control tissues in which no stimulus was given). The cat tracheal glycoconjugates released by the rhamnolipids differed from those released by pilocarpine 50 µM, in having a higher 3H:35S ratio (P < 0.001). After gel chromatography on a Sepharose CL-4B column, the void volume fractions of the glycoconjugates also had different profiles in a cesium chloride density gradient. Those released by rhamnolipid banded at 1.62 g/ml, while those released by pilocarpine banded mainly at 1.50 g/ml, with some of the higher density material also present. The rhamnolipids released glycoconjugates without causing ultra-structural damage to the cat tracheal epithelium, as shown by transmission electron microscopy. Monorhamnolipid, at a concentration similar to those releasing glycoconjugates in the cat trachea, was present in lung secretions from a patient with cystic fibrosis, colonized by P. aeruginosa, undergoing heart-lung transplantation. The rhamnolipids may contribute to the increased mucus output and respiratory morbidity associated with P. aeruginosa colonization.
Streptococcus pneumoniae infections are common, but how they cause host tissue injury and death is incompletely understood. Immunization with pneumolysin, a thiol-activated toxin produced by the pneumococcus, partially protects animals during subsequent infection. The mechanism by which pneumolysin contributes to disease is not known. The aim of the present investigation was to determine the histologic changes induced by recombinant pneumolysin in the rat lung and to compare them with the changes induced by live organisms.Injection of either toxin (200 or 800 ng) or bacteria into the apical lobe bronchus was associated with the development of a severe lobar pneumonia restricted to the apical lobe. The changes induced by the toxin were greater at the higher concentration, and changes were most severe in those animals in which there was partial ligation of the apical lobe bronchus. The pneumonitis was less severe following injection of a modified toxin with decreased hemolytic activity, generated by site-directed mutagenesis of the cloned pneumolysin gene, indicating that this property of the toxin was important in generating pulmonary inflammation. There was still considerable pneumonitis after injection of a modified toxin with decreased capacity to activate complement.
Chronic bronchial inflammation is associated with migration of large numbers of granulocytes into the bronchial tree. A study was designed to find out whether products of bacteria commonly isolated in chronic bronchial infection stimulate neutrophil migration in vitro. Neutrophils from healthy donors were studied by a modified Boyden chamber technique. Bacterial culture filtrates stimulated neutrophil migration over a wide dilution range and the chemotactic activity was heat stable. Culture filtrates derived from Pseudomonas aeruginosa, Streptococcus pneumoniae, and Haemophilus influenzae were significantly chemokinetic and directionally chemotactic, whereas filtrates from Staphylococcus aureus were only chemotactic. Gel filtration of S aureus and P aeruginosa culture filtrates yielded high, medium, and low molecular weight fractions showing chemotactic activity. S pneumoniae and H influenzae yielded fractions with only low molecular weight chemotactic activity. Neutrophil chemotactic responses, occurring in response to all bacterial species tested, appear to represent a defence mechanism by the host. Chemoattractant activity may, however, contribute to bronchial damage mediated by products released from continuously attracted, activated neutrophils.
A total of 11 of 15 Streptococcus pneumoniae culture filtrates and all five bacterial autolysates produced by cell death in the stationary phase caused slowed ciliary beating and disruption of the surface integrity of human respiratory epithelium in organ culture. This effect was inhibited by cholesterol and was heat labile and reduced by standing at room temperature but was stable at -40 degrees C. The activity was detected at the late stationary phase of culture and was associated with the presence of hemolytic activity. Gel filtration of a concentrated culture filtrate and autolysate both yielded a single fraction of approximately 50 kilodaltons which slowed ciliary beating and were the only fractions with hemolytic activity. Rabbit antiserum to pneumolysin, a sulfhydryl-activated hemolytic cytotoxin released by S. pneumoniae during autolysis, neutralized the effect of the culture filtrate on respiratory epithelium. Both native and recombinant pneumolysin caused ciliary slowing and epithelial disruption. Electron microscopy showed a toxic effect of pneumolysin on epithelial cells: cytoplasmic blebs, mitochondrial swelling, cellular extrusion, and cell death, but no change in ciliary ultrastructure. Recombinant pneumolysin (10 micrograms/ml) caused ciliary slowing in the absence of changes in cell ultrastructure. Release of pneumolysin in the respiratory tract during infection may perturb host defenses, allowing bacterial proliferation and spread.
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