alginate facilitates infection in mice cystic fibrosis transmembrane conductance regulator knockout

alginate facilitates infection in mice cystic fibrosis transmembrane conductance regulator knockout mice were contaminated with strain BC7 suspended in either phosphate-buffered saline (BC7/PBS) or alginate (BC7/alginate) and the pulmonary bacterial weight and inflammation were monitored. specific to alginate because enzymatically degraded alginate TWS119 or other polyuronic acids did not facilitate bacterial persistence. These observations suggest that alginate may facilitate contamination by interfering with host innate defense mechanisms. Respiratory failure due to lung contamination is the major cause of mortality in cystic fibrosis (CF) patients. CF airways are colonized by more than one opportunistic bacterial pathogen and is a major pathogen. The other opportunistic bacterial pathogens that are frequently isolated from CF airways include complex (BCC) (7). Most individuals with CF experience a characteristic age-related pattern of pulmonary colonization and intermittent exacerbations including and (4 5 Similarly accumulating evidence suggests that can promote colonization by less commonly observed bacteria such as (43). has also been implicated in promoting BCC pathogenesis by increasing the adherence of BCC to respiratory epithelial cells and upregulating the expression of BCC virulence factors (17 31 32 Chronic infections are often associated with a mucoid phenotype due to the production of large quantities of the acidic exopolysaccharide alginate (5). Alginate is an important extracellular virulence factor and has been shown to impair host innate defenses related to phagocytes (1 13 15 18 26 30 In CF airways is found in the airway lumen and hence one may expect large amounts of alginate in airways along with host products. Sputum samples from CF patients have been shown to contain 50 to 200 μg/ml alginate (23 30 In fact it is likely that there are much higher concentrations of alginate in CF airways as sputum samples are mixed with host secretions and hence the concentration of alginate may be DCN underestimated. Since BCC contamination generally occurs in patients who have been chronically colonized with mucoid strain BC7 suspended in either phosphate-buffered saline (PBS) (BC7/PBS) alginate (BC7/alginate) or enzymatically TWS119 degraded alginate (BC7/ED-alginate) and examined the persistence of bacteria and the associated lung inflammation. We also examined the effects of alginate on phagocytosis of by macrophages and neutrophils and the proinflammatory responses of airway epithelial cells to contamination. MATERIALS AND METHODS Bacteria and growth conditions. strains BC7 and BC45 were isolated from your TWS119 sputum of CF sufferers and also have been defined previously (36 37 and had been kindly supplied by J. LiPuma (School of Michigan Ann Arbor MI). Nontypeable was supplied by T. Murphy (School of Buffalo Buffalo NY) and continues to be defined previously (38). ATCC 25416 was bought from American Type TWS119 Tissues Lifestyle (Manassas VA). A medical strain of mucoid alginate in the concentration required. The actual concentration of bacteria inside a suspension was determined by plating. For measurement of phagocytosis bacteria were labeled with fluorescein isothiocyanate (FITC) (Pierce Biotechnology Rockford IL) as explained previously (36). Isolation and purification of alginate. alginate was isolated from a single clinical strain of mucoid amoebocyte lysate assay kit (Lonza) and was found to be <0.1 endotoxin unit/mg of alginate. The alginate preparation was treated with alginate lyase for 6 h at 37°C (Sigma Aldrich Inc. St. Louis MO) to enzymatically degrade the alginate polymer into small oligomers to obtain ED-alginate which was used like a control for alginate. Illness of animals. Ten- to 12-week-old strain Cftrtm1/Unc-TgN(FABPCFTR)iJaw/J homozygous cystic fibrosis transmembrane conductance regulator (CFTR) TWS119 knockout mice were purchased from your Case Western Reserve University or college Animal Resource Center (Cleveland OH). C57BL/6 mice were purchased from Charles River. All mice were maintained inside a specific-pathogen-free barrier facility in microisolator cages throughout the experiment. All methods performed with the mice were approved by the Animal Care and Use Committee of the University or college of Michigan. Mice were anesthetized with ketamine/xylazene (41) and sham infected with 50 μl of PBS or PBS comprising 180 μg of alginate or ED-alginate or were infected with BC7 (1 × 106 to 5 × 106 CFU) suspended in either PBS (BC7/PBS) alginate (BC7/alginate) or ED-alginate TWS119 (BC7/ED-alginate) from the intratracheal route. In some experiments mice were infected with BC7 suspended in low-melting-point agarose polygalacturonic acid or seaweed alginate (Sigma Aldrich Inc. St. Louis MO) at.

Leave a Reply

Your email address will not be published. Required fields are marked *

Proudly powered by WordPress
Theme: Esquire by Matthew Buchanan.