Ian Geary

Induction of Neutrophil Apoptosis by the Pseudomonas aeruginosa Exotoxin Pyocyanin: A Potential Mechanism of Persistent Infection

Pseudomonas aeruginosa colonizes and infects human tissues, although the mechanisms by which the organism evades the normal, predominantly neutrophilic, host defenses are unclear. Phenazine products of P. aeruginosa can induce death in Caenorhabditis elegans. We hypothesized that phenazines induce death of human neutrophils, and thus impair neutrophil-mediated bacterial killing. We investigated the effects of two phenazines, pyocyanin and 1-hydroxyphenazine, upon apoptosis of neutrophils in vitro. Pyocyanin induced a concentration- and time-dependent acceleration of neutrophil apoptosis, with 50 μM pyocyanin causing a 10-fold induction of apoptosis at 5 h (p < 0.001), a concentration that has been documented in sputum from patients colonized with P. aeruginosa. 1-hydroxyphenazine was without effect. In contrast to its rapid induction of neutrophil apoptosis, pyocyanin did not induce significant apoptosis of monocyte-derived macrophages or airway epithelial cells at time points up to 24 h. Comparison of wild-type and phenazine-deleted strains of P. aeruginosa showed a highly significant reduction in neutrophil killing by the phenazine-deleted strain. In clinical isolates of P. aeruginosa pyocyanin production was associated with a proapoptotic effect upon neutrophils in culture. Pyocyanin-induced neutrophil apoptosis was not delayed either by treatment with LPS, a powerfully antiapoptotic bacterial product, or in neutrophils from cystic fibrosis patients. Pyocyanin-induced apoptosis was associated with rapid and sustained generation of reactive oxygen intermediates and subsequent reduction of intracellular cAMP. Treatment of neutrophils with either antioxidants or synthetic cAMP analogues significantly abrogated pyocyanin-induced apoptosis. We conclude that pyocyanin-induced neutrophil apoptosis may be a clinically important mechanism of persistence of P. aeruginosa in human tissue.

Interleukin-1β Regulates CXCL8 Release and Influences Disease Outcome in Response to Streptococcus pneumoniae, Defining Intercellular Cooperation between Pulmonary Epithelial Cells and Macrophages

ABSTRACT The success of Streptococcus pneumoniae (the pneumococcus) as a pulmonary pathogen is related to its restriction of innate immune responses by respiratory epithelial cells. The mechanisms used to overcome this restriction are incompletely elucidated. Pulmonary chemokine expression involves complex cellular and molecular networks, involving the pulmonary epithelium, but the specific cellular interactions and the cytokines that control them are incompletely defined. We show that serotype 2 or 4 pneumococci induce only modest levels of CXCL8 expression from epithelial cell lines, even in the absence of a polysaccharide capsule. In contrast, coculture of A549 cells with the macrophage-like THP-1 cell line, differentiated with vitamin D, or monocyte-derived macrophages enhanced CXCL8 release. Supernatants from the THP-1 cell line prime A549 cells to release CXCL8 at levels similar to cocultures. Interleukin-1Ra (IL-1Ra) inhibits CXCL8 release from cocultures and reduces the activity of macrophage-conditioned media, but inhibition of tumor necrosis factor alpha (TNF-α) had only a minimal effect on CXCL8 release. Release of IL-1β but not TNF-α was upregulated in cocultures. IL-1 type 1 receptor knockout C57BL/6 and BALB/c mice confirmed the importance of IL-1 signaling in CXC chemokine expression and neutrophil recruitment in vivo. In fulminant disease, increased IL-1 signaling resulted in increased neutrophils in the airway and more invasive disease. These results demonstrate that IL-1 is an important component of the cellular network involving macrophages and epithelial cells, which facilitates CXC chemokine expression and aids neutrophil recruitment during pneumococcal pneumonia. They also highlight a potential clinical role for anti-IL-1 treatment to limit excessive neutrophilic inflammation in the lung.

Role of Activins and Inducible Nitric Oxide in the Pathogenesis of Ectopic Pregnancy in Patients with or without Chlamydia trachomatis Infection

ABSTRACT Chlamydia trachomatis infection can lead to pelvic inflammatory disease, ectopic pregnancy (EP), infertility, and chronic pelvic pain in women. Activins and inducible nitric oxide synthase (iNOS) are produced by the human fallopian tube, and we speculate that tubal activins and iNOS may be involved in the immune response to C. trachomatis in humans and their pathological alteration may result in tubal pathology and the development of EP. Blood and fallopian tubes were collected from 14 women with EP. Sera were analyzed by enzyme-linked immunosorbent assay to detect antibodies against chlamydial heat shock protein 60 (chsp60) and the major outer membrane protein of C. trachomatis. Confirmation of C. trachomatis serology was made using the microimmunofluorescence test. The patients were classified into three groups according to their serological results, and immunohistochemistry and quantitative reverse transcription-PCR were performed to investigate the expression of candidate molecules by tubal epithelial cells among the three groups. This is the first study to show an increase in the expression of activin βA subunit, type II receptors, follistatin, and iNOS within the human fallopian tube of EP patients who were serologically positive for C. trachomatis. A similar expression profile was observed in the fallopian tubes with detectable antibodies only against chsp60. These results were shown at the mRNA and protein levels. We suggest that tubal activin A, its type II receptors, follistatin, and NO could be involved in the microbial-mediated immune response within the fallopian tube, and their pathological expression may lead to tubal damage and the development of EP.

Growth of Aeromonas spp. at 4°C and related toxin production

Over the last few years Aeromonas spp. have been considered to be potential intestinal pathogens (Altwegg & Geiss 1989). Recently, detailed evidence has linked a strain from contaminated shrimp with a clinical isolate from a patient with diarrhoea and found them to be the same (Altwegg et al. 1991). It is now known that Aeromonas spp. are present in many foods, especially those of animal origin (Palumbo et al. 1989). Furthermore, previous studies have shown that these organisms are quite capable of growing at refrigeration temperatures. For example, Palumbo et al. (1985) found that seven out of 14 strains showed significant growth within 14 d at 5°C. However, most studies have only examined a relatively small number of strains, which were not speciated and were simply called A. hydrophila.

Effect of Storage Temperature on Survival of Chlamydia trachomatis after Lyophilization

ABSTRACT Lyophilized preparations of Chlamydia trachomatis were made to investigate how well they would survive storage at four relevant incubation temperatures for 1 week and 1 month. Good viability was maintained by storage at either 4°C or 20°C for 1 week. If the ambient temperature is not too high, short-term transportation of C. trachomatis is achievable through lyophilization.