Detection of intact influenza viruses using biotinylated biantennary S-sialosides.

We report the modular synthesis of robust, biotinylated biantennary sialylglycoconjugates and their ability to differentiate between two type A influenza strains. This is the first demonstration of glycoconjugate-based discriminatory capture and detection of two strains of intact influenza virus, in the presence of the innate enzymatic activity of viral neuraminidases. We also demonstrate a "carboassay" using glycoconjugates as capture and reporter elements, which therefore, does not require antibodies. The capture of intact influenza viruses is of potential benefit for clinical diagnostics.

Resolution of LPS-induced airway inflammation and goblet cell hyperplasia is independent of IL-18.

BACKGROUND: The resolution of inflammatory responses in the lung has not been described in detail and the role of specific cytokines influencing the resolution process is largely unknown. METHODS: The present study was designed to describe the resolution of inflammation from 3 h through 90 d following an acute injury by a single intratracheal instillation of F344/N rats with LPS. We documented the inflammatory cell types and cytokines found in the bronchoalveolar lavage fluid (BALF), and epithelial changes in the axial airway and investigated whether IL-18 may play a role in the resolution process by reducing its levels with anti-IL-18 antibodies. RESULTS: Three major stages of inflammation and resolution were observed in the BALF during the resolution. The first stage was characterized by PMNs that increased over 3 h to 1 d and decreased to background levels by d 6-8. The second stage of inflammation was characterized by macrophage influx reaching maximum numbers at d 6 and decreasing to background levels by d 40. A third stage of inflammation was observed for lymphocytes which were elevated over d 3-6. Interestingly, IL-18 and IL-9 levels in the BALF showed a cyclic pattern with peak levels at d 4, 8, and 16 while decreasing to background levels at d 1-2, 6, and 12. Depletion of IL-18 caused decreased PMN numbers at d 2, but no changes in inflammatory cell number or type at later time points. CONCLUSION: These data suggest that IL-18 plays a role in enhancing the LPS-induced neutrophilic inflammation of the lung, but does not affect the resolution of inflammation.

Bcl-2 sustains increased mucous and epithelial cell numbers in metaplastic airway epithelium.

Bcl-2, an inhibitor of apoptosis, is expressed in LPS-induced metaplastic goblet cells of rat airways. The present study investigated expression of Bcl-2 in airway mucous cells of persons with cystic fibrosis and tested in rats and mice whether its expression is responsible for sustaining metaplastic mucous cells. A significantly higher percentage of mucous cells expressed Bcl-2 in humans with cystic fibrosis compared with control subjects with no disease or subjects with other diseases. In LPS-instilled F344/N rats, the percentage of Bcl-2-positive mucous cells was decreased to background levels before the resolution of goblet cell metaplasia. Furthermore, intraperitoneal injection of rats with antisense oligonucleotides significantly reduced Bcl-2 expression and goblet cell metaplasia in nasal and pulmonary airway epithelia in rats. In contrast, sustained expression of Bcl-2 in transgenic mice by a metallothionein promoter caused increased LPS-induced goblet cell metaplasia over 8 days compared with wild-type mice. These studies demonstrate that Bcl-2 expression sustains goblet cell metaplasia in various species, that epithelial cell numbers are directly linked to the regulation of the numbers of goblet cells, and that downregulating Bcl-2 expression reduces goblet cell metaplasia.

DNA synthesis and Bcl-2 expression during development of mucous cell metaplasia in airway epithelium of rats exposed to LPS.

Exposure of pulmonary airways to environmental toxins and allergens may cause proliferation of airway epithelial cells and mucous cell metaplasia (MCM); however, it is unclear to what extent proliferating cells differentiate into mucus-storing cells and contribute to MCM. Our previous studies demonstrated that Bcl-2, an inhibitor of apoptosis with cell cycle regulatory functions, is expressed in metaplastic mucous cells. The purpose of the present study was to investigate the number of metaplastic mucous cells that are derived from proliferating epithelial cells and whether Bcl-2 has a role in cell cycle entry in these cells. Rats were intratracheally instilled with 100 microg of LPS from Pseudomonas aeruginosa in 500 microl of saline, and proliferating airway cells were labeled with bromodeoxyuridine (BrdU) by implanting a subcutaneous osmotic pump 24 h before instillation. The volume of stored mucosubstance and the number of mucous cells were increased 10- and 3-fold, respectively, from 24-48 h after instillation. The number of total epithelial cells per millimeter of basal lamina increased, and the number of serous cells per millimeter of basal lamina decreased during this time. Approximately 50% of Alcian blue-periodic acid Schiff-stained mucous cells were labeled with BrdU at 48 h after instillation, suggesting that one-half of the secretory cells were derived from proliferating cells. Furthermore, 50% of the Bcl-2-positive mucous cells were BrdU negative and therefore derived from nonproliferating, preexisting cells. Our findings demonstrate that preexisting and proliferating cells differentiate into mucous cells and compose LPS-induced metaplasia and that Bcl-2 does not have cell cycle regulatory function in these cells.